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 OCULAR
PATHOLOGY
 OCULAR
PATHOLOGY
EIGHTH EDITION

MYRON YANOFF MD
Chair Emeritus, Ophthalmology
Professor of Ophthalmology & Pathology
Departments of Ophthalmology & Pathology
College of Medicine
Drexel University
Philadelphia, PA, USA

JOSEPH W. SASSANI MD MHA

Professor of Ophthalmology and Pathology
Pennsylvania State University
The Milton S. Hershey Medical Center
Hershey, PA, USA
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© 2020, Elsevier Inc. All rights reserved.

First edition 1975

Second edition 1982
Third edition 1989
Fourth edition 1996
Fifth edition 2002
Sixth edition 2009
Seventh edition 2015

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F O R E WO R D
Myron Yanoff did his residency at the Scheie Eye Institute of the
University of Pennsylvania in Ophthalmology followed by a
residency in the Department of Pathology. He then did a fel-
lowship at the Armed Forces Institute of Pathology (AFIP) in
Washington, DC, under the directorship of Lorenz Zimmerman.
Yanoff ’s colleague, Ben Fine, was also Zimmerman’s student.
Ben Fine was an excellent electron microscopist, and he and
Yanoff authored the book, Ocular Histology: A Text and Atlas.
Dr. Yanoff developed a series of lectures presented at the Annual
Postgraduate Course in Ophthalmology at the Scheie Eye Institute
and the Lancaster Course in Colby College, Maine, as well as
the Biannual Course in Ophthalmic Pathology at the AFIP. These
lectures led to the first edition of Ocular Pathology: A Text and
Atlas by Drs. Yanoff and Fine, which was published in 1975. The
text was presented in outline form, similar to the lecture series,
with ample illustrations in black and white and a few color
plates. This book became the standard ocular pathology text for
residents in ophthalmology and, indeed, I used this textbook
when I was an ophthalmology resident. Dr. Yanoff went on to
be Chair of the Department of Ophthalmology at the University
of Pennsylvania, then Chair at Hahnemann University and, sub-
sequently, Drexel University, where he maintained a compre-
hensive ophthalmology practice. He and Dr. Fine updated their
textbook every several years, with the second edition in 1982,
third edition in 1989, fourth edition in 1996, and fifth edition
in 2002.
By that time, Yanoff’s resident, Joe Sassani, was ready to replace
Ben Fine as the coauthor of this textbook. Dr. Sassani completed
his ophthalmology residency and fellowship in Ophthalmic
Pathology at the University of Pennsylvania, and developed a
practice focused on glaucoma. Dr. Sassani is currently on the
faculty at Penn State University in Hershey, Pennsylvania. Drs.
Yanoff and Sassani completed the sixth edition of Ocular Pathol-
ogy in 2009 and the seventh edition in 2015. The textbook retained
its outline format; however, virtually all of the illustrations are
now in color, and the book is replete with references. The text-
book has kept up with the times, as it has added new information
including immunohistochemistry, molecular biology, and confocal
microscopy over the years.
The story of ocular pathology is one of successive waves of
confluences of technology, clinicopathologic correlations and,
most importantly, people. An important confluence of technolo-
gies occurred in the mid-1800s when Hermann von Helmholtz
in Heidelberg developed the ophthalmoscope and Rudolf Virchow
in Berlin established cellular pathology as the basis of disease.
vi
This enabled correlation of findings in the eye as seen with the
ophthalmoscope with cellular pathology viewed under the micro-
scope. This led to important clinicopathologic correlations in
ocular pathology, including tumors such as retinoblastoma and
melanoma. As time progressed, more and more disease entities
were defined by clinicopathologic correlations. Zimmerman and
his students, Yanoff being one of them, described the pathology
of most ocular diseases during the so-called “golden age of eye
pathology” from the late 1950s through the 1980s. Subsequently,
many of Zimmerman’s students, and in turn, their students,
Sassani being one of them, applied newer technologies to the
descriptions of these ocular diseases. This enabled updates of
their book, Ocular Pathology. Ocular pathology has advanced
with the confluence of technologies now being molecular biology
and digital technology, including imaging technology such as
confocal microscopy. The most important element in advancing
knowledge and teaching of ocular pathology are individuals, in
this case Drs. Yanoff and Sassani. Remarkably, they have updated
their textbook, now in its eighth edition, keeping up with new
discoveries in ocular pathology and clinicopathologic correla-
tions, including using modern methods of investigation, such
as ocular coherence tomography. Some examples of the updates
in the eighth edition include the ocular manifestations of Zika
virus infection, descriptions of the pathology of intravitreal
injections, ocular injuries associated with terrorism, stem cells
in the conjunctiva, the latest genetic information regarding corneal
dystrophies, the genetics of retinal dystrophies, the TNM clas-
sification in the latest edition of the AJCC Cancer Staging Manual,
and others.
Indeed, Drs. Yanoff and Sassani have kept up with the times
in a remarkable fashion. The definition of a classic textbook is
that it endures the test of time and builds upon itself to a point
where it becomes a standard text that remains current. In this
case, the text began as an outgrowth from the long-standing and
storied history of the venerable AFIP, including Yanoff, a disciple
of Zimmerman, and Sassani, a student of Yanoff. Ocular Pathol-
ogy has stood the test of time, remains current, and remains a
standard textbook for the study of ocular pathology, the basis
of ocular disease. I congratulate Drs. Yanoff and Sassani for their
continued efforts in the production of this beautiful textbook,
which is now the classic textbook for ophthalmology residents
and fellows and pathology residents and fellows.
Hans E. Grossniklaus MD
Professor of Ophthalmology and Pathology
Emory University School of Medicine
 F O R E WO R D S T O T H E F I R S T E D I T I O N
During the year of the observance of the 100th anniversary
(1874–1974) of the University of Pennsylvania’s Department of
Ophthalmology, it is exciting to have the publication of a volume
whose coauthors have contributed significantly to the strides in
ocular pathology taken by the Department in the past several
years.
Myron Yanoff, a highly regarded member of our staff, began
a residency in ophthalmology in 1962, upon graduating from
the University’s School of Medicine. The residency continued
for the next five years, during the first two of which he also held
a residency in the Department of Pathology. His keen interest
and ability in ocular pathology were readily apparent, and I
encouraged him to apply for a fellowship at the Armed Forces
Institute of Pathology (AFIP), Washington, DC. From July 1964
through June 1965, he carried out exceptional research at the
AFIP in both ophthalmology and pathology. He returned to our
Department in July 1965, where the caliber both of his clini-
cal and research work was of the highest. When he completed
his residency in June 1967, I invited him to join the staff, and
he has recently attained the rank of full professor. During the
ensuing years, he has contributed substantially to the litera-
ture, particularly in the fields of ophthalmic and experimen-
tal pathology. He is Board certified in ophthalmology and in
pathology.
Ben Fine, noted for his work in electron microscopy at AFIP
and at George Washington University, has shared his expertise
in the field through lectures presented as part of the curriculum
of the annual 16-week Basic Science Course in the Department’s
graduate program.
It can be said that 100 years ago ophthalmology was a specialty
that had been gradually evolving during the preceding 100 years,
dating from the time of the invention of bifocals by Benjamin
Franklin in 1785. Few American physicians of that era, however,
knew how to treat diseases of the eye, but as medical education
became more specialized it was inevitable that ophthalmology
would also become a specialty.
With the invention of the ophthalmoscope in 1851, great
advances were made in the teaching and practice of ophthalmol-
ogy. This contributed greatly, of course, to setting the scene for
the establishment of the University’s Department of Ophthal-
mology. It was on February 3, 1874, that Dr. William F. Norris
was elected First Clinical Professor of Diseases of the Eye. Similar
chairs had been established earlier in only three other institu-
tions. The chair at the University of Pennsylvania later became
known as the William F. Norris and George E. de Schweinitz
Chair of Ophthalmology.
Both Dr. Norris and Dr. de Schweinitz actively engaged in
the study of ocular pathology. Dr. Norris stressed the importance
of the examination of the eye by microscopy and of the correla-
tion of findings from pathology specimens with the clinical signs.
Dr. de Schweinitz was instrumental in having a member of his
staff accepted as ophthalmic pathologist with the Department
of Pathology.
In the years that followed under succeeding chairmen of the
Department, other aspects of ophthalmology were stressed. Then,
in 1947, during the chairmanship of Dr. Francis Heed Adler, Dr.
Larry L. Calkins was appointed to a residency. Dr. Calkins, like
Dr. Yanoff, displayed a keen interest in ocular pathology. Accord-
ingly, he was instrumental in its study being revitalized during
the three years of his residency. Another resident, Dr. William
C. Frayer, who came to the Department in 1949, joined Dr.
Calkins in his interest in ocular pathology. Dr. Frayer received
additional training in the Department of Pathology and then
became the ophthalmic pathologist of the Department.
The importance of ocular pathology was increasingly evident,
but facilities for carrying out the work in the Department of Oph-
thalmology were unfortunately limited. Until 1964, the pathology
laboratory had been confined to a small room in the outpatient
area of the Department. Then we were able to acquire larger
quarters in the Pathology Building of the Philadelphia General
Hospital located next door to the Hospital of the University of
Pennsylvania. Although the building was earmarked for eventual
demolition, the space was fairly adequate for research and also for
conducting weekly ophthalmic pathology teaching conferences.
Despite the physical aspects, we saw to it that Dr. Yanoff and his
team of workers had a well-equipped laboratory.
During the next several years as I saw that my dream for an
eye institute with facilities for patient care, teaching, and research
under one roof was to become a reality, I was delighted to be
able to include prime space on the research floor for the ever
enlarging scope of ocular pathology. In addition to all that Dr.
Yanoff has had to build upon from the past tradition of our
Department of Ophthalmology, I would like to think that the
new facilities at the Institute have in some measure contributed
to the contents of this excellent volume. With grateful apprecia-
tion, therefore, I look upon this book as the authors’ birthday
present to the Department. From these same facilities, as Dr.
Yanoff and Dr. Fine continue to collaborate, I can hope will
come insights and answers for which all of us are ever searching
in the battle against eye disease.
Harold G. Scheie, MD
Chairman, Department of Ophthalmology
University of Pennsylvania
Director, Scheie Eye Institute
From their earliest days in ophthalmology, Myron Yanoff and
Ben Fine impressed me as exceptional students. As they have
matured and progressed up the academic ladder, they have become
equally dedicated and effective teachers. Their anatomical studies
of normal and diseased tissues have always been oriented toward
providing meaningful answers to practical as well as esoteric
clinical questions. Their ability to draw upon their large personal
experience in clinical ophthalmology, ocular pathology, and
laboratory investigation for their lectures at the Armed Forces
Institute of Pathology and at the University of Pennsylvania has
contributed immeasurably to the success of those courses. Now
vii
viii Forewords to the First Edition
they have used the same time-tested approach in assembling
their material for this book. Beginning with their basic lecture
outlines, then expanding these with just enough text to substitute
for what would have been said verbally in lecture, adding a
remarkable amount of illustrative material for the amount of
space consumed, and then providing pertinent references to get
the more ambitious student started in the pursuit of a subject,
Drs. Yanoff and Fine have provided us with a sorely needed
teaching aid for both the student and the teacher of ocular pathol-
ogy. It should prove to be especially popular among medical
students and residents in both ophthalmology and ocular pathol-
ogy. With it one gets good orientation from the well-conceived
outlines and fine clinicopathologic correlations from the selection
of appropriate illustrations.
It is with considerable pride and admiration that I’ve watched
the evolution of the authors’ work and its fruition in the form
of this latest book. I am proud that both authors launched their
respective careers with periods of intensive study at the Armed
Forces Institute of Pathology and that ever since, they have
remained loyal, dedicated, and highly ethical colleagues. I admire
their youthful energy, their patient, careful attitude, their friendly
cooperative nature, and their ability to get important things
accomplished. I’m appreciative of this opportunity to express
my gratitude for the work they have been doing. If it is true that
“by his pupils, a teacher will be judged,” I could only wish to
have had several dozen more like Drs. Yanoff and Fine.
Lorenz E. Zimmerman, MD
Chief, Ophthalmic Pathology Division
Armed Forces Institute of Pathology
Washington, DC

This edition of Ocular Pathology has been revised extensively to
reflect the many developments in the field since the publication
of the 7th edition. While maintaining a focus on histopathologic
and immunohistopathologic features upon which most diagnoses
are made, we have expanded coverage of supplemental and cor-
relative techniques such as clinical confocal microscopy and
optical coherence tomography. Moreover, we have placed addi-
tional emphasis on the pathobiology underlying established and
new diagnoses. This emphasis is reflected particularly in expanded
coverage of genetics as it relates to disease entities. For a more
in-depth analysis of the latest developments in genetics please
see: Wiggs JL: Part 1 Genetics, in Yanoff M, Duker JS: Ophthal-
mology (5th Ed). London: Elsevier 2018. There are many online
resources to catalog these conditions, including Online Mendelian
Inheritance in Man (OMIM, http://www.ncbi.nlm.nih.gov/omim),
RetNet (https://sph.uth.edu/Retnet/), and Retina International
(http://www.retina-international.org/).
Virtually every chapter has seen extensive revision including
the addition of salient new material. Chapter 1, on the Basic
Principles of Pathology, incorporates an expanded discussion
of the role of the complement system in ocular homeostasis and
disease. The section on immunobiology includes the concept of
the inflammasome as a component of innate immunity. A section
on autoimmunity and autoinflammation has been added. The
discussion of HIV infection has been expanded including newer
developments relative to its complications. There are entirely
new sections on epigenetics and on modern molecular pathology
diagnostic techniques. All of these changes are found in only
the first chapter!
Chapter 2, Congenital Anomalies, revises multiple topics
including the phakomatoses, chromosomal anomalies, and syn-
dromes such as Noonan syndrome and Walker–Warburg syn-
drome. Relevant genetic alterations are cited throughout the
chapter.
Chapter 3, Nongranulomatous Inflammation: Uveitis, End-
ophthalmitis, Panophthalmitis, and Sequelae, includes new atten-
tion on the ocular manifestations of Zika virus infection.
Chapter 4, Granulomatous Inflammation, updates the discus-
sion of sympathetic uveitis (ophthalmia, ophthalmitis) and
nontraumatic infectious causes, such as tuberculosis.
Chapter 5, Surgical and Nonsurgical Trauma, includes an
expanded discussion of ophthalmic operative and postoperative
surgical complications. A section on intravitreal injections has
been added. A discussion of ocular injuries associated with
modern warfare and terrorism has been added to the section
on nonsurgical trauma. Numerous sections have been expanded
in scope including the information on radiation injuries.
Chapter 6, Skin and Lacrimal Drainage System, has an
expanded discussion of congenital lesions and anomalies. The
new classification system for ichthyosis has been added, as has
information regarding its genetic correlates. The section on aging
has been expanded significantly, as has the discussion of numer-
ous individual entities, such as pseudoxanthoma elasticum,
P R E FA C E
epidermolysis bullosa, and erythema multiforme. There is an
extensive revision of the sections on degenerative diseases, col-
lagen diseases, and other inflammatory skin conditions, such as
the vasculitides. Particular attention has been given to the section
on adnexal tumors.
Chapter 7, Conjunctiva, contains an enhanced discussion of
stem cells. The congenital anomalies section is expanded sig-
nificantly, with new entities added. The degenerations section
has been revised to include the new classification of amyloidosis.
The information regarding multiple types of cystic and neoplastic
lesions has been expanded significantly, with a particular emphasis
on cancerous epithelial lesions.
Chapter 8, Cornea and Sclera, contains an extensive revision
of the section on congenital lesions. The ever-changing classifi-
cation of corneal dystrophies is reflected in further revisions to
that section that also include the latest genetic information
impacting our understanding of these disorders. The section on
nonheredofamilial disorders also has been revised extensively.
Significant new information is found in the section on sclera.
Chapter 9, Uvea, includes updates on aniridia, coloboma, and
choroidal dystrophies such as those involving choriocapillaris
atrophy.
Chapter 10, Lens, reflects particular attention on congenital
cataracts and those associated with syndromes. The section on
pseudoexfoliation has been revised, as have other sections includ-
ing lens-related complications and ectopic lens.
Chapter 11, Neural (Sensory) Retina, reflects updates in
congenital and hereditary retinal disorders, vascular diseases,
and inflammatory disorders. Particular attention has been directed
to retinal degenerations. Multiple modifications have been made
to the section on retinal dystrophies with a particular emphasis
on the genetics of these disorders.
Chapter 12, Vitreous, has seen a revision on the amyloid section
and on familial exudative vitreoretinopathy and other familial
disorders.
Chapter 13, Optic Nerve, reflects updates in the section on
congenital and familial disorders including relevant syndromes.
The section on ischemic optic neuropathies has been revised, as
has been the section on optic nerve tumors.
Chapter 14, Orbit, has an extensively revised discussion of
thyroid orbitopathy and muscular disorders. The section on the
reticuloendothelial system and related disorders, including rela-
tive genetic anomalies, has been revised extensively, and the
section on Lymphomas and related disorders has been signifi-
cantly expanded.
Chapter 15, Diabetes Mellitus, includes a new section on ocular
surface disease secondary to diabetes. Additional new informa-
tion and pertinent diagnostic techniques are discussed relative
to diabetic complications for each anatomic region of the eye.
The information on the pathobiology of ocular diabetic com-
plications is expanded greatly.
Chapter 16, Glaucoma, contains a comprehensively revised
discussion of the anatomic basis for aqueous outflow and the
ix
x Preface
histopathologic correlates in glaucoma. The information on the
genetics of glaucoma is revised extensively as is the discussion
of pathobiology for each of the glaucomas where appropriate.
Particular attention has been paid to the discussion of pseudo-
exfoliation. Much information has been added regarding the
pathobiology of optic nerve damage in glaucoma.
Chapter 17, Ocular Melanocytic Lesions, reflects the TMN
classification as found in the 8th edition of the AJCC Cancer
Staging Manual. Additionally, particular emphasis has been placed
on genetic and chromosomal correlates to prognosis in ocular
melanoma. The pathobiology underlying the correlations also
is discussed.
Chapter 18, Retinoblastoma and Simulating Lesions, is revised
extensively, including the chapter title itself, which drops refer-
ence to “pseudoglioma.” The latest classifications for retinoblas-
toma are presented including the principles on which they are
based. Genetic mutations and chromosomal abnormalities relative
to retinoblastoma have been revised. Prognostic factors for
retinoblastoma and the latest information on overall survival
are included. The section on simulating lesions includes new
entities and the latest terminology. Particular attention has been
paid to the latest developments in retinopathy of prematurity.
Adjunctive diagnostic techniques are discussed.
The 8th edition of Ocular Pathology is replete with new infor-
mation reflecting the rapidly evolving world of ophthalmic
pathology. Nevertheless, as we state in the very first line of our
textbook, “The most important tool that the pathologist has at
his/her disposal is meaningful communication with the patient’s
clinician regarding the suspected diagnosis so that the patholo-
gist can choose the appropriate strategy for processing whatever
tissue or other samples are received.” No matter how sophisticated
our techniques become, accurate communication remains the
bedrock for accurate pathologic diagnoses in support of the best
care for our patients.
MY, JS
 AC K N OW L E D G M E N T S

This book could not have been completed without the understanding and patience of our wives
Karin L. Yanoff, PhD, and Gloria Sassani, MA. We also wish to acknowledge the help of our
assistants, Kelly McAnally and Sherri Maslasics. Finally, the members of the Elsevier production
and editorial team lead by Russell Gabbedy, Kayla Wolfe and Sharon Nash, and including project
manager Joanna Souch, designer Brian Salisbury and illustration managers Paula Catalano and
Teresa McBryan all have provided invaluable help and guidance in the production of this 8th
edition of Ocular Pathology.

xi
We dedicate this book to our wives, Karin and Gloria, and to our children.

Basic Principles of Pathology
The most important tool that the pathologist has at his/her dis-
posal is meaningful communication with the patient’s clinician
regarding the suspected diagnosis so that the pathologist can
choose the appropriate strategy for processing whatever tissue
or other samples are received. As will be seen in the discussion
under Modern Molecular Pathology Diagnostic Techniques, there
is a dizzying array of techniques at the pathologist’s disposal;
however, it is only through communication with the clinician
that the pathologist can determine which of these techniques to
utilize to best serve the patient.
INFLAMMATION
Definition
I. Inflammation is the response of a tissue or tissues to a
noxious stimulus.
A. The tissue may be predominantly cellular (e.g., retina),
composed mainly of extracellular materials (e.g., cornea),
or a mixture of both (e.g., uvea).
B. The response may be localized or generalized, and the
noxious stimulus may be infectious or noninfectious.
II. In a general way, inflammation is a response to a foreign
stimulus that may involve specific (immunologic) or non-
specific reactions. Immune reactions arise in response to
specific antigens, but they may involve other components
(e.g., antibodies, T cells) or nonspecific components (e.g.,
natural killer [NK] cells, lymphokines).
III. There is an interplay between components of the inflam-
matory process and blood clotting factors that shapes the
inflammatory process.
Causes
I. Noninfectious causes
A. Exogenous causes: originate outside the eye and body,
and include local ocular physical injury (e.g., perforating
trauma), chemical injuries (e.g., alkali), or allergic reac-
tions to external antigens (e.g., conjunctivitis secondary
to pollen).
B. Endogenous causes: sources originating in the eye and
body, such as inflammation secondary to cellular immu-
nity (phacoanaphylactic endophthalmitis [phacoantigenic
uveitis]); spread from continuous structures (e.g., the
sinuses); hematogenous spread (e.g., foreign particles);
and conditions of unknown cause (e.g., sarcoidosis).
1
II. Infectious causes include viral, rickettsial, bacterial, fungal,
and parasitic agents.
Phases of Inflammation
(Table 1.1 lists the actions of the principal mediators of
inflammation.)
I. Acute (immediate or shock) phase (Fig. 1.1)
A. Five cardinal signs: (1) redness (rubor) and (2) heat
(calor)—both caused by increased rate and volume of
blood flow; (3) mass (tumor)—caused by exudation of
fluid (edema) and cells; (4) pain (dolor) and (5) loss
of function (functio laesa)—both caused by outpour-
ing of fluid and irritating chemicals. Table 1.2 lists the
roles of various mediators in the different inflammatory
reactions.
B. The acute phase is related to histamine release from
mast cells and factors released from plasma (kinin,
complement, and clotting systems).
1. Histamine is found in the granules of mast cells, where
it is bound to a heparin–protein complex. Serotonin
(5-hydroxytryptamine), found in platelets and some
neuroendocrine cells, has a similar effect to histamine.
2. The kinins are peptides formed by the enzymatic
action of kallikrein on the α2-globulin kininogen.
Kallikrein is activated by factor XIIa, which is the
active form of the coagulation factor XII (Hageman
factor). Factor XIIa converts plasma prekallikrein
into kallikrein. Plasmin also can activate Hageman
factor.
3. Plasmin, the proteolytic enzyme responsible for fibri-
nolysis, has the capacity to liberate kinins from their
precursors and to activate kallikrein, which brings
about the formation of plasmin from plasminogen.
Plasmin cleaves C3 complement protein, resulting
in the formation of C3 fragments. It also breaks down
fibrin to form fibrin split products.
4. The complement system (see Table 1.3, which lists the
complement molecules found in the normal eye, and
Table 1.4, which lists the complement molecules found
in diseased eyes) consists of almost 60 proteins present
in blood plasma, on the cell surfaces, or within the
cell. Its vital nature is evidenced by the fact that it
has been preserved by evolution for more than a
billion years.
1
2 CHAPTER 1 Basic Principles of Pathology

TABLE 1.1 The Actions of the Principal Mediators of Inflammation

Mediator Principal Sources Actions
Cell-Derived
Histamine Mast cells, basophils, platelets Vasodilation, increased vascular permeability, endothelial activation
Serotonin Platelets Vasodilation, increased vascular permeability
Prostaglandins Mast cells, leukocytes Vasodilation, pain, fever
Leukotrienes Mast cells, leukocytes Increased vascular permeability, chemotaxis, leukocyte adhesion and
activation
Platelet-activating factor Leukocytes, mast cells Vasodilation, increased vascular permeability, leukocyte adhesion,
chemotaxis, degranulation, oxidative burst
Reactive oxygen species Leukocytes Killing of microbes, tissue damage
Nitric oxide Endothelium, macrophages Vascular smooth muscle relaxation, killing of microbes
Cytokines (TNF, IL-1) Macrophages, endothelial cells, Local endothelial activation (expression of adhesion molecules), fever/
mast cells pain/anorexia/hypotension, decreased vascular resistance (shock)
Chemokines Leukocytes, activated macrophages Chemotaxis, leukocyte activation

Plasma Protein-Derived
Complement products (C5a, C3a, C4a)
Kinins
Proteases activated during coagulation
Plasma (produced in liver)
Plasma (produced in liver)
Plasma (produced in liver)
Leukocyte chemotaxis and activation, vasodilation (mast cell
stimulation)
Increased vascular permeability, smooth muscle contraction,
vasodilation, pain
Endothelial activation, leukocyte recruitment

IL-1, interleukin-1; MAC, membrane attack complex; TNF, tumor necrosis factor.
(Reproduced from Table 2.4, Kumar R, Abbas A, DeLancey A et al.: Robbins and Cotran Pathologic Basis of Disease, 8th edn. Philadelphia,
Saunders. © 2010 by Saunders, an imprint of Elsevier Inc.)

A B

C D

Fig. 1.1 Acute inflammation. A, Corneal ulcer with hypopyon (purulent exudate). Conjunctiva hyperemic.
B, Polymorphonuclear leukocytes (PMNs) adhere to corneal endothelium and are present in the anterior
chamber as a hypopyon (purulent exudate). C, Leukocytes adhere to limbal, dilated, blood-vessel wall (mar-
gination) and have emigrated through endothelial cell junctions into edematous surrounding tissue. D, PMNs
in corneal stroma do not show characteristic morphology but are recognized by “bits and pieces” of nuclei
lining up in a row. (C and D are thin sections from rabbit corneas six hours post-corneal abrasion.)
 Inflammation 3

TABLE 1.2 Role of Mediators in Different TABLE 1.3 Complement Molecules Found
Reactions of Inflammation in the Normal Eye
Role in Inflammation Mediators Complement Molecules
Vasodilation Prostaglandins Expressed in the
Nitric oxide Healthy Eye Eye-Associated Remarks
Histamine Complement System Activators
Increased vascular permeability Histamine and serotonin Amyloid precursor proteins (APP) Retina
C3a and C5a (by liberating vasoactive C-reactive protein (CRP) Retina
amines from mast cells, other cells)
Bradykinin Complement Proteins
Leukotrienes C4, D4, E4 C1q, C2, C3 Cornea, choroid, inner retina, sclera,
PAF optic nerve, retinal pigmented
Substance P epithelium (RPE) cell
Chemotaxis, leukocyte TNF, IL-1 C4 Sclera
recruitment and activation Chemokines C5–8 Cornea, scleral tissue
C3a, C5a C9 Soft drusen from non-AMD eyes,
Leukotriene B4 retina, optic nerve
(Bacterial products; e.g., N-formyl C5b–9 Bruch’s membrane, increase with age
methyl peptides) in non-AMD eyes
Fever IL-1, TNF Factor B Cornea, sclera
Prostaglandins
Pain Prostaglandins Complement Regulators
Bradykinin Factor H Cornea, sclera, iris, ciliary body, retina,
Tissue damage Lysosomal enzymes of leukocytes choroidal tissue outside Bruch’s
Reactive oxygen species membrane, optic nerve
Nitric oxide Factor H-like protein 1 (FHL-1) Bruch’s membrane
C1 inhibitor (C1-INH) Cornea
IL-1, interleukin-1; PAF, platelet-activating factor; TNF, tumor necrosis CD46 (MCP) Cornea and corneal limbus, vitreous
factor. humor, RPE basolateral surface,
(Reproduced from Table 2.7, Kumar R, Abbas A, DeLancey A et al.: photoreceptors
Robbins and Cotran Pathologic Basis of Disease, 8th edn. CD55 (DAF) Cornea and corneal limbus, conjunctiva,
Philadelphia, Saunders. © 2010 by Saunders, an imprint of Elsevier
iris, ciliary body, vitreous humor,
Inc.)
retinal nerve fiber layer (NFL) and
photoreceptors
a. Initially named because it was seen to “comple- CD59 (protectin) Cornea and corneal limbus, conjunctiva,
ment” antibody and cell-mediated immune iris, ciliary body, choroid, vitreous
defenses against microbes. humor, vessels in the inner retina
b. Classic functions: Fig. 1.2 highlights some of the Vitronectin Soft drusen from non-AMD eyes
myriad functions performed by complement. Clusterin Soft drusen from non-AMD eyes
1) Removal of immune (antigen–antibody) Complement Receptors
complexes. Complement receptor-1 (CR1) RPE apical surface
2) Labeling (opsonization) of foreign antigens C3aR Retinal ganglion cells, NFL
for enhanced removal by phagocytes. C5aR Inner plexiform layer (IPL), Müller cells,
3) Recruitment and activation of nearby NFL
leukocytes.
AMD, age-related macular degeneration; RPE, retinal pigment
4) Direct cytolysis of invading microorganisms. epithelium.
c. Performs multiple functions in addition to those (From Mohlin et al.: The link between morphology and complement
“classically” ascribed to it. in ocular disease. Mol Immunol 89:84–99, 2017. Table 1. Elsevier.)
d. Complement achieves its effect through a cascade
of the separate components working in coordina-
tion and in specific sequences leading through 2) Cleavage of C3 produces the active fragments
activation of C3. (Fig. 1.3 is a schematic repre- C3a and C3b.
sentation of the three primary routes or pathways a) C3a is anaphylatoxin leading to chemotactic
of complement cascade activation through C3.) and proinflammatory responses.
1) The three pathways leading to activation of b) C5a also is an anaphylatoxin.
C3 are: c) C3b results in opsonization of foreign
a) Classical pathway. surfaces.
b) Lectin pathway. 3) Thus, C3 has a major role in complement acti-
c) Alternative pathway. vation and generation of immune responses.
4 CHAPTER 1 Basic Principles of Pathology

TABLE 1.4 Complement Molecules Found in the Human Diseased Eye, i.e., in Age-Related
Macular Degeneration (AMD), Glaucoma, Neuromyolitis Optica (NMO) and in Uveitis
Complement Molecules Complement Molecules
Expressed in the Eye Disease–Associated Expressed in the Eye Disease–Associated
Diseased Eye Remarks Diseased Eye Remarks
Complement System Age-Related Macular Complement System Activators
Activators Degeneration (AMD) Immunoglobulin Retina, optic nerve
Amyloid precursor proteins (APP) Drusen
C-reactive protein (CRP) Drusen, choroid Complement Proteins/Activation Products
Immunoglobulin Drusen C1q Retina, ganglion cells (GCL) and
Lipoprotein Drusen nerve fiber layer (NFL)
C3, C3b Retina, GCL and NFL
Complement Proteins/Activation Products C5b-9 (MAC) Retina, GCL
C1q Drusen
Mannose binding protein (MBL) Drusen Complement Regulators
C2a Factor H GCL
C3a, C3c, C3d, C3dg, C3b, iC3b, Bb Choroid, drusen, retinal pigmented Uveitis
epithelial (RPE) cell
C5b–9 (MAC) and sC5b−9a Drusen, RPE, choroid, macula Complement System Activators
Factor Ba Drusen, choroid Immunoglobulin Ocular proteins
Factor Da Drusen, retina
Complement Proteins/Activation Products
Complement Regulators C3c, C3d Aqueous humor
Factor Ia Drusen, inner retina C4a Aqueous humor
Factor Ha Drusen, retinal pigmented epithelial Factor B and Bb Aqueous humor
(RPE) cell, choroid, macula
Complement Anaphylatoxins
FHL-1 Drusen, choroid
C3a, C5a Aqueous humor
Complement receptor 1 (CR1, CD35) Drusen, RPE
Neuromyelitis optica (NMO)
CD46 (MCP) Drusen, choroidal vessels,
basolateral RPE Complement System Activators
Vitronectin Drusen, RPE Immunoglobulin Optic nerve
Clusterin Drusen

Complement Anaphylatoxins
C3a Aqueous humor, drusen
C5a Drusen
Glaucoma
a
Complement-associated genes connected with AMD: (Adamus et al., 2017; Edwards et al., 2005; Hageman et al., 2005; Haines et al., 2005;
Heckner et al., 2010; Klein et al., 2005; Gold et al., 2006; Maller et al., 2007; Park et al., 2009) and uveitis: (Thompson et al., 2013; Yang et al.,
2011, 2013; Xu et al., 2015).
(From Mohlin et al., The link between morphology and complement in ocular disease. Mol Immunol 89:84–99, 2017. Table 2. Elsevier.)

e. C1 has been called the “defining component” of
the classical complement pathway.
1) Functions as a molecular scaffold for binding
of other complement components.
2) Activates and cleaves complement components
to continue the complement cascade.
3) Helps to trigger Wnt receptor signaling.
4) Participates in the process of apoptosis.
5) Cleaves MHC class I molecule and other
proteins.
6) Can adapt to multiple molecular and cellular
processes besides the complement system.
f. Complement plays major roles in immune defense
against microorganisms and in clearing damaged
host components.
1) It responds to recognition of pathogen-
associated molecular patterns (PAMPs) when
they bind to host pattern-recognition receptors
(PRRs) and/or internally produced danger-
associated molecular patterns (DAMPs).
g. Activation of complement pathways results in a
proinflammatory response that includes the gen-
eration of membrane attack complexes (MACs),
which mediate cell lysis, the release of chemokines
to attract inflammatory cells to the site of damage,
and the enhancement of capillary permeability.
(See Fig. 1.3 for the steps leading to activation of
MAC.)
1) Composed of five terminal complement pro-
teins: C5b, C6, C7, C8, and C9. Multiple C9
molecules may be involved.
2) There are numerous levels regulating the
activity of MAC and protecting heathy cells
from attack. In fact, control of the system
is the responsibility of almost half of its
components.
 Inflammation 5

m. Helps maintain tissue homeostasis and cellular

integrity, and functions in tissue regeneration.
Also functions in early sperm–egg interactions
in fertilization, regulation of epiboly and organo-
Increased
vascular genesis, and in refinement of cerebral synapses.
Lysis of permeability Smooth n. The complement system is implicated in mul-
foreign muscle tiple ocular diseases including age-related macular
cells contraction
1 degeneration, glaucoma, and neuromyelitis optica
8 2 (Table 1.4 lists elements of the complement system
Lysis of Mast cell and how they may be involved in these disorders).
7 Complement 3
bacteria degranulation
6 4
o. Complement system, components and their genetic
5 deficiency.
Neutrophil Localization
activation and of complexes
1) Deficiency of early components of the classical
chemotaxis in germinal pathway (C1q, C1r/s, C2, C4, and C3) is asso-
Opsonization centers
and phagocytosis
ciated with autoimmune diseases resulting
of bacteria from failure of clearance of immune complexes
and apoptotic materials and impairment of
humoral response.
2) Deficiencies of mannan-binding lectin and
Fig. 1.2 Summary of the actions of complement and its role in the the early components of the alternative (factor
acute inflammatory reaction. Note how the elements of the reaction D and properdin) and terminal pathways (from
are induced. Increased vascular permeability (1) due to the action of C3 onward components C5, C6, C7, C8, and
C3a and C5a on smooth muscle (2) and mast cells (3) allows exudation C9) increase susceptibility to infections and
of plasma protein. C3 facilitates both the localization of complexes in
to their recurrence.
germinal centers (4) and the opsonization and phagocytosis of bacteria
(5). Neutrophils, which are attracted to the area of inflammation by 3) See also the discussion of monogenic autoin-
chemotaxis (6), phagocytose the opsonized microorganisms. The mem- flammatory syndromes later in this chapter.
brane attack complex, C5–C9, is responsible for the lysis of bacteria (7) p. Activation of complement in the tumor micro-
and other cells recognized as foreign (8). (Adapted with permission from environment enhances tumor growth and increases
Roitt IM, Brostoff J, Male DK: Immunology, 2nd edn. London, Gower
metastasis.
Medical. © Elsevier 1989.)
5. Prostaglandins (prostanoids), which have both inflam-
matory and anti-inflammatory effects, are 20-carbon,
cyclical, unsaturated fatty acids with a 5-carbon ring
a) Disorders resulting from impaired regulation of and two aliphatic side chains.
complement are termed complementopathies. a. They are produced by mast cells, macrophages,
h. Complement proteins opsonize or lyse cells. There- endothelial cells, and others.
fore, they may injure healthy tissue, particularly b. With leukotrienes, they are designated eicosanoids.
when there is a defect in complement regulation. Leukotrienes are metabolized through the lipoxy-
i. Complement is important in such diseases as genase pathway and prostaglandins through the
macular degeneration, rheumatoid arthritis, mul- cyclooxygenase pathway.
tiple sclerosis, Alzheimer’s disease, schizophrenia, c. Active in vascular and systemic reactions of inflam-
and angioedema. mation, oxidative stress, and physiologic functions.
j. T cells and other cell types contain multi­ d. Cyclooxygenase helps catalyze the biosynthesis
ple complement components, which have been of prostaglandins from arachidonic acid.
called the “complosome” in analogy to the in- e. Prostaglandins, cytokines, and leukotrienes func-
flammasome, which will be discussed later in tion to dilate lymphatics at a site of injury.
this chapter. (Fig. 1.4 provides an overview of f. Prostaglandins play an important role in nocicep-
the multiple ways in which the cell complosome tion and pain.
and other complement components may im- 6. Major histocompatibility complex (MHC), called the
pact key cell processes when faced with various human leukocyte antigen (HLA) complex in humans,
challenges.) is critical to the immune response.
k. Other immune system cells that may produce or a. HLAs are present on all nucleated cells of the
be involved in complement function are poly- body and platelets.
morphonuclear leukocytes, mast cells, monocytes,
macrophages, dendritic cells, natural killer (NK)
cells, and B cells.
l. Plays a role in adaptive immune response involv- The HLA region is on autosomal chromosome
6. In practice, the blood lymphocytes are the
ing T and B cells, and functions as a bridge
cells tested for HLA.
between innate and adaptive immunity.
6 CHAPTER 1 Basic Principles of Pathology

Fig. 1.3 Schematic of the complement cascade. The three primary routes for activation of complement are:
(1) the lectin pathway (LP), (2) the classical pathway (CP), and (3) the alternative pathway (AP). The LP and
CP are activated when specific triggers are recognized by host pattern-recognition receptors (PRRs). The AP
is constitutively active. Initial activation through the LP or CP generates a shared C3 convertase (C4b•C2a).
In the AP, C3b pairs with factor B (FB) to form the AP proconvertase (C3b•B), which is processed by factor
D (FD) to form the AP C3 convertase (C3b•Bb). Both types of C3 convertases cleave C3 to generate C3a
and C3b. C3a is an anaphylatoxin, a substance that promotes an inflammatory response. C3b that lands on
the surface of a healthy host cell is quickly inactivated; C3b that attaches to the surface of a pathogen or
altered host cell triggers a rapid amplification loop to generate more C3b, resulting in opsonization. C3b also
complexes with the C3 convertases to form the C5 convertases (C4b•C2a•C3b and C3b•Bb•C3b). In the
terminal complement cascade, C5 convertases cleave C5 into C5a (an anaphylatoxin) and C5b. C5b combines
with C6–9 to form the membrane attack complex (MAC), also referred to as the terminal complement
complex (TCC). Regulatory factors act at various stages of the cascade to control complement activation via
their decay accelerating activity and/or cofactor activity. Additional abbreviations: MASPs, mannose-binding
lectin-associated serine proteases; MBL, mannose-binding lectin; PAMPs, pathogen-associated molecular
patterns. (From Baines AC, Brodsky RA: Complementopathies. Blood Rev 31:213–223, 2017. Figure 1.
Elsevier.)

b. The three genetic loci belonging to HLA class I
are designated by the letters HLA-A, HLA-B, and
HLA-C. Class II MHC molecules are encoded at
the locus HLA-D with three subregions HLA-DP,
HLA-DQ, and HLA-DR.
1) Class I MHC molecules display proteins
derived from foreign antigens, which are rec-
ognized by CD8+ T lymphocytes.
2) Class II MHC molecules present antigens that
are contained in intracellular vesicles and
derived from foreign organisms and soluble
proteins.
c. A tentatively identified specificity carries the
additional letter “W” (workshop) and is inserted
between the locus letter and the allele number—
for example, HLA-BW 15.
d. The HLA system is the main human leukocyte
isoantigen system and the major human histo-
compatibility system.
1) HLA-B 27 is positive in a high percentage of
young women who have acute anterior uveitis
and in young men who have ankylosing spon-
dylitis or Reiter’s disease.
2) HLA-B 51 is strongly associated with Behçet’s
disease.
7. Nonspecific soluble mediators of the immune system
include cytokines, such as interleukins, which
are mediators that act between leukocytes, inter-
ferons (IFNs), colony-stimulating factors (CSFs),
tumor necrosis factor (TNF), transforming
growth factor-β, and lymphokines (produced by
lymphocytes).
 Inflammation 7

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Fig. 1.4 Suggestions on the potential impact of complosome-derived and/or pathogen-shunted intracellular
complement on key cell processes during the host/pathogen interaction. Pathogens trigger an array of
responses when interacting with complement during cell infection processes – some of which are beneficial
for the microbe and some of which support host protection. For example, infection of human papillomavirus
(HPV) triggers globular C1q receptor signaling (gC1qR), which leads to mitochondrial dysfunction and apoptosis
(1). Opsonized bacteria trigger mitochondrial antiviral signaling, which increases the expression of AP-1- and
NF-κB-controlled genes and proinflammatory cytokine responses. C3-opsonized viruses, on the other hand,
are targeted for degradation via the proteosome (2). Opsonized Listeria is also targeted in an intracellular
complement-dependent fashion for degradation after cell entry through v-set immunoglobulin domain con-
taining 4 (VSIG4)-driven autophagosome formation (3). Supporting viral and bacterial propagation, gC1R
signaling on mitochondria was also shown to block retinoic acid-inducible gene I (RIG-I) activation in a process
that promoted the replication of vesicular stomatitis virus (4), while opsonized Klebsiella and other species
use vitronectin to gain entry in nonphagocytic cells (5). Although in most of these processes, complement
fragments were “dragged” into the cell by microbes, we propose that there will also be (subsequent) inter-
actions of invading intracellular pathogens with components of the complosome, for example C3 and C5
activation fragments (6). In line with the “scheme” observed for the role of serum-derived complement, we
further predict that in some cases the complosome will mediate clearance of the pathogen while in other
cases, it will be utilized by the pathogen to promote its survival. (From Arbore G et al.: Intracellular comple-
ment – the complosome – in immune regulation. Mol Immunol 89:2–9, 2017. Figure 2. Elsevier.)

a. The TNF ligand family encompasses a large group
of secreted and cell surface proteins (e.g., TNF and
lymphotoxin-α and -β) that may affect the regula-
tion of inflammatory and immune responses.
b. The actions of the TNF ligand family are some-
what of a mixed blessing in that they can protect
against infection, but they can also induce shock
and inflammatory disease.
C. Immediately after an injury, the arterioles briefly
contract (for approximately five minutes) and then
gradually relax and dilate because of the chemical medi-
ators discussed previously and from antidromic axon
reflexes.
After the transient arteriolar constriction terminates,
blood flow increases above the normal rate for a
variable time (up to a few hours) but then diminishes
to below normal (or ceases) even though the vessels
are still dilated. Part of the decrease in flow is caused
by increased viscosity from fluid loss through the
capillary and venular wall. The release of heparin by
mast cells during this period probably helps to prevent
widespread coagulation in the hyperviscous intra-
vascular blood.
D. During the early period after injury, the leukocytes (pre-
dominantly the PMNs) stick to the vessel walls, at first
8 CHAPTER 1 Basic Principles of Pathology
momentarily, but then for a more prolonged time; this
is an active process called margination (see Fig. 1.1C).
1. Ameboid activity then moves the PMNs through
the vessel wall (intercellular passage) and through
the endothelial cell junctions (usually taking 2–12
minutes); this is an active process called emigration.
2. PMNs, small lymphocytes, macrophages, and imma-
ture erythrocytes may also pass actively across endo-
thelium through an intracellular passage in a process
called emperipolesis.
3. Mature erythrocytes escape into the surrounding
tissue, pushed out of the blood vessels through open-
ings between the endothelial cells in a passive process
called diapedesis.
E. Chemotaxis, a positive unidirectional response to a chemi-
cal gradient by inflammatory cells, may be initiated by
lysosomal enzymes released by the complement system,
thrombin, or the kinins.
F. PMNs (neutrophils; Fig. 1.5) are the main inflammatory
cells in the acute phase of inflammation.
All blood cells originate from a small, common pool
of multipotential hematopoietic stem cells. Regulation
of the hematopoiesis requires locally specialized bone
marrow stromal cells and a coordinated activity of a
group of regulatory molecules—growth factors con-
sisting of four distinct regulators known collectively
as CSFs.
1. PMNs are born in the bone marrow and are consid-
ered “the first line of cellular defense.”
2. CSFs (glycoproteins that have a variable content of
carbohydrate and a molecular mass of 18–90 kDa)
control the production, maturation, and function of
A B
Fig. 1.5 Polymorphonuclear leukocyte (PMN). A, Macroscopic appearance of abscess—that
is, a localized collection of pus (purulent exudate)—in vitreous body. B, PMNs are recog-
nized in abscesses by their segmented (usually three parts or trilobed) nucleus. C, Electron
micrograph shows segmented nucleus of typical PMN, and its cytoplasmic spherical and
oval granules (storage granules or primary lysosomes).
PMNs, macrophages, and eosinophils mainly, but
also of megakaryocytes and dendritic cells.
3. PMNs are the most numerous of the circulating leu-
kocytes, making up 50–70% of the total.
4. PMNs function at an alkaline pH and are drawn to
a particular area by chemotaxis (e.g., by neutrophilic
chemotactic factor produced by human endothelial
cells).
5. The PMNs remove noxious material and bacteria by
phagocytosis and lysosomal digestion.
PMNs produce highly reactive metabolites, includ-
ing hydrogen peroxide, which is metabolized to
hypochlorous acid and then to chlorine, chlora-
mines, and hydroxyl radicals—all important in
killing microbes. Lysosomes are saclike cytoplas-
mic structures containing digestive enzymes and
other polypeptides. Lysosomal dysfunction or lack
of function has been associated with numerous
heritable storage diseases: Pompe’s disease (gly-
cogen storage disease type 2) has been traced
to a lack of the enzymes α-1,4-glucosidase in liver
lysosomes (see Chapter 11); Gaucher’s disease
is caused by a deficiency of the lysosomal enzyme
β-glucosidase (see Chapter 11). Metachromatic
leukodystrophy is caused by a deficiency of the
lysosomal enzyme arylsulfatase-A (see Chapter
11). Most of the common acid mucopolysaccha-
ride, lipid, or polysaccharide storage diseases are
caused by a deficiency of a lysosomal enzyme
specific for the disease (see under appropriate
diseases in Chapters 8 and 11). Chédiak–Higashi
syndrome may be considered a general disorder
of organelle formation (see section on congenital
anomalies in Chapter 11) with abnormally large
and fragile leukocyte lysosomes.
C
 Inflammation 9

A B

Fig. 1.6 A, Eosinophils are commonly seen in allergic conditions

such as this case of vernal catarrh. B, Eosinophils are character-
ized by bilobed nucleus and granular, pink cytoplasm. C, Electron
micrograph shows segmentation of nucleus and dense cytoplasmic
crystalloids in many cytoplasmic storage granules. Some granules
appear degraded.

6. PMNs are end cells; they die after a few days and
liberate proteolytic enzymes, which produce tissue
necrosis.
G. Eosinophils and mast cells (basophils) may be involved
in the acute phase of inflammation.
1. Eosinophils (Fig. 1.6) originate in bone marrow,
constitute 1% or 2% of circulating leukocytes, increase
in number in parasitic infestations and allergic reac-
tions, and decrease in number after steroid admin-
istration or stress. They elaborate toxic lysosomal
components (e.g., eosinophil peroxidase) and generate
reactive oxygen metabolites.
2. Mast cells (basophils; Fig. 1.7) elaborate heparin,
serotonin, and histamine, and they are imperative
for the initiation of the acute inflammatory reaction.
H. The acute phase is an exudative phase (i.e., an outpour-
ing of cells and fluid from the circulation) in which the
nature of the exudate often determines and characterizes
an acute inflammatory reaction.
1. Serous exudate is primarily composed of protein (e.g.,
seen clinically in the aqueous “flare” in the anterior
chamber or under the neural retina in a rhegmatog-
enous neural retinal detachment).
2. Fibrinous exudate (Fig. 1.8) has high fibrin content
(e.g., as seen clinically in a “plastic” aqueous).
3. Purulent exudate (see Figs. 1.1 and 1.5) is composed
primarily of PMNs and necrotic products (e.g., as
seen in a hypopyon).
The term “pus” as commonly used is synonymous
with a purulent exudate.

Except for location, mast cells appear identical

to basophils; mast cells are fixed-tissue cells,
whereas basophils constitute approximately 1%
of circulating leukocytes. Basophils are usually
recognized by the presence of a segmented
nucleus, whereas the nucleus of a mast cells is
large and nonsegmented.
4. Sanguineous exudate is composed primarily of
erythrocytes (e.g., as in a hyphema).
II. Subacute (intermediate or reactive countershock and adap-
tive) phase.
A. The subacute phase varies greatly and is concerned
with healing and restoration of normal homeostasis
10 CHAPTER 1 Basic Principles of Pathology

A B

C D

Fig. 1.7 A, Mast cell seen in center as round cell that contains slightly basophilic cytoplasm and round to
oval nucleus. B, Mast cells show metachromasia (purple) with toluidine blue (upper right and left and lower
right) and C, positive (blue) staining for acid mucopolysaccharides with Alcian blue. D, Electron microscopy
of granules in cytoplasm of mast cell often shows typical scroll appearance.

(formation of granulation tissue and healing) or with
the exhaustion of local defenses, resulting in necrosis,
recurrence, or chronicity.
B. PMNs at the site of injury release lysosomal enzymes
into the area.
1. The enzymes directly increase capillary permeability
and cause tissue destruction.
2. Indirectly, they increase inflammation by stimulating
mast cells to release histamine, by activating the kinin-
generating system, and by inducing the chemotaxis
of mononuclear (MN) phagocytes.
C. Mononuclear (MN) cells (Fig. 1.9) include lymphocytes
and circulating monocytes.
1. Monocytes constitute 3%–7% of circulating leuko-
cytes, are bone marrow-derived, and are the progenitor
of a family of cells (monocyte–histiocyte–macrophage
family) that have the same fundamental character-
istics, including cell surface receptors for complement
and the Fc portion of immunoglobulin, intracellular
lysosomes, and specific enzymes; production of mono-
kines; and phagocytic capacity.
2. Circulating monocytes may subsequently become
tissue residents and change into tissue histiocytes,
macrophages, epithelioid histiocytes, and inflamma-
tory giant cells.
3. CSFs (glycoproteins that have a variable content of
carbohydrate and a molecular mass of 18–90 kDa)
control the production, maturation, and function of
MN cells.
4. These cells are the “second line of cellular defense,”
arrive after the PMN, and depend on release of che-
motactic factors by the PMN for their arrival.
a. Once present, MN cells can live for weeks, and
in some cases even months.
b. MN cells cause much less tissue damage than do
PMNs, and they are more efficient phagocytes.
5. Monocytes have an enormous phagocytic capacity
and are usually named for the phagocytosed material
(e.g., blood-filled macrophages [erythrophagocytosis]
and lipid-laden macrophages; Fig. 1.10).
6. Monocytes replace neutrophils as the predominate
cell 24–48 hours after the onset of inflammation.
D. Lysosomal enzymes, including collagenase, are released
by PMNs, MN cells, and other cells (e.g., epithelial cells
and keratocytes in corneal ulcers) and result in consider-
able tissue destruction.
In chronic inflammation, the major degradation of
collagen may be caused by collagenase produced
by lymphokine-activated macrophages.
 Inflammation 11

E. If the area of injury is tiny, PMNs and MN cells alone

can handle and “clean up” the area with resultant healing.
F. In larger injuries, granulation tissue is produced.
1. Granulation tissue (Fig. 1.11) is composed of leuko-
cytes, proliferating blood vessels, and fibroblasts.
2. MN cells arrive after PMNs, followed by an ingrowth
of capillaries that proliferate from the endothelium
of pre-existing blood vessels.

The new blood vessels tend to leak fluid and

A leukocytes, especially PMNs.

3. Fibroblasts (see Fig. 1.11), which arise from fibrocytes

B C
Fig. 1.8 A, Cobweb appearances of fibrinous exudate, stained with
periodic acid–Schiff. Cells use fibrin as scaffold to move and to lay down
reparative materials. B, Electron micrograph shows periodicity of fibrin
cut in longitudinal section. C, Fibrin cut in cross-section.
and possibly from other cells (monocytes), prolifer-
ate, lay down collagen (Table 1.5), and elaborate
ground substance.
4. With time, the blood vessels involute and disappear,
the leukocytes disappear, and the fibroblasts return
to their resting state (fibrocytes). This involutionary
process results in shrinkage of the collagenous scar
and a reorientation of the remaining cells into a par-
allel arrangement along the long axis of the scar.
5. If the noxious agent persists, the condition may not
heal as described previously, but instead may become
chronic.
6. If the noxious agent that caused the inflammation
is immunogenic, a similar agent introduced at a future
date can start the cycle anew (recurrence).

Histiocyte/macrophage Activated macrophage Activated macrophages

? ?
?

Multinucleated
inflammatory
?
giant cell

Langhans Foreign body Touton

Epithelioid cells
A B
Fig. 1.9 A, Monocytes have lobulated, large, vesicular nuclei and moderate amounts of cytoplasm, and they
are larger than the segmented polymorphonuclear leukocytes and the lymphocytes, which have round nuclei
and scant cytoplasm. B, Possible origins of multinucleated inflammatory giant cells and of epithelioid cells.
12 CHAPTER 1 Basic Principles of Pathology
A B
Fig. 1.10 A, Foamy and clear lipid-laden macrophages in subneural retinal space. B, Cytoplasm of macro-
phages stains positively for fat with oil red-O technique.
A B
Fig. 1.11 Granulation tissue. A, Pyogenic granuloma, here in region of healing chalazion, is composed of
granulation tissue. B, Three components of granulation tissue are capillaries, fibroblasts, and leukocytes.
III. Chronic phase
A. The chronic phase results from a breakdown in the pre-
ceding two phases, or it may start initially as a chronic
inflammation (e.g., when the resistance of the body and
the inroads of an infecting agent, such as the organisms
of tuberculosis or syphilis, nearly balance; or in condi-
tions of unknown cause such as sarcoidosis).
B. Chronic nongranulomatous inflammation is a prolifera-
tive inflammation characterized by a cellular infiltrate
of lymphocytes and plasma cells (and sometimes PMNs
or eosinophils).
1. The lymphocyte (Fig. 1.12) constitutes 15%–30% of
circulating leukocytes and represents the competent
immunocyte.
a. All lymphocytes probably have a common stem
cell origin (perhaps in the bone marrow) from
which they populate the lymphoid organs: the
thymus, spleen, and lymph nodes.
b. Two principal types of lymphocytes are recognized:
(1) The bone marrow-dependent (or bursal equiv-
alent) B-lymphocyte is active in humoral immu-
nity, is the source of immunoglobulin production
(Fig. 1.13), and is identified by the presence of
immunoglobulin on its surface; (2) the thymus-
dependent T lymphocyte participates in cellular
immunity, produces a variety of lymphokines,
and is identified by various surface antigens.
1) Helper-inducer T lymphocytes (CD4-positive)
initiate the immune response in conjunction
with macrophages and interact with (helper)
B lymphocytes.
CD4+ T cells are activated after interaction
with antigen–MHC complex and differenti-
ate into Helper subsets. These functionally
distinct T-helper subsets participate in host
defense and immunoregulation. Classically,
T-helper 1 (Th1) and T-helper 2 (Th2) cells
secrete a distinctive suite of cytokines:
Th1 express T-bet and produce interferon-γ
and are involved predominantly in cell-
mediated immunity (e.g., cytotoxic T-cell
response); Th2 express Gata3 and produce
interleukins-4, -5, and -13. Regulatory T
(Treg) cells also are CD4+-derived cells,
 Inflammation 13

TABLE 1.5 Heterogeneity of Collagens in the Cornea*

Type Polypeptides Monomer Polymer
I [α1(I)]2α2(I)

II [α1(II)]3

III [α1(III)]3

IV [α1(IV)]2α2(IV)

V [α1(V)]2α2(V)

VI [α1(VI)]2α2(VI)α3(VI)

VII [α1(VII)]3?

VIII [α1(VIII)]2α2(VIII)?

IX [α1(IX)]2α2(IX)α3(IX)

XII [α1(XII)]3

*At least 10 genetically distinct collagens have been described in the corneas of different animal species, ages, and pathologies. Types I, II, III,
and V collagens are present as fibrils in tissues. Types IV, VI, VII, and VIII form filamentous structures. Types IX and XII are fibril-associated
collagens. The sizes of the structures are not completely known. Type II collagen is found only in embryonic chick collagen associated with the
primary stroma. Type III collagen is found in Descemet’s membrane and in scar tissue. Types I and V form the heterotypic fibrils of lamellar
stroma. Type VII has been identified with the anchoring fibrils, and type VIII is present only in Descemet’s membrane. Type IX collagen,
associated with type II fibrils in the primary stroma, and type XII collagen, associated with type I/V fibrils, are part of a family of fibril-associated
collagens with interrupted triple helices. Both type IX and type XII are covalently associated with a chondroitin sulfate chain.
(Reproduced from Cintron C: The molecular structure of the corneal stroma in health and disease. In Podos SM, Yanoff M, eds: Textbook of
Ophthalmology, vol. 8. London, Mosby. © Elsevier 1994.)

serve an immunosuppressive function, and 2) Suppressor-cytotoxic T lymphocytes (CD8-

express the master transcription factor
FoxP3. There are thymic-derived natural,
nTreg cells and peripherally induced iTreg
cells that relate to autoimmunity. T-helper
17 (Th17) cells participate in protective
tumor immunity; however, Th17-associated
cytokines may be associated with tumor
initiation and growth and also with auto-
immune diseases. Finally, there are fol-
licular T-helper (Tfh) cells that are in
proximity to B cells in the germinal centers
of lymphoid tissue. They promote class
switching of B cells and express the
master regulator Bc16 and the effector
cytokine IL-21 as well as other surface
molecules. Fig. 1.14 illustrates the com-
plexity, flexibility and plasticity of the rela-
tionships between T-helper cells.
positive) suppress the immune response and
are capable of killing target cells (e.g., cancer
cells) through cell-mediated cytotoxicity.
3) MHC molecules present antigenic peptides
to CD8+ T cells, thereby providing the founda-
tion for immune recognition.
2. The plasma cell (Fig. 1.15) is produced by the bone
marrow–derived B lymphocyte, elaborates immuno-
globulins (antibodies), and occurs in certain modified
forms in tissue sections.
After germinal center B cells undergo somatic
mutation and antigen selection, they become
either memory B cells or plasma cells. CD40 ligand
directs the differentiation of germinal center B
cells toward memory B cells rather than toward
plasma cells.
14 CHAPTER 1 Basic Principles of Pathology

rbc

Fig. 1.12 Lymphocyte. A, Low magnification shows cluster of many lymphocytes appearing as a deep blue
infiltrate. Cluster appears blue because cytoplasm is scant and mostly nuclei are seen. B, Electron micrograph
shows lymphocyte nucleus surrounded by small cytoplasmic ring containing several mitochondria, diffusely
arrayed ribonucleoprotein particles, and many surface protrusions or microvilli (rbc, red blood cell). C, Lym-
phocytes seen as small, dark nuclei with relatively little cytoplasm. Compare with polymorphonuclear leuko-
cytes (segmented nuclei) and with larger plasma cells (eccentric nucleus surrounded by halo and basophilic
cytoplasm).

N VL
N
a. Plasmacytoid cell (Fig. 1.16A and B): This has a
CL
single eccentric nucleus and slightly eosinophilic
VH granular cytoplasm (instead of the normal baso-
C
philic cytoplasm of the plasma cell).
Antigen- CH1 C b. Russell body (Fig. 1.16C and D): This is an inclu-
binding CH2 CH3
sites
sion in a plasma cell whose cytoplasm is filled
C
and enlarged with eosinophilic grapelike clusters
C Heavy chain
(morular form), with single eosinophilic globular
structures, or with eosinophilic crystalline struc-
tures; usually the nucleus appears as an eccentric
N
Light chain
rim or has disappeared.
N
Fig. 1.13 The basic immunoglobulin structure. The unit consists of two
identical light polypeptide chains linked together by disulfide bonds (gray). The eosinophilic material in plasmacytoid cells
The amino-terminal end (N) of each chain is characterized by sequence and in Russell bodies appears to be immuno-
variability (VL, VH), whereas the remainder of the molecule has a relatively globulin that has become inspissated, as if
constant structure (CL, CH1–CH3). The antigen-binding sites are located the plasmacytoid cells can no longer release
at the N-terminal end. (Adapted with permission from Roitt IM, Brostoff the material because of defective transport
J, Male DK: Immunology, 2nd edn. London, Gower Medical. © Elsevier by the cells (“constipated” plasmacytoid cells).
1989.)
 Inflammation 15

C. Chronic granulomatous inflammation is a proliferative b. They are often found oriented around necrosis as
inflammation characterized by a cellular infiltrate of large polygonal cells that contain pale nuclei and
lymphocytes and plasma cells (and sometimes PMNs abundant eosinophilic cytoplasm whose borders
or eosinophils). blend imperceptibly with those of their neighbors
1. Epithelioid cells (Fig. 1.17) are bone marrow–derived in a pseudosyncytium (“palisading” histiocytes in
cells in the monocyte–histiocyte–macrophage family a granuloma).
(Fig. 1.18). c. All cells of this family interact with T lymphocytes,
a. In particular, epithelioid cells are tissue monocytes are capable of phagocytosis, and are identified by
that have abundant eosinophilic cytoplasm, some- the presence of surface receptors for complement
what resembling epithelial cells. and the Fc portion of immunoglobulin.
2. Inflammatory giant cells, probably formed by fusion
of macrophages rather than by amitotic division,
Tfh predominate in three forms:
a. Langhans’ giant cell (Fig. 1.19; see Fig. 1.17): This
Plasticity Bcl6 is typically found in tuberculosis, but it is also seen
in many other granulomatous processes. When
Th1 Th2 sectioned through its center, it shows a perfectly
Bcl6
Bcl6 FoxP3 Bcl6
homogeneous, eosinophilic, central cytoplasm
T-bet T-bet Gata3 Gata3 with a peripheral rim of nuclei.
Flexibility
T-bet Gata3 If the central portion is not homogeneous,
RORyt FoxP3
foreign material such as fungi may be present:
RORyt T-bet the cell is then not a Langhans’ giant cell but
FoxP3 FoxP3
a foreign-body giant cell. When a Langhans’
giant cell is sectioned through its periphery,
RORyt FoxP3
it simulates a foreign-body giant cell.
Th17 Tregs

b. Foreign-body giant cell (Fig. 1.20): This has its

Fig. 1.14 Flexibility and plasticity of helper T cells. Recent studies con-
tinue to reveal surprising flexibility in expression of “master regulator”
transcription factors. In addition, there are now many examples in which
helper T cell phenotypes can change their pattern of expression of sig-
nature cytokines and gene expression. Striking examples exist in which
apparently fully committed “lineages” readily switch their phenotype,
and there are now many circumstances in which helper T cells have
been shown to express more than one master regulator. This may be
advantageous in terms of host defense, but it needs to be borne in
mind in thinking about effective therapies for immune-mediated disease
and vaccine development. (From Nakayamada S, Takahashi H, Kanno Y
et al.: Helper T cell diversity and plasticity. Curr Opin Immunol 24:297,
2012.)
nuclei randomly distributed in its eosinophilic
cytoplasm and contains foreign material.
c. Touton giant cell (Fig. 1.21), frequently associated
with lipid disorders such as juvenile xanthogranu-
loma, appears much like a Langhans’ giant cell
with the addition of a rim of foamy (fat-positive)
cytoplasm peripheral to the rim of nuclei.
3. Three patterns of inflammatory reaction may be found
in granulomatous inflammations:
a. Diffuse type (Fig. 1.22A): This typically occurs in
sympathetic uveitis, disseminated histoplasmosis

A B

Fig. 1.15 Plasma cell. A, Plasma cells are identified by eccentrically located nucleus containing clumped
chromatin and perinuclear halo in basophilic cytoplasm that attenuates opposite to nucleus. Plasma cells are
larger than small lymphocytes, which contain deep blue nuclei and scant cytoplasm. B, Electron microscopy
shows exceedingly prominent granular endoplasmic reticulum that accounts for cytoplasmic basophilia and
surrounds nucleus. Mitochondria are also present in cytoplasm.
16 CHAPTER 1 Basic Principles of Pathology

A B

C D

Fig. 1.16 Altered plasma cells. A, Electron micrograph shows that left plasmacytoid cell contains many small
pockets of inspissated material (γ-globulin) in segments of rough endoplasmic reticulum; right cell contains
large globules (γ-globulin), which would appear eosinophilic in light microscopy. B, Plasmacytoid cell in center
has eosinophilic (instead of basophilic) cytoplasm that contains tiny pink globules (γ-globulin). C, Russell body
appears as large anuclear sphere or D, multiple anuclear spheres.

Activated ?
macrophage Langerhans’
cell

Lymphokine
?
Monocyte/ Giant cell Foreign
macrophage body

? ?

Epithelioid Touton
cell
Fig. 1.17 Epithelioid cells in conjunctival, sarcoidal granuloma, here
Fig. 1.18 Proposed scheme for the terminal differentiation of cells of
forming three nodules, which are identified by eosinophilic color resem-
the monocyte/macrophage system. The pathologic changes result from
bling epithelium. Giant cells, simulating Langhans’ giant cells, are seen
the inability of the macrophage to deal effectively with the pathogen.
in nodules.
Lymphokines from active T cells induce monocytes and macrophages
to become activated macrophages. Where prolonged antigenic stimula-
tion exists, activated macrophages may differentiate into epithelioid
cells and then into giant cells in vivo, in granulomatous tissue. The
multinucleated giant cell may be derived from the fusion of several
epithelioid cells. (Adapted with permission from Roitt IM, Brostoff J,
Male DK: Immunology, 2nd edn. London, Gower Medical. © Elsevier
1989.)
 Inflammation 17

and other fungal infections, lepromatous leprosy,
juvenile xanthogranuloma, Vogt–Koyanagi–Harada
syndrome, cytomegalic inclusion disease, and toxo-
plasmosis. The epithelioid cells (sometimes with
macrophages or inflammatory giant cells or both)
Fig. 1.19 Langhans’ giant cells have homogeneous central cytoplasm
surrounded by rim of nuclei.
are distributed randomly against a background
of lymphocytes and plasma cells.
b. Discrete type (sarcoidal or tuberculocidal; see Fig.
1.22B): This typically occurs in sarcoidosis, tuber-
culoid leprosy, and miliary tuberculosis. An accu-
mulation of epithelioid cells (sometimes with
inflammatory giant cells) forms nodules (tuber-
cles) surrounded by a narrow rim of lymphocytes
(and perhaps plasma cells).
c. Zonal type (see Fig. 1.22C): This occurs in caseation
tuberculosis, some fungal infections, rheumatoid
scleritis, chalazion, phacoanaphylactic (phacoanti-
genic) endophthalmitis, toxocara endophthalmitis,
and cysticercosis.
1) A central nidus (e.g., necrosis, lens, and foreign
body) is surrounded by palisaded epithelioid
cells (sometimes with PMNs, inflammatory
giant cells, and macrophages) that in turn are
surrounded by lymphocytes and plasma cells.
2) Granulation tissue often envelops the entire
inflammatory reaction.

A B

Fig. 1.20 A, Foreign-body giant cell (FBGC) simulating Langhans’ giant cells, except that homogeneous
cytoplasm is interrupted by large, circular foreign material. B, Anterior-chamber FBGCs, here surrounding
clear clefts where cholesterol had been, have nuclei randomly distributed in cytoplasm.

A B

Fig. 1.21 A, Touton giant cells in juvenile xanthogranuloma closely resemble Langhans’ giant cells except
for the addition of peripheral rim of foamy (fat-positive) cytoplasm in the former. B, Increased magnification
showing fat positivity of peripheral cytoplasm with oil red-O technique. (Case presented by Dr. M Yanoff to
the Eastern Ophthalmic Pathology Society, 1993, and reported in Arch Ophthalmol 113:915, 1995.)
18 CHAPTER 1 Basic Principles of Pathology
A B
Fig. 1.22 Patterns of granulomatous inflammation. A, Diffuse type
in sympathetic uveitis. B, Discrete (sarcoidal or tuberculocidal) type
in sarcoidosis. C, Zonal type in phacoanaphylactic endophthalmitis.
Staining Patterns of Inflammation
I. Patterns of inflammation are best observed microscopically
under the lowest (scanning) power.
II. With the hematoxylin and eosin (H&E) stain, an infiltrate
of deep blue tint (basophilia) usually represents a chronic
nongranulomatous inflammation. The basophilia is pro-
duced by lymphocytes that have blue nuclei (when stained
with hematoxylin) and practically no cytoplasm (if it were
present, it would stain pink with eosin) and by plasma cells
that have blue nuclei and blue cytoplasm.
III. A deep blue infiltrate with scattered gray (pale pink) areas
(“pepper and salt”) usually represents a chronic granulo-
matous inflammation, with the blue areas lymphocytes and
plasma cells, and the gray areas islands of epithelioid cells.
IV. A “dirty” gray infiltrate usually represents a purulent reac-
tion with PMNs and necrotic material.
A. If the infiltrate is diffuse (Fig. 1.23; e.g., filling the vitre-
ous [vitreous abscess]), the cause is probably bacterial.
B. If the infiltrate is localized into two or more small areas
(Fig. 1.24; i.e., multiple abscesses or microabscesses),
the cause is probably fungal.
IMMUNOBIOLOGY
Background
I. There are three levels of human defense against invading
organisms:
A. Anatomical and physiological barriers
1. Examples are the skin, enzymes in secretions, mucoid
surface secretions, surfactant, and gastric pH.
B. Innate immunity (See also discussion of complement
earlier in this chapter.)
1. This system has few receptors for antigens, but ones
that are widespread among potential invaders.
2. Inflammasome (Fig. 1.25) illustrates how activation
of one of the upstream sensors precipitates inflam-
masome formation leading to cell membrane breach
and cell death through pyroptosis.
a. It is a multiprotein complex composed of a sensor
protein, the adapter protein ASC (apoptosis-
associated speck-like protein containing caspase
recruitment domain), and the inflammatory pro-
tease caspase-1.
b. Downstream substrates are gasdermin D, IL-1β,
and IL-18 and are responsible for an inflamma-
tory form of cell death called pyroptosis with
gasdermin D functioning as the actual instru-
ment of cell death by forming pores in the cell
membrane.
1) Following its activation, caspase-1 induces
activation of IL-1β, and IL-18 thereby resulting
in inflammation.
c. Upstream sensors include NLRP (nucleotide-
binding domain and leucine-rich repeat contain-
ing) 1, NLRP3, NLRC4, AIM2, and pyrin.
 Immunobiology 19

A B C

Fig. 1.23 Staining patterns of inflammation. A, Macroscopic appearance of diffuse vitreous abscess.
B, Diffuse abscess, here filling vitreous, characteristic of bacterial infection. C, Special stain shows Gram
positivity of bacterial colonies in this vitreous abscess.

A B C

Fig. 1.24 Staining patterns of inflammation. A, Macroscopic appearance of multiple vitreous microabscesses,
characteristic of fungal infection. B, One vitreous microabscess contiguous with detached retina. C, Septate
fungal mycelia (presumably Aspergillus) from same case stained with Gomori’s methenamine silver.

1) Activated by stimuli such as infection and
changes in cell homeostasis.
d. Involved in monogenic autoinflammatory disorders
in which there is apparently spontaneous inflam-
mation in the absence of inciting auto-antibodies
or antigen-specific T cells. (See section on auto-
inflammation later in this chapter).
1) Abnormal response to endogenous or exog-
enous factors that results in exaggerated acti-
vation of inflammation and usually mediated
by the inflammasome.
2) May involve the eye in idiopathic granuloma-
tous disorders, familial Mediterranean fever,
tumor necrosis factor receptor-associated peri-
odic syndrome, mevalonate kinase deficiency,
and cryopyrin-associated periodic syndrome.
e. Involved in the pathogenesis of glaucoma, age-
related macular degeneration, diabetic retinopathy,
dry eye, and ocular infections.
f. Activity by certain inflammasomes is associate
with susceptibility to infections, autoimmunity,
and tumorigenesis.
g. When the cell is in a steady state, inflammasome
components are present in the cytosol, but their
assembly is prevented by auto-inhibitory mecha-
nisms mediated by chaperone protein.
h. Autophagy inducers reduce symptoms of
inflammasome-related diseases, while deficien-
cies in autophagy-related proteins may induce
aberrant activation of inflammasome-mediated
tissue damage.
C. Adaptive immunity
1. The main components of this system are B and T
lymphocytes. Their strength is in their ability to gen-
erate a response to a diverse population of potential
pathogens.
The immune system provides the body with a
mechanism to distinguish “self” from “nonself.”
The distinction, made after a complex, elaborate
process, ultimately relies on receptors on the only
immunologically specific cells of the immune
system, the B and T lymphocytes.
20 CHAPTER 1 Basic Principles of Pathology
Fig. 1.25 Inflammasome. (From Place DE, Kanneganti TD: Recent advances in inflammasome biology. Curr
Opin Immunol 50:32–38, 2018. Figure 1. Elsevier.)
II. Table 1.6 lists the major effector elements in our immu-
nologic defense system.
III. All lymphocytes in mammalian lymph nodes and spleen
have a remote origin in the bone marrow. Those that have
undergone an intermediate cycle of proliferation in the
thymus (thymus-dependent, or T lymphocytes) mediate
cellular immunity, whereas those that seed directly into
lymphoid tissue (thymus-independent, or B lymphocytes)
provide the precursors of cells that produce circulating
antibodies.
A. Thus, mediators of immune responses can be either
specifically reactive lymphocytes (cell-mediated immu-
nity) or freely diffusible antibody molecules (humoral
immunity).
B. Antibody-producing B cells or killer T-type cells are
only activated when turned on by a specific antigen.
When an antigen (immunogen) penetrates the body,
it binds to an antibody-like receptor on the surface
of its corresponding lymphocyte that proliferates and
generates a clone of differentiated cells. Some of
the cells (large B lymphocytes and plasma cells)
secrete antibodies; T cells secrete lymphokines; and
other lymphocytes circulate through blood, lymph,
and tissues as an expanded reservoir of antigen-
sensitive (memory) cells. When the immunogen
encounters the memory cells months or years later,
it evokes a more rapid and copious secondary anam-
nestic response. Other immune cells (e.g., NK) are
less specific and eliminate a variety of infected or
cancerous cells.
IV. T lymphocytes derive from lymphoid stem cells in the bone
marrow and mature under the influence of the thymus.
A. T lymphocytes are identified by surface antigens (T3,
T4, T8, and T11).
1. T lymphocytes are divided into two major subsets
that express either CD4 or CD8 protein on their
surface. CD4+ and CD8+ T cells depend on different
signaling pathways to support their development and
survival.
B. T lymphocytes are the predominant lymphocytes in the
peripheral blood and reside in well-defined interfollicular
areas in lymph nodes and spleen.
 Immunobiology 21

TABLE 1.6 Host Effector Mechanisms

Name Properties Effector Mechanisms
Soluble Effectors
Complement system Proteolytic cascade, activated by antibody, directly by Direct destruction of pathogens via pore formation; recruit
microbial components, or via PRRs inflammatory cells; enhance phagocytosis and killing
Coagulation system Proteolytic cascade, activated by tissue and vascular Prevents blood loss; bars access to bloodstream;
damage proinflammatory
Kinin system Proteolytic cascade triggered by tissue damage Proinflammatory; causes pain response; increases vascular
permeability to allow increased access to plasma proteins
Antibodies Antigen-specific proteins produced by B cells; Directly neutralize pathogens; activate complement; opsonize
recognize a broad range of antigens pathogens to enhance phagocytosis and killing

Cellular Effectors
Monocyte/macrophage Have PRRs to recognize pathogens; activated by Phagocytosis and microbial killing via multiple mechanisms;
dendritic cell specific T cells and chemokines antigen presentation
Neutrophil Have PRRs to recognize pathogens, activated antibody Phagocytosis and microbial killing via multiple mechanisms
and complement
Eosinophil Recognize antibody-coated parasites Killing of multicellular pathogens
Basophil/mast cell Associated with IgE-mediated responses Release of granules containing histamine and other mediators
of anaphylaxis
NK cell Lymphocyte lacking antigen-specific reactivity; Induce death of infected cells via membrane pores and
recognize PAMPs of intracellular pathogens, induced apoptosis
activated by chemokines and by membrane proteins
of infected cells
B lymphocyte Recognize antigens presented by APCs; regulated by T Produce antibody
cells and chemokines
T lymphocyte Recognize antigens presented by APCs; regulate major Directly kill infected cells via membrane pores and induced
portions of both adaptive and innate immunity apoptosis; activate macrophages; many other functions

APCs, antigen-presenting cells; PAMPs, pathogen-associated molecular patterns; PRRs, pattern-recognition receptors.
(Reproduced from Table 3.1, Coleman WB, Tsongalis GJ, eds: Molecular Pathology. Burlington, MA, Academic Press. © 2009, Elsevier Inc. All
rights reserved.)

C. The T-lymphocyte system is responsible for the recogni-

Thymus-derived
tion of antigens on cell surfaces and, thus, monitors self precommitted
from nonself on live cells (Fig. 1.26). lymphocyte
Aggregated
D. The MHC (HLA) system allows T cells to recognize antigen Macrophage
foreign antigen in cells and then, aided by macrophages, A
mobilizes helper T cells to make killer T cells to destroy
the antigen-containing cells.
E. T lymphocytes, therefore, initiate cellular immunity
(delayed hypersensitivity), are responsible for graft-
versus-host reactions, and initiate the reactions of the B C D
body against foreign grafts such as skin and kidneys
(host-versus-graft reactions).
F. When activated (by an antigen), they liberate lympho-
kines such as macrophage inhibition factor (MIF), mac-
Sensitized
rophage activation factor (MAF), interferon (IFN), and Lymphoblast lymphocyte
E
interleukins IL-2 (previously called T-cell growth factor),
Fig. 1.26 Cellular immunity. A, The participants in the cellular immune
IL-3, and IL-15 (Fig. 1.27).
response include the thymus-derived precommitted lymphocyte (T cell),
bone marrow-derived monocyte (macrophage), and the aggregated anti-
The proliferation and differentiation of T lymphocytes gens. B, Aggregated antigen is seen attaching to the surface of the
macrophage. C, The T cell is shown as it attaches to the aggregated
are regulated by cytokines that act in combination
antigen. D, The substance originating in the macrophage passes into
with signals induced by the engagement of the T-cell the T cell, which is attached to the antigen. E, The combined T cell,
antigen receptor. A principal cytokine is IL-2, itself a antigen, and macrophagic material causes the T cell to enlarge into a
product of activated T cells. IL-2 also stimulates B lymphoblast. Sensitized or committed T lymphocytes arise from lym-
cells, monocytes, lymphokine-activated killer cells, and phoblasts. (From Yanoff M, Fine BS: Ocular Pathology: A Color Atlas,
glioma cells. Another growth factor that stimulates 2nd edn. New York, Gower Medical. © Elsevier 1992.)
22 CHAPTER 1 Basic Principles of Pathology
Sensitized T lymphocyte
Uncommitted lymphocyte
Polymorphonuclear lymphocyte
Monocyte Capillary
Antigen
Tuberculosis
organisms
within
macrophage
A B
C D
Fig. 1.27 Cellular immunity. A, Sensitized T lymphocytes (SL) are seen
in a capillary. Along with the SL are other leukocytes, including mono-
cytes, at an antigenic site. A macrophage, which contains tubercle bacilli
and antigen, may be seen in the surrounding tissue. B, Monocytes
become sensitized when cytophilic antibody from SL is transferred to
them. They migrate toward the antigenic stimulus. C, Biologically active
molecules, which cause the monocytes and leukocytes to travel to the
area, are released by SL when they have encountered a specific antigen.
D, Monocytes arriving at the site are immobilized by migration inhibitory
factor (MIF), which is released by SL, which also release cytotoxin and
mitogenic factor. Cytotoxin causes tissue necrosis (caseation), and mito-
genic factor causes proliferation of cells. Some of these cells undergo
transformation, becoming epithelioid cells, causing the formation of a
tuberculoma. (From Yanoff M, Fine BS: Ocular Pathology: A Color Atlas,
2nd edn. New York, Gower Medical. © Elsevier 1992).
the proliferation of T lymphocytes, the cytokine IL-15,
competes for binding with IL-2 and uses compo-
nents of the IL-2 receptor. T lymphocytes will not
go “into action” against an “enemy” unless they
are triggered by several signals at once. When one
of the signals needed is lacking, the T cell becomes
“paralyzed” (anergy).
G. T lymphocytes also regulate B-cell responses to antigens
by direct contact and by the release of diffusible factors
that act as short-range stimulators of nearby B cells.
H. Many reactions in cellular immunity are mediated by
lymphocyte-derived soluble factors known collectively
as lymphokines, which exert profound effects on inflam-
matory cells such as monocytes, neutrophils, and lym-
phocytes. Such action falls into three main categories:
(1) effects on cell motility (migration inhibition, che-
motaxis, and chemokinesis); (2) effects on cell prolifera-
tion or cellular viability; and (3) effects on cellular
activation for specific specialized functions.
V. The B lymphocyte also arises from lymphoid stem cells in
the bone marrow, but it is not influenced by the thymus.
A. It resides in follicular areas in lymphoid organs distinct
from the sites of the T lymphocyte.
Thymus-derived
Aggregated antigen lymphocyte
Bone marrow-
Macrophage derived
lymphocyte
A
Immunoglobulin
(antibody)
Plasma cell
B C
Fig. 1.28 Humoral immunity. A and B, Four prerequisites for immuno-
globulin formation are demonstrated, including thymus-derived lymphocyte
(T cell), thymus-independent bone marrow-derived lymphocyte (B cell),
bone marrow-derived monocyte (macrophage), and aggregated antigen.
In A, aggregated antigens are seen attached to macrophages. In B, T
and B cells are seen attached to different determinants on the aggre-
gated antigen. C, Cooperative interaction that occurs between T and B
cells causes the B cells to differentiate into plasma cells. (From Yanoff
M, Fine BS: Ocular Pathology: A Color Atlas, 2nd edn. New York, Gower
Medical. © Elsevier 1992.)
B. The B-lymphocyte system is characterized by an enor-
mous variety of immunoglobulins having virtually all
conceivable antigenic specificities that are capable of
being recognized by at least a few B-lymphocyte clones.
After germinal center B cells undergo somatic muta-
tion and antigen selection, they become either
memory B cells or plasma cells. CD40 ligand directs
the differentiation of germinal center B cells toward
memory B cells rather than toward plasma cells.
C. The system is well designed to deal with unpredictable
and unforeseen microbial and toxic agents.
D. The B lymphocyte can be stimulated by antigen to enlarge,
divide, and differentiate to form antibody-secreting
plasma cells (Fig. 1.28).
In most circumstances, T lymphocytes collaborate
with B lymphocytes during the induction of antibody-
forming cells by the latter (see the section on humoral
immunoglobulin, later).
VI. Null lymphocytes, which constitute approximately 5% of
lymphocytes in peripheral blood, lack the surface markers
used to identify T and B lymphocytes.
A. Most null cells carry a surface receptor for the Fc portion
of the immunoglobulins, can function as killer cells in
antibody-dependent cell-mediated cytotoxicity, and are
called NK cells.

B. When stimulated, NK cells release perforin, which forms
pores in the cell membrane.
C. They also release substances through the pores that can
precipitate apoptosis in the target cell.
VII. Initially, the sheep red blood cell resetting test (especially
with fixed, embedded tissue) and the immunofluorescence
or immunoperoxidase techniques that demonstrate surface
immunoglobulins were the principal techniques for iden-
tification of T or B lymphocytes, respectively.
A. Now, monoclonal antibodies (especially with fresh tissue)
are used for the localization of lymphocyte subsets in
tissue sections, and their use has revolutionized research
in immunology, cell biology, molecular genetics, diag-
nosis of infectious diseases, tumor diagnosis, drug and
hormone assays, and tumor therapy.
B. A myriad of different types of monoclonal antibod-
ies now exist, and new ones are continuously being Humoral Immunoglobulin (Antibody)
created.
C. Monoclonal antibodies can be obtained against B and
T lymphocytes, monocytes, Langerhans’ cells, keratins,
type IV collagen, retinal proteins (e.g., human S-100),
and tumor antigens (e.g., factor VIII and intermedi-
ate filaments—cytokeratins, vimentin, desmin, neuro-
filaments, and glial filaments—neuron-specific enolase,
and glial fibrillary acidic protein; all may be found in
tumors).
Cellular Immunity (Delayed Hypersensitivity)
I. Two distinct cell types participate in cellular immunity: the
T lymphocyte and the macrophage (histiocyte).
A. Phagocytic cells of the monocytic line (monocytes,
reticuloendothelial cells, macrophages, Langerhans’ den-
dritic cells, epithelioid cells, and inflammatory giant
cells—all are different forms of the same cell) are devoid
of antibody and immunologic specificity.
1. Macrophages, however, have the ability to process
proteins (antigens) and activate the helper T cells.
2. Macrophages also secrete proteases, complement
proteins, growth-regulating factors (e.g., IL-1), and
arachidonate derivatives.
B. All lymphocytes seem to be pre-committed to make
only one type of antibody, which is cell-bound.
TABLE 1.7 Antibody Classes and Functions
Class Location* Structure
IgD Surface of B cells only 2 κ or λ light chains, 2 δ heavy chains
IgM Plasma 2 κ or λ light chains, 2 µ heavy chains, arranged
in pentamers with 1 J chain
IgG Widely distributed in 2 κ or λ light chains, 2 γ heavy chains
extracellular fluid
IgA Mucosal tissues, surfaces, 2 κ or λ light chains, 2 α heavy chains, arranged
and secretions in dimers with 1 J chain
IgE Bound to mast cells and 2 κ or λ light chains, 2 ε heavy chains
basophils
*All immunoglobin classes are found on B cells as antigen receptors.
(Reproduced from Table 3.2, Coleman WB, Tsongalis GJ, eds: Molecular Pathology. Burlington, MA, Academic Press. © 2009, Elsevier Inc. All
rights reserved.)
Immunobiology 23
II. The delayed hypersensitivity reaction begins with perivenous
accumulation of sensitized lymphocytes and other MN cells
(i.e., monocytes, which constitute 80%–90% of the cells
mobilized to the lesion). The infiltrative lesions enlarge
and multiply (e.g., in tuberculosis, where the lesions take
a granulomatous form), and cellular invasion and destruc-
tion of tissue occur.
III. Delayed hypersensitivity is involved in transplantation
immunity; in the pathogenesis of various autoimmune
diseases (e.g., sympathetic uveitis); and in defense against
most viral, fungal, protozoal, and some bacterial diseases
(e.g., tuberculosis and leprosy). Perhaps the most important
role is to act as a natural defense against cancer—that is,
the immunologic rejection of vascularized tumors and
immunologic surveillance of neoplastic cells.
I. Four distinct cell types participate in humoral immu-
noglobulin (antibody) formation: the T lymphocyte,
the B lymphocyte, the monocyte (macrophage), and the
plasma cell.
A. Macrophages process antigen in the early stage of the
formation of cellular immunity and secrete IL-1.
B. Specifically, pre-committed cells of both T and B lym-
phocytes attach to different determinants of the antigen;
T cells then secrete a B-cell growth factor (BCGF).
C. BCGF and IL-1 evoke division of triggered B cells, which
then differentiate and proliferate into plasma cells that
elaborate specific immunoglobulins. All humoral immu-
noglobulins (antibodies) are made up of multiple poly-
peptide chains and are the predominant mediators of
immunity in certain types of infection, such as acute
bacterial infection (caused by streptococci and pneu-
mococci) and viral diseases (hepatitis).
II. The B lymphocyte, once a specific antigen causes it to become
committed (sensitized) to produce an immunoglobulin,
makes that immunoglobulin and none other, as does its
progeny. It, or its progeny, may produce immunoglobulin
or become a resting memory cell to be reactivated at an
accelerated rate (anamnestic response) if confronted again
by the same antigen. Table 1.7 enumerates the immuno-
globulin classes and functions produced by B cells.
Function
Unknown; expressed early in differentiation along with IgM
Activated complement; first functional immunoglobulin formed
in immune response
Complement activation, transfer to neonate via placenta,
opsonization, neutralization of viruses and other pathogens
Important in mucosal immunity; has opsonizing activity
Binds to and activates mast cells and basophils; important in
defense versus parasites
24 CHAPTER 1 Basic Principles of Pathology
Autoimmunity and Autoinflammation
I. Autoimmune diseases are caused by abnormalities in adap-
tive immunity regulation, while autoinflammatory disorders
are attributed to defects in innate immunity proteins and
are characterized by the absence of pathogenic autoanti-
bodies or autoreactive T cells. In both situations, the patient’s
own immunological systems becomes a source of tissue
damage rather than its protector.
II. Monogenic autoinflammatory syndromes have been defined
as inherited conditions caused by mutations in one or both
copies of a single gene that result in over-activation of the
innate immune system causing inappropriate inflammation.
A. Appear to be unprovoked attacks of inflammation most
commonly directed at the eye, skin, joints, and gut.
B. Mechanisms by which genetic defects cause autoinflam-
matory disease:
1. Affect intracellular sensor function.
2. Lead to accumulation of intracellular triggers that
cause cell stress and activate intracellular sensors.
3. Cause loss of a negative regulator of inflammation.
4. Affect signaling molecules that upregulate innate
immune cell function.
C. Mediated by IL-1 secretion stimulated by monocytes
and macrophages.
D. Induction of inflammation in many of these disorders
is triggered by the inflammasome pathway (see discus-
sion above regarding inflammasomes, pyroptosis, and
associated monogenic ocular disorders).
E. IL-1 secretion in response to Toll-like receptor stimula-
tion, and ultimately, the triggering of NLRP3 inflam-
masome may occur not only in response to exogenous
microbial stimulation, but also to “endogenous stress
molecules” by setting off an autoinflammatory process.
F. Other monogenic autoinflammatory disorders arise from
perturbations in signaling by the transcription factor
NF-κB, ubiquitination, cytokine signaling, protein
folding, and type I interferon production, and comple-
ment activation.
G. Some immunologic diseases have combined fea-
tures of autoinflammation, autoimmunity, and/or
immunodeficiency.
A B
Fig. 1.29 Immunocytochemistry. A, Cathepsin-D, which here stains cytoplasm of conjunctival submucosal
glands (shown under increased magnification in B), is an excellent stain for lipofuscin.
H. Monogenic autoinflammatory syndromes include
idiopathic granulomatous diseases, familial Mediter-
ranean fever (FMF), TNF receptor–associated periodic
syndrome (TRAPS), deficiency of mevalonate kinase
(MKD), cryopyrin-associated periodic fever syndrome
(CAPS, consisting of familial cold autoinflammatory
syndrome or FCAS, Muckle–Wells syndrome or MWS,
and chronic infantile neurologic cutaneous articular
syndrome or CINCA syndrome).
III. The eye is considered an immunologically privileged site due
to 1) absence of blood and lymphatic vessels in the anterior
chamber, 2) anterior chamber–associated immune deviation
(ACAID) that controls the proinflammatory milieu, and 3)
retinal protection identified in phagocytosis of damaged
receptors and retinal pigment epithelium, which also helps
construct part of the blood–retinal barrier through its tight
intercellular junctions. Another important component of
the blood–retinal barrier resides in the tight junctions
between retinal vascular endothelial cells. Nevertheless,
autoimmune disorders do occur and may have devastating
consequences.
A. A particularly devastating but, fortunately, rare auto-
immune disorder is sympathetic ophthalmia in which
the ocular immune barriers are breached, and autoim-
munity develops to uveal protein resulting in a delayed
hypersensitivity reaction characterized by diffuse granu-
lomatous inflammation involving the entire uveal tract
in both the inciting and sympathizing eyes.
Immunohistochemistry
I. As stated previously, monoclonal antibodies can be obtained
against B and T lymphocytes, monocytes, Langerhans’ cells,
keratins, type IV collagen, retinal proteins, and so forth
(Figs. 1.29 and 1.30). Table 1.8 lists antibody tests that
are helpful in differentiating various tumors when they
lack sufficient differentiation for accurate microscopic
diagnosis.
II. Commonly used antibodies include the following:
A. Cytokeratins (AE1/AE3, CAM 5.2, CK7, and CK20) and
epithelial membrane antigen are markers for epithelia.
B. Factor VIII and Ulex europaeus-1 are markers for vascular
endothelia.
 Immunobiology 25

A B

C D

Fig. 1.30 Immunocytochemistry. A, Monoclonal antibody against desmin, one of the cytoskeletal filaments,
reacts with both smooth and striated muscles, and it helps to identify tumors of muscular origin. B, Mono-
clonal antibody against λ chains in plasma cells. C and D, Polyclonal antibody against S-100 protein in mela-
nocytes and Langerhans’ cells in epidermis (C) and in malignant melanoma cells (D). (From Schaumberg-Lever
G, Lever WF: Color Atlas of Pathology of the Skin. Philadelphia, Lippincott, 1988, with permission.)

C. Intermediate filaments: Vimentin is a marker for mes-
enchymal cells, including smooth muscle, Schwann cells,
histiocytes, and fibrocytes; desmin is a marker for smooth
and striated muscles; cytokeratin is a marker for epithelia;
neurofilament is a marker for neurons; and glial fibrillary
acidic protein is a marker for astrocytes and Schwann
cells.
D. Neuron-specific enolase is a marker for Schwann cells,
neurons, smooth muscle, and neuroendocrine cells.
E. S-100 is a marker for neural crest-derived tissues includ-
ing melanocytes, but only melanocytic tumors should
be positive for HMB45 and Mel-A.
F. Smooth muscle actin (SMA) is a marker for smooth
muscles and myoepithelial cells.
G. Many antibodies are available for immunophenotyping
of lymphomas and leukemias, both on fresh and on
paraffin-embedded tissue. Table 1.9 lists the immuno-
histochemical paradigm for differentiating hematolym-
phoid neoplasms.
H. Many other markers are available, and new markers seem
to appear almost weekly.
1. Useful websites for further information regarding
immunohistochemical stains and techniques include
the following:
a. http://www.immunoquery.com
2. Throughout this textbook, appropriate key immu-
nohistochemical markers are cited where appropriate
for each histopathologic diagnosis.
3. Similarly, although genetics is not the focus of this
textbook, critical genetic abnormalities are highlighted
as appropriate.
Immunodeficiency Diseases
I. The following are disorders associated with immunodefi-
ciency discussed elsewhere in this textbook:
A. Wiskott–Aldrich syndrome (see Chapter 6)
B. Ataxia–telangiectasia (see Chapter 2)
C. Chédiak–Higashi syndrome (see Chapter 11)
II. Severe combined immunodeficiencies (SCIDs)—hetero-
geneous group of inherited disorders characterized by 1)
absence or very low number of T cells (<300 CD3 T cells/
mm3) and no or very low T-cell function (<10% of the
lower limit of normal) as measured by response to PHA
or 2) presence of T cells of maternal origin. The pres-
ence or absence of B and NK cells has permitted clini-
cians to direct attention to certain genetic defects. SCIDs
occur in approximately 1 : 50 000 live births, and they are
more common in males because of the prevalence of
X-linked SCID.
III. Chronic granulomatous disease of childhood (see Chapter 4)
26 CHAPTER 1 Basic Principles of Pathology

TABLE 1.8 Antibodies Useful in Determining the Origin of Undifferentiated Tumors and
Tumors of Uncertain Primary Site
Panel Antibodies Panel Antibodies
Undifferentiated tumors Pan-keratin Liver panel α-Fetoprotein
CD45 (CLA) α1-Antitrypsin
S-100 α1-Antichymotrypsin
Vimentin Nonsquamous keratin
Carcinoma panel Pan-keratin Hepatitis B surface antigen
CK5/6 Hepatitis B core antigen
AE1 Mesothelioma panel Squamous keratin
AE3 Nonsquamous keratin
CAM5.2 CEA-negative
MAK6 EP4 (epithelial antigen)-negative
Squamous keratin (HMW) CD15 (Leu M1)-negative
Nonsquamous keratin (LMW) Epithelial membrane antigen
Sarcoma panel Pan-keratin B72.3
Vimentin Secretory component
S-100 Vimentin
Desmin OC-125
CD45 (CLA) Melanoma panel S-100
Actin (muscle/HHF-35-MSA) HMB-45
Actin (smooth muscle-specific—SMA) Melan A
Myoglobin (skeletal muscle) Vimentin
LN-5 (histiocytes) Pan-keratin
Lysozyme (histiocytes) Central nervous system/neural panel Glial fibrillary acidic protein (GFAP)
LN-6 (nonlymphoid vimentin) Neurofilament
Factor VIII antigen (endothelial cells) S-100
CD34 (vascular antigen) NSE
CD31 (vascular antigen) Vimentin
Ulex (vascular antigen) Pan-keratin
O13 (Ewing’s sarcoma/PNET) Synaptophysin
Breast panel BRST-2 (GCDFP) Neuroendocrine panel NSE
Mammaglobin Chromogranin
Cu-18 (breast-related antigen) Serotonin
Lactalbumin Neuron-endocrine
Prognosis (breast carcinoma) Estrogen receptor (monoclonal) Synaptophysin
Progesterone receptor (monoclonal) Nonsquamous keratin
Her-2/neu (c-erb B-2) CD57 (Leu 7, HMK 1)
p52 (luminal epithelial antigen) Vasointestinal peptide
p53 Pituitary hormone panel Adrenocorticotropic hormone (ACTH)
Factor VIII antigen Follicle-stimulating hormone (FSH)
Lung panel CK7 Growth hormone
CK20 Luteinizing hormone
TTF-1 Prolactin
Prostate panel Prostate-specific antigen (PSA) Thyroid-stimulating hormone (TSH)
Prostatic acid phosphatase (PAP) α-Subunit
Androgen receptor PIT-1
34BE12 (SK) Pancreatic panel Amylase
Gastrointestinal panel CEA Insulin
COTA Glucagon
CDX-2 Gastrin
CK7 Somatostatin
CK20 CDX-2
Kidney/bladder panel Renal antigen Urothelial panel p63
CEA Uroplakin
p53
Ovary panel COTA
OC-125 (CA-125)
Estrogen receptor
Progesterone receptor

CEA, carcinoembryonic antigen; CLA, common leukocyte antigen; COTA, colonic ovarian tumor antigen; HMW, high molecular weight; LMW,
low molecular weight; NSE, neuron-specific enolase; PNET, primitive neuroectodermal tumor.
(Reproduced from Table 5.2, Weidner N, Cote RJ, Suster S et al., eds: Modern Surgical Pathology, 2nd edn. Philadelphia, Saunders. © 2009,
2003 by Saunders, an imprint of Elsevier Inc.)
 Immunobiology 27

TABLE 1.9 Diagnostic Algorithm for Hematolymphoid Neoplasms

First Choice Consistent With Tumor
Antibody Panel Second Choice Antibody Panel Additional Antibodies Type
CD3−; CD20−/+ CD10 CD34 CD79a Precursor B-Cell Neoplasms
− + −/+ CD22−/+; TdT+ Pro-B-ALL (B-1)
+ + + CD22−/+; TdT+ Common ALL (B-II, early pre-B)
+ −/+ + CD22−/+; TdT−/+ Pre-B-ALL (B-III, late pre-B)

CD3−/CD20+ CD5 CD10 CD23 CD43 Mature B-Cell Neoplasms

+ − − + Cyclin D1+ Mantle cell lymphoma
+ − + + Cyclin D1−; CD38+/− CLL/SLL
− + +/− −/+ BCL2+/−; BCL6+ Follicular cell lymphoma
− + − +/− BCL2−; Ki67+; BCL6+ Burkitt’s lymphoma
− −/+ − −/+ CD25+; DBA44+; TRAP+; Hairy cell leukemia
− − −/+ −/+ Correlation with morphology is critical MALT lymphoma
− − − − B-cell prolymphocytic leukemia
Lymphoplasmacytic lymphoma
CD45+; BCL6+/−; Bob1 or OCT2+ NLPHD

CD3/CD20− CD30 CD38 CD45

+/− + + CD138+; K/L (Clonal) Plasma cell neoplasm
+ − − ALK+; BCL6+/−; EMA+/−; CD4+/− Anaplastic large cell lymphoma
+ − R/− (focal) CD15+; Fascin+; EBV+/−; Bob1/OCT2+ Classic Hodgkin’s lymphoma
−/+ − +/− S100+; CD1a+; CD68−/+ Langerhans’ cell histiocytosis
CD68+; CD4+; CD43+ True histiocytic neoplasm
CD21+; CD23+; CD35+; Follicular dendritic cell
S100−/+; CD4− neoplasm
S100+; CD4+/−; Dendritic cell neoplasm other
CD68−/+ than follicular
− − +/− MPO+; Lysozyme; CD34−/+; C-kit+; Acute myeloid leukemia
TdT−/+

(Reproduced from Figure 5.5, Weidner N, Cote RJ, Suster S et al., eds: Modern Surgical Pathology, 2nd edn. Philadelphia, Saunders. © 2009,
2003 by Saunders, an imprint of Elsevier Inc.)

IV. Acquired immunodeficiency syndrome (AIDS) (Table 1.10)
A. The first five cases of what came to be called “AIDS” in
the United States presented with Pneumocystis carinii
(now termed Pneumocystis jiroveci) pneumonia, cyto-
megalovirus (CMV) infections, and candidiasis. Thus,
ocular manifestations were among the presenting find-
ings in these first AIDS patients.
B. In 2013 there were 1.2 million people living with HIV
and there were 40,630 new cases in the U.S. that year.
In 2017, worldwide, an estimated 33 million people were
HIV-positive and 30% didn’t know their status. More
than two-thirds of HIV-infected individuals are in sub-
Saharan Africa.
C. From an ocular perspective, AIDS is the most important
immunodeficiency disease.
D. AIDS is caused by a retrovirus, the human immunode-
ficiency virus (HIV), which is highly lethal if not treated
with HAART (highly active antiretroviral therapy).
Worldwide, 77% of individuals diagnosed HIV-positive
are on antiretroviral therapy.
1. Excess mortality associated with AIDS has been
reduced 50% since the introduction of HAART
therapy. Nevertheless, excess mortality is five times
higher in AIDS patients than in HIV-infected patients
without AIDS.
2. HIV-1 and HIV-2 have an affinity for the CD4 antigen
on T lymphocytes, macrophages, and other cells.
HIV-2 typically is less virulent than HIV-1 and gen-
erates a more effective host response.
E. HIV-1 consists of an electron-dense core surrounding
a single-stranded RNA genome, both enveloped by a
cell membrane. Retroviruses contain DNA polymerase
(reverse transcriptase) complexed to the RNA in the
viral core. Reverse transcriptase catalyzes the transcrip-
tion of the RNA genome into DNA form (the provirus).
The provirus migrates from the host cell’s cytoplasm to
the nucleus, assumes a double-stranded circular form,
integrates into the host cell DNA, and may remain
throughout the life of the host cell.
F. Ironically, although immunodeficiency is the hallmark
of HIV-AIDS, the use of inflammatory pathways by the
virus are critical to its establishment and spread within
its host.
1. HIV immune activation is an essential component
of HIV pathobiology by increasing proinflammatory
mediators, dysfunctional regulatory T cells, and a
28 CHAPTER 1 Basic Principles of Pathology
TABLE 1.10 Human Immunodeficiency
Virus-Related Ophthalmic Disorders
I. Opportunistic infections
A. Retina
1. CMV retinitis
a. Complications
(1) Immune recovery uveitis
2. Other retinal infections (caused by various agents, VZV, and
Toxoplasma gondii being most common; most occur in less than
1% of patients with AIDS)
B. Choroid (uncommon; caused by various agents, fungi and
mycobacteria being most common)
C. Ocular surface and adnexa (important agents include VZV,
microsporidia, molluscum contagiosum virus)
II. Vascular abnormalities
A. Microvasculopathy
1. HIV retinopathy (cotton-wool spots, retinal hemorrhages)*
B. Retinal arteriolar and venular occlusions (uncommon)
III. Neoplasia†
A. Kaposi sarcoma (conjunctiva, eyelids)
B. Lymphoma (intraocular)
C. Squamous cell carcinoma (conjunctiva)
IV. Other disorders of uncertain pathogenesis
A. Intraocular inflammation
1. Chronic anterior uveitis (uncommon)
2. Chronic multifocal retinal infiltrates (uncommon)‡
3. Iatrogenic uveitis (drug related: cidofovir; rifabutin)
B. Blepharitis
C. Dry eye
V. Neuro-ophthalmic disorders associated with orbital or intracranial
disease
AIDS, acquired immunodeficiency syndrome; CMV, cytomegalovirus;
HIV, human immunodeficiency virus; VZV, varicella–zoster virus.
*Clinical signs reflect focal ischemia, attributable to undetermined
factors, on a background of the retinal microvasculopathy of HIV
disease.
†
Infection has been shown to be involved in the pathogenesis of
tumors in severely immunodeficient individuals.
‡
As described in Levinson RD, Vann R, Davis JL, et al.: Chronic
multifocal retinal infiltrates in patients infected with human
immunodeficiency virus. Am J Ophthalmol 125:312, 1998.
(Reproduced from Holland GN: AIDS and ophthalmology: The first
quarter century. Am J Ophthalmol 145:397, 2008.)
pattern of T-cell senescent phenotypes as are seen
in elderly individuals.
2. Chemokines, which are small chemotactic cytokines,
and their receptors contribute to the “cytokine storm”
that is one of the salient features of HIV infection.
3. Apoptosis contributes significantly to T-cell depletion
and this process is mediated by caspase-3. The process
of pyroptosis also plays a role in CD4+ T-cell deple-
tion with dying T cells releasing inflammatory signals
that perpetuate the destructive process (see discussion
of the inflammasome earlier in this chapter).
G. Patients with AIDS are prone to life-threatening oppor-
tunistic infections, wasting, central nervous system dys-
function, generalized lymphadenopathy, and Kaposi’s
sarcoma and other malignancies, particularly squamous
neoplasia.
H. Cytomegalovirus is the most common ocular opportunis-
tic agent in AIDS even following the advent of HAART
therapy. Individuals with CD4+ T-cell counts of 50 cells/µl
or less are at particular risk for ocular complications of
CMV infection. The other agents include most of the
viral, bacterial, fungal, and parasitic agents customarily
associated with cellular immunodeficiency, with herpes
simplex virus, Mycobacterium tuberculosis and Mycobacte-
rium avium-intracellulare, cat-scratch bacillus (Bartonella
henselae), Candida albicans, Cryptococcus neoformans,
Pneumocystis carinii, and Toxoplasma gondii heading
the list.
I. HIV-associated neurologic disorders (HAND) have been
found in up to 70% of HIV-infected individuals. For-
tunately, new cases of HAND have decreased by 75%
since the advent of HAART. From the ocular perspective,
approximately 10%–15% of patients with AIDS, but
without ocular opportunistic infection have a presumed
neuroretinal disorder as evidenced by reduced contrast
sensitivity and abnormal visual fields.
1. Abnormal contrast sensitivity is associated with
increased mortality in these patients and may indicate
the presence of life-threatening microvascular disease.
2. Narrowed retinal vascular caliber is present in AIDS
and is associated with an increased mortality risk.
3. Neuroretinal disorders in these patients are associated
with specific mitochondrial haplogroups.
J. The histologic appearance depends on the site of involve-
ment and the causative agent (see appropriate sections
of this book).
K. HIV also predisposes to malignancies, particularly
Kaposi’s sarcoma; however, other tumors, such as con-
junctival lymphoma, are increased in frequency in these
patients.
L. HIV-associated complications also include dyslipidemia,
hyperglycemia, and loss of bone mineral density, and
autoimmune disorders such as autoimmune thrombo-
cytopenia, connective tissue disorders, spondyloarthritis,
and others.
M. Immune recovery uveitis (IRU).
1. Usually occurs when previously immune-deficient
patients with AIDS have their immune response to
CMV restored by HAART therapy. It also may be
associated with other pathogens such as T. gondii,
M. avium complex, and Leishmania sp.
2. Uveitis may occur within several weeks of starting
HAART therapy.
3. Represents a change in inflammatory reaction rather
than an absolute level of inflammation or specific
complications.
4. It may develop in as many as 25% of patients with
advanced immunodeficiency who start HAART
therapy.
a. Larger CMV retinitis lesions are associated with
a greater risk of IRU.
5. Ocular findings may include anterior segment inflam-
mation, cataract, vitritis, papillitis, cystoid macular

edema, epiretinal membrane, vitreous hemorrhage,
retinal neovascularization, and proliferative vitreo-
retinopathy. Severe visual loss may result.
6. The presence of HLA-B 8–18 may be associated with
IRU. Other genetic factors may modulate the impact
of HIV infection on a given patient.
7. Cytomegalovirus is the most common underlying
infection leading to IRU.
Transplantation Terminology
I. Autograft: transplantation of tissue excised from one place
and grafted to another in the same individual.
II. Syngraft (isograft): transplantation of tissue excised from
one individual and grafted to another who is identical
genetically.
III. Allograft (homograft): transplantation of tissue excised from
one individual and grafted to another of the same species.
IV. Xenograft (heterograft): transplantation of tissue excised
from one individual and grafted to another of a different
species.
V. Orthotopic graft: transplantation to an anatomically correct
position in the recipient.
VI. Heterotopic graft: transplantation to an unnatural
position.
CELLULAR AND TISSUE REACTIONS
Hypertrophy
Hypertrophy is an increase in size of individual cells, fibers, or
tissues without an increase in the number of individual elements
(e.g., retinal pigment epithelium [RPE] in RPE hypertrophy).
Hyperplasia
Hyperplasia is an increase in the number of individual cells in
a tissue; their size may or may not increase. Therefore, hyperplasia
is cellular proliferation in excess of normal, but the growth even-
tually reaches an equilibrium and is never indefinitely progressive
(e.g., RPE hyperplasia secondary to trauma; see section on neo-
plasia, later).
Aplasia
Aplasia is the lack of development of a tissue during embryonic
life (e.g., aplasia of the optic nerve).
Hypoplasia
Hypoplasia is the arrested development of a tissue during embry-
onic life (e.g., hypoplasia of the iris [aniridia]).
Metaplasia
Metaplasia is the transformation of one type of adult tissue into
another type (e.g., fibrous metaplasia of lens epithelium [in
anterior subcapsular cataract]).
Atrophy
Atrophy is a diminution of size, a shrinking of cells, fibers, or
tissues that had previously reached their full development (e.g.,
retinal vascular atrophy in retinitis pigmentosa).
Cellular and Tissue Reactions 29
Dysplasia
Dysplasia is an abnormal growth of tissue during embryonic
life (e.g., retinal dysplasia).
Neoplasia
I. Neoplasia is a continuous increase in number of cells in a
tissue, caused by unregulated proliferation and, in some
cases, failure of mechanisms (e.g., apoptosis) that lead to
cell death.
A. The neoplastic proliferation is probably caused by either
excessive or inappropriate activation of oncogenes or
reduced activity of genes that downregulate growth
(anti-oncogenes).
B. It differs from hyperplasia in that its growth never attains
equilibrium.
C. The neoplasm may be benign or malignant.
II. A malignant neoplasm differs from a benign one in being
invasive (it infiltrates and actively destroys surrounding
tissue), in having the ability to metastasize (develop sec-
ondary centers of neoplastic growth at a distance from the
primary focus), and in showing anaplasia (histologically,
the features of a malignancy that include variation from
the normal structure [Fig. 1.31] or behavior in the sense
of a loss of specialized or “adult” characteristics of the cell
or tissue, e.g., loss of cellular or tissue polarity, or inability
to form photoreceptors).
Mutations in the p53 tumor suppressor gene, located
on the short arm of chromosome 17 at position 17p13.1,
represent the most frequent genetic alteration detected
in human solid malignancies. In more than half of all
human cancer cases, p53 is inactivated by mutations
and other genomic alterations. Mutations in p53 also
can result in altered p53 proteins and endow these
mutant proteins with novel activities. The p53 gene
encodes a 53-kDa nucleophosphoprotein that binds DNA,
is involved in the regulation of transcription and the
induction of programmed cell death (apoptosis), and
negatively regulates cell division, preventing progression
from G to S phase. Normally, p53 responds to a complex
network of stress signals. When functioning properly,
it has two responses to stress: either cell-cycle arrest
Fig. 1.31 Abnormal tripolar mitotic figure in a sebaceous gland carcinoma.
30 CHAPTER 1 Basic Principles of Pathology

until DNA damage is repaired or apoptotic cell death if

TABLE 1.11 Features of Necrosis and
such repair has failed. It appears to be a marker of tumor
progression (i.e., a direct correlation seems to exist
Apoptosis
between mutations at the p53 locus and increasing Feature Necrosis Apoptosis
histologic grade).
Cell size Enlarged (swelling) Reduced (shrinkage)
Nucleus Pyknosis → Fragmentation into
karyorrhexis → nucleosome-size
Degeneration and Dystrophy karyolysis fragments
Plasma membrane Disrupted Intact; altered structure,
I. A dystrophy is a primary (bilateral), inherited disorder that
especially orientation of
has distinct clinicopathologic findings. The individual dys-
lipids
trophies are discussed elsewhere under their individual Cellular contents Enzymatic digestion; Intact; may be released in
tissues. may leak out of apoptotic bodies
II. A degeneration (monocular or binocular) is a secondary cell
phenomenon resulting from previous disease. It occurs in Adjacent Frequent No
a tissue that has reached its full growth. inflammation
A. Cloudy swelling is a reversible change in cells secondary Physiologic or Invariably pathologic Often physiologic, means
to relatively mild infections, intoxications, anemia, or pathologic role (culmination of of eliminating unwanted
circulatory disturbances. The cells are enlarged and filled irreversible cell cells; may be pathologic
with granules or fluid and probably represent an intra- injury) after some forms of cell
injury, especially DNA
cellular edema.
damage
B. Hydropic degeneration is a reversible change in cells also
secondary to relatively mild infections, intoxications, (Reproduced from Table 1.2, Kumar R, Abbas A, DeLancey A et al.:
anemia, or circulatory disturbances. The cells are enlarged Robbins and Cotran Pathologic Basis of Disease, 8th edn.
Philadelphia, Saunders. © 2010 by Saunders, an imprint of Elsevier
and contain cytoplasmic vacuoles and probably represent
Inc.)
an early stage of swelling of the endoplasmic reticulum.
C. Fatty change results when fat accumulates in cells for
unknown reasons or after damage by a variety of agents
(e.g., chloroform and carbon tetrachloride). 2. Organelle dissolution and rupture of the plasma
D. Glycogen infiltration results from diseases such as diabetes membrane.
mellitus (e.g., lacy vacuolation of iris pigment epithelium; 3. Leakage of cellular contents into the extracellular
see Chapter 15) and from a lack of nutrition (e.g., in space.
long-standing neural retinal detachment and in prolif- 4. Inflammatory response to the released cellular debris.
erating retinal pigment epithelial cells). B. No inflammation occurs in apoptosis—see later.
E. Amyloid may be found in ocular tissues in primary amy- II. Coagulative necrosis: This is a firm, dry necrosis generally
loidosis (see Chapters 7, 8 and 12), such as in primary formed in tissue that has been shut off from its blood supply.
familial amyloidosis and lattice corneal dystrophy (in A. The gray, opaque clinical appearance of the retina after
which case it is a dystrophic change), or in secondary a central retinal artery occlusion is caused by coagula-
amyloidosis (see Chapter 7), in which case it is a degen- tive necrosis (ischemic necrosis). As seen by electron
erative change. microscopy, coagulative necrosis (e.g., after a laser burn)
F. Hyaline degeneration is quite common; consists of acel- is produced by widespread focal densification of mem-
lular, amorphous, and eosinophilic material; and may branes in the necrotic cell.
be found in places such as the walls of arteriolosclerotic B. Caseation, characteristic of tuberculosis, is a combination
vessels or in the ciliary processes in elderly people. of coagulative and liquefaction (see later) necrosis.
III. Hemorrhagic necrosis: This type is caused by occlusion of
Necrosis (Table 1.11) venous blood flow but with retention of arterial blood flow,
I. Necrosis occurs when cells die an “accidental” death, such as seen classically in central retinal vein thrombosis.
as from severe and sudden injury (e.g., ischemia), sustained IV. Liquefaction necrosis: Necrosis of this type results from
hyperthermia, physical or chemical trauma, complement autolytic (see section on Autolysis and Putrefaction, later)
attack, or metabolic poisons. decomposition, usually in tissue that is rich in proteolytic
enzymes (e.g., suppuration is a form of liquefaction necrosis
Necrosis should be differentiated from apoptosis—see in which rapid digestion is brought about by the proteolytic
later. enzymes from the leukocytes, especially PMNs, present in
the area). It also occurs from complete dissolution of all
A. Necrosis is accompanied by the following: cell components, as in ultraviolet photocomposition.
1. Swelling of the cytoplasm and organelles (especially V. Fat necrosis: Necrosis causes liberation of free fatty
the mitochondria) and only mild changes in the acids and glycerol that results in a lipogranulomatous
nucleus. reaction.
 Cellular and Tissue Reactions 31

a. In the intrinsic pathway, Apaf-1-like molecules

Apoptosis
(See also multiple references to apoptosis throughout this chapter,
particularly relative to complement, the inflammasome, and HIV
infections.)
I. Apoptosis is “physiologic” or programmed cell death, unre-
lated to “accidental death” (necrosis)—see earlier.
A. There are intrinsic and extrinsic pathways of apoptosis,
and a substitute pathway for cell killing (see description,
below, and Fig. 1.32).
1. The extrinsic pathway also, is termed the “death recep-
tor pathway,” and is mediated by the binding of cell-
surface death receptors and their natural ligands.
2. The intrinsic pathway, also termed the “mitochon-
drial pathway,” is strictly regulated by BCL-2 family
proteins.
assemble into a ring-like platform known as the
“apoptosome” that, in turn, activates procaspases.
b. Caspase-9 is activated on the apoptosome, and
then breaks up cells into apoptotic bodies (see
below).
1) Inability to activate caspase-9 can lead to
degenerative and developmental disorders, and
even to cancer.
3. There is a third “substitute pathway” to cell killing
that involves cytotoxic T-cell and natural killer cell–
mediated and perforin–granzyme-dependent func-
tions. In this pathway, granzymes A and B are involved.
B. Two steps accompany apoptosis:
1. The cell undergoes nuclear and cytoplasmic conden-
sation, eventually breaking up into a number of

Caspase-10
Initiator
Caspase-8/10

Caspase-9
Cell destruction
Apoptosis

Fig. 1.32 Illustration of major apoptotic signaling pathways. A, The extrinsic pathway is initiated by the
interaction of death receptors (DRs) and death-inducing ligands via their FADD/TRADD domains. A death-
inducing signaling complex (DISC) is formed, which then recruits inactivated initiator caspase-8 and caspase-
10. After that, BID is cleaved into tBID and translocates to mitochondria. B, In intrinsic apoptotic pathway,
the stimuli first activate BH3-only proteins, and then pass the apoptotic signals via activating interactions
between BCL-2 members, forming a mitochondrial outer membrane permeabilization (MOMP) complex and
eventually releasing cytochrome c. Cytochrome c induces the assembly of the apoptosome, and caspase-9
is activated. The extrinsic and intrinsic apoptotic pathways converge in which caspase-3, caspase-6 and
caspase-7 function as executioner caspases that trigger various biochemical and morphological alterations
in apoptotic cells that are eventually taken up by phagocytes. Inhibitors of apoptosis (IAP) proteins inhibit
both initiator and executioner caspases in apoptotic pathways. C, The substitute pathway is mediated by
cytotoxic T lymphocytes or NK cells which involve perforin and granzyme. (From Wu et al.: Apoptosis signal-
ing and BCL-2 pathways provide opportunities for novel targeted therapeutic strategies in hematologic
malignancies. Blood Rev 32(1):8-28, 2018. Figure 1. Elsevier.)
32 CHAPTER 1 Basic Principles of Pathology

membrane-bound fragments containing structurally II. When certain bacteria (especially clostridia) invade necrotic
intact organelles. (autolytic) tissue, the changes catalyzed by destructive bac-
terial enzymes are called putrefaction.
Cells undergoing apoptosis demonstrate shrink-
age, nuclear condensation associated with DNA Pigmentation
fragmentation, a relatively intact cell membrane, I. In ocular histologic sections stained with H&E, some com-
loss of viability, and absence of inflammation. monly found pigments may resemble each other closely:
(1) melanin and lipofuscin, (2) hemosiderin, (3) exogenous
2. The cell fragments, termed apoptotic bodies, are phago- iron, and (4) acid hematin.
cytosed by neighboring cells and rapidly (within II. Melanin is found in uveal melanocytes as fine, powdery,
minutes) degraded. brown granules barely resolvable with the light microscope
and also in pigment epithelial cells of the retina, ciliary
body, and iris as rather large, black granules. Lipofuscin
The apoptotic bodies are membrane-encapsulated,
thus preventing exposure of cellular contents to
occurs in aged cells and in the RPE and may be difficult to
the extracellular space and possible inflammatory identify by conventional light microscopy, but by electron
reaction. microscopy it differs considerably in structure and density
from melanin.
III. Hemosiderin results from intraocular hemorrhage when
C. Apoptosis appears to play a major role in regulating cell hemoglobin is oxidized to hemosiderin.
populations. A. It occurs as an orange–brown pigment in macrophages
D. Defective apoptosis may play a role in the genesis of and, when plentiful in the eye, is called hemosiderosis
cancer, AIDS, autoimmune diseases, degenerative and bulbi.
dystrophic diseases of the central nervous system (includ- B. Systemic hemochromatosis (see Chapter 6) consists of
ing the neural retina), and diabetic retinopathy. portal cirrhosis and elevated iron content in parenchymal
E. Apoptosis is significant in the pathobiology of such cells of multiple organs. When increased amounts of iron
ocular disorders as glaucoma, cataracts, ocular tumors, are deposited in tissues of multiple organs but cirrhosis
macular degeneration, and diabetic retinopathy. and its complications are lacking, systemic hemosiderosis
F. Although apoptosis frequently is viewed as a pathologic is present.
process, it is vital to successful ocular embryologic C. The distribution of iron in the eye differs in local ocular
development. disease (hemosiderosis bulbi and siderosis bulbi) and
systemic disease (Table 1.12).
Calcification
I. Dystrophic (degenerative) calcification: This occurs when
calcium is deposited in dead or dying tissue (e.g., in
long-standing cataracts, in band keratopathy, and in
retinoblastoma). TABLE 1.12 Deposition of Iron in the Eye
II. Metastatic calcification: This type of calcification occurs of Local* and Systemic (Hemo) Siderosis
when calcium is deposited in previously undamaged tissue Local Siderosis
(e.g., in the cornea of people with high serum calcium and Systemic
levels [hyperparathyroidism and vitamin D intoxication], Tissue Hemosiderosis Hemochromatosis
where it shows as a horizontal band, and in the sclera, Corneal epithelium Yes No
where it shows as a senile plaque). Trabecular meshwork Yes No
Iris epithelium Yes No
Iris dilator and Yes No
An unusual cause of metastatic calcification is Werner’s sphincter muscles
syndrome, a heredofamilial disorder characterized by Ciliary epithelium Yes Yes
premature graying and baldness, short stature, gracile Lens epithelium Yes No
build, and “bird face.” Ocular findings include blue Vitreous body Yes No
sclera, bullous keratopathy, presenile posterior sub- Sclera No Yes
capsular cataract, degenerative corneal changes post Blood vessels Yes No
cataract surgery, retinitis pigmentosa–like features, and Sensory retina Yes No
paramacular degeneration. Retinal pigment Yes Yes
epithelium

Autolysis and Putrefaction
I. Autolysis is partly the self-digestion of cells using their own
cellular digestive enzymes contained in lysosomes (“suicide
bags”) and partly other unknown factors.
*With local iron foreign body, iron is usually deposited in all adjacent
(contiguous) tissues.
(Modified from Roth AM, Foos RY: Ocular pathologic changes in
primary hemochromatosis. Arch Ophthalmol 87:507, 1972. ©
American Medical Association.)

IV. Exogenous iron results from an intraocular iron foreign
EPIGENETICS AND OCULAR DISEASE
body. The resultant ocular iron deposition is called siderosis
bulbi (see Table 1.12).
V. Acid hematin is an artifact produced by action of acid fixa-
tives, particularly formaldehyde, on hemoglobin.
VI. Differentiation of the pigments:
A. Only acid hematin is birefringent to polarized light.
B. Only melanin bleaches with oxidizing agents, such as
hydrogen peroxide.
C. The cathepsin-D reaction is helpful in identifying
lipofuscin.
D. Only iron stains positively with the common stains
for iron.
Hemosiderin and exogenous iron cannot be differ-
entiated on their staining properties and sometimes
may not be differentiated on structural grounds.
Growth and Aging
I. In general, ocular tissue in infants and young people is
quite cellular. Cellularity decreases with aging as the col-
lagenization of tissues increases.
II. The eye is at least two-thirds of its adult size at birth, and
it usually reaches full size by the end of the second decade
of life.
Although the eyeball reaches full size, the lens, an
inverted epithelial structure, continues to grow throughout
life. Nuclear cataract results from the increased density
of the central (nuclear) lens cells (and other factors) and
can be considered an aging change.
III. Certain chemicals may be deposited in ocular tissues during
the aging process, including calcium in the insertion of
the rectus muscles (senile plaque; Fig. 1.33) and in Bruch’s
membrane (calcification of Bruch’s membrane), and sorbitol
in the lens.
IV. The important growth and aging changes of individual
tissues are taken up in the appropriate sections in the remain-
ing chapters.
A B
Fig. 1.33 A, Scleral calcium plaques present where horizontal rectus muscles insert. Plaques appear trans-
lucent gray. B, Calcium deposited through full thickness of sclera in region of insertion of rectus muscles.
Epigenetics and Ocular Disease 33
I. Throughout Ocular Pathology we will provide information
regarding the role of classic genetic alterations in the patho-
genesis of many ocular disorders. Nevertheless, epigenetic
mechanisms increasingly are being demonstrated to play
a role in ocular disease processes.
II. Epigenetics involves the mitotically and meiotically heritable
potential for gene expression that does not involve variation
in the DNA sequence.
III. Chromatin consists of a DNA-protein-RNA complex.
IV. Epigenetics involves chemical reactions that modulate chro-
matin accessibility, regulate gene expression, and the envi-
ronmental factors that coordinate these chemical reactions.
V. Epigenetic mechanisms may be reversible, heritable, and
influenced by the environment.
VI. Common epigenetic mechanisms include DNA methylation,
covalent modification of histones, nucleosome remodeling,
nuclear dynamics, and chromatin interaction with regula-
tory noncoding RNAs.
A. DNA methylation
1. Catalyzed by DNA methyltransferase.
2. In most cases CpG dinucleotides (region of DNA
where cytosine nucleotide is followed by a guanine
nucleotide in the 5′ to 3′ direction) are methylated
at the cytosine resulting in 5-methyl-cytosine.
3. Methylation silences the promoter associated with
this genetic region. Conversely, hypomethylation leads
to increased gene expression.
4. Epigenetic mechanisms may have a pathogenic role
in ocular diseases such as corneal dystrophy, cata-
ract, glaucoma, diabetic retinopathy, ocular neoplasia,
uveitis, and age-related macular degeneration.
5. For example, in retinoblastoma the RAS-associated
domain family IA (RASSFIA) tumor suppressor gene
is silenced by DNA methylation in 82% of tumors.
B. Histone modification
1. Histones are the primary protein components of chro-
matin and condense the DNA into nucleosomes that
are said to form a “beads-on-a-string” conformation
and based on their location along the DNA helix,
the histones impact the ability of other regulatory
34 CHAPTER 1 Basic Principles of Pathology
DNA-binding proteins to access specific sites along
the DNA.
2. Can include acetylation, methylation, ubiquitination,
phosphorylation, and sumoylation.
a. Writers, readers, and erasers are involved in adding,
interpreting, and/or removing epigenetic modi-
fications on chromatin.
b. Acetylation is a very common histone modifica-
tion strategy.
1) It opens up the chromatin structure allowing
recruitment and binding of the transcription
factor and RNA polymerase II.
3. Methylation may facilitate or depress gene expression
depending upon the target site.
4. Occur primarily within the N-terminal tails of histones
protruding from the surface of the nucleosome or
on its core region.
C. Noncoding RNA includes long noncoding RNA,
microRNA, short interference RNA, and Piwi-interacting
RNA. Only long noncoding RNA and microRNA will
be discussed.
1. Long noncoding RNA (LncRNA)
a. RNA transcripts longer than 200 nucleotides and
structurally resembling RNA but have little or no
protein coding potential.
b. Most are located in the nucleus.
c. Termed sense or antisense if they overlap with
protein coding genes.
d. Intronic long noncoding RNA (lincRNA) are genes
derived from intron of protein coding genes.
e. Circular RNA (circRNA) are longer than 200
nucleotides.
f. May impact DNA transcription through multiple
mechanisms.
g. May be involved in glaucoma, proliferative vit-
reoretinopathy, diabetic retinopathy and ocular
tumors.
2. Small noncoding RNAs (micro RNAs), which are
about 22 nucleotides in length, bind to untranslated
regions of mRNA and cleave mRNA resulting in
decreased protein synthesis and gene expression.
D. Chromosome remodeling proteins modulate chroma-
tin compaction and DNA accessibility to proteins in
a non-covalent manner. They modulate DNA-histone
interactions.
MODERN MOLECULAR PATHOLOGY
DIAGNOSTIC TECHNIQUES
The scope of molecular pathology is expanding at a rapid rate.
Only a few of the most common techniques can be highlighted
here.
I. Fluorescence in situ hybridization (FISH) (Fig. 1.34).
A. Commonly employed to determine the presence of a
specific mutation, or a particular chromosomal presence,
absence or rearrangement.
B. Targets DNA in cells, nuclei or chromosomes.
C. Uses a DNA probe that is labeled with a nucleotide that
is either conjugated to fluorescein (direct labeling) and/
or a non-fluorescent hapten (indirect labeling).
1. If a non-fluorescent hapten is used, the detection
depends on the addition of a fluorescence-coupled
anti-hapten reporter molecule.
D. The probe binds with the specific complementary region
on the DNA under investigation.
E. Three basic types of probes are utilized:
1. Chromosome painting probes that contain multiple
DNA fragments that are complementary with regions
along the entire length of the chromosome resulting
in fluorescence of the entire chromosome.
2. Repeat sequence probes are used to detect the gain
or loss of specific chromosomes.
a. They usually employ chromosome-specific peri-
centromeric and/or subtelomeric probes.
3. Unique sequence probes usually are employed to
identify a gene or fragment related to a specific disease.
a. Subtelomeric probes are used to identify subtelo-
meric deletions or rearrangements.
F. One use for FISH in ophthalmic oncology has been in
the cytogenetic identification of abnormalities, such as
those involving chromosomes 3, 4, 6, 8, 15 and 18 in
uveal melanoma.
1. Other cytogenetic techniques, such as comparative
genomic hybridization (CGH) and spectral karyotyp-
ing (SKY) also may be employed for these purposes.
G. It can be performed rapidly (1–2 days) and can be per-
formed on both dividing and static cells (i.e., cells in
metaphase or interphase).
H. There are numerous variations on the basic FISH tech-
nique that will not be discussed here.
II. Flow cytometry (Fig. 1.35).
A. The flow cytometer evaluates characteristics of individual
cells as they flow in fluid through a laser beam.
1. The cells usually are labeled with a fluorescently con-
jugated monoclonal antibody.
2. Fluorescence in the laser beam occurs in response to
a specific wavelength of light stimulation, and the
intensity of the fluorescence is measured.
3. By using different fluorochromes for different anti-
bodies, flow cytometers may be capable of 5- to
10-color immunophenotypic analysis.
B. Multicolor flow cytometry is often utilized in evaluating
vitreous samples, particularly for the detection and
immunotyping of lymphoma.
1. In one report, it has an 82.4% sensitivity and 100%
specificity.
C. It has been used to characterize vitreous inflammatory
cells.
III. Polymerase chain reaction (PCR) (Fig. 1.36)
A. Tremendously amplifies very small quantities of DNA so
that they can be characterized for diagnostic purposes.
B. Fundamentally, a three-step process that requires:
1. A DNA template that contains the target DNA
region.
Another random document with
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tungettelevilta suojelijoilta, ja löytääkseni halvan huoneen, suuntasin
nyt kulkuni Cadem-nimistä kaupunginosaa kohti, pieniä katuja pitkin,
jotka usein kulkevat kuin tunnelit ainakin talojen läpi. Kreikkalainen
majatalon isäntä vastasi minulle, että jos halusin nukkua halvalla, oli
minun asuttava toisen kanssa. Suostuin tähän. Nähtyäni huoneen ja
jätettyäni mytyn kainalostani, minä tiedustelin, kuka nukkuisi toisessa
vuoteessa.

»Mies samaa maata kuin sinäkin», vastasi kreikkalainen töykeästi.

Tuska ahdisti rintaani. Maani, Kira, äiti, häipyivät kauas sumuun,
kadoten saavuttamattomiin. Ja minä, joka olin kiskaistu juurineni
omasta maaperästäni, mitä etsin minä tästä synkästä kaupungista?
Millä keinoin luulin vielä voivani löytää sisareni? Ja kuinka ansaitsisin
elatukseni, kun rahani olisivat lopussa?

Lisäksi olin ilman passia. Siinä toinen pulma. Minut voitaisiin

pidättää. Ja kuka auttaisi minut vankilasta? Majatalon pihalla istui
miehiä turkkilaiseen tapaan kukkien reunustaman suihkulähteen
ympärillä, poltellen, juoden maidonvalkeaa viinaa ja näyttäen
onnellisilta. Nuo ihmiset olivat kotonaan. He tunsivat toisensa,
auttoivat toinen toistaan, jakoivat keskenään ilot ja surut. Entä minä?
Mitä minä olin heille? Tuntematon. Kukapa tulisi huoneeseen, missä
tuntematon kuolee (kuolee sairauteen tai suruun), kysymään
häneltä, kaipaako hänen sielunsa jotakin?

Vaistomaisesti tunnustelin kemiriäni, missä minulla oli rahani,

ainoa ystäväni. Mutta raha on ystävä, joka jättää meidät petollisesti,
ilman kaipausta ja surua, enkä tietänyt, millä tavoin sen jälleen saisi
palaamaan kemiriin. Toista oli Kira! Hän ei olisi jättänyt minua
mistään hinnasta maailmassa. Me olimme eroittamattomat. Oliko
toista sellaista Kiraa kaikkien näiden ihmisten joukossa, jotka
täyttivät kaupungit ja kylät? Ehkä. Mutta heillä oli omat Dragomirinsa,
ja heille minä olin vain muukalainen, joka kulkee ohi, johon luodaan
utelias katse, ja joka unohdetaan.

Lohdutuksekseni tilasin lasin viinaa, sitten vielä toisen. Tuli

päivällisaika. Söin jotakin ja join lasin viiniä, sitten vielä toisen. Ja
sydän epävarmana nousin huoneeseeni viimein.

Noin kolmekymmenvuotias mies istui siellä puolipukeissa

vuoteensa laidalla. Pöydällä paloi öljylamppu. Vuoteet siisteyteen
nähden epäilyttäviä. Sumea peili. Ei mitään pesutelinettä.

Toivotin kreikankielellä hyvää iltaa ja tarkastelin vuodettani.

»Se on siirrettävä hiukan etemmäs seinästä», sanoi mies minulle

kuin vanhalle tutulle. »Täällä on lutikoita. Ja lamppu on jätettävä
palamaan yöksi, sillä lutikat pelkäävät valoa, kuten pöllötkin».

»Lutikat?» sanoin kysyvästi, sillä en lainkaan tietänyt, millaisia

eläviä ne olivat. »Mitä ne ovat?»

»Etkö tiedä, mitä lutikat ovat? No niin, saat nähdä tänä yönä.
Mutta sanohan, missä olet nukkunut tähän asti, kun et niitä tunne.
Minä puolestani en tiedä, millainen on vuode ilman lutikoita!»

»Purevatko lutikat?» tiedustelin minä, tätä uutta vihollista peläten.

»Hiukan», vastasi hän välinpitämättömästi.

Olin väsynyt ja halusin riisuutua ja panna maata, mutta kumma

kainous esti minua tekemästä sitä tuon muukalaisen nähden. Hän
huomasi sen, sillä hän meni pienentämään lamppua. Kun olin puoli
pimeässä pujahtanut peiton alle, nousi hän jälleen ja väänsi valon
suuremmaksi.

»Luulisipa, että olet nuori tyttö!» sanoi hän nauraen. Tämä

ystävällisyys herätti minussa luottamusta, ja minä nukahdin tuona
iltana melko tyytyväisenä, kemiri pääni alla.

Seuraavana aamuna en tietänyt enempää kuin edellisenä

iltanakaan, mitä lutikan purema oli, mutta toverini osoitti veritahraa
tyynylläni. Melkein iloisena minä pukeuduin rivakasti hänen
nähtensä.

Pihalta kuului puhetta ja naurun remakkaa. Katsoin ulos ikkunasta

ja näin suihkukaivon ympärillä joukon miehiä, jotka imivät suuria
tshibukeja ja hörppivät kahvia. Piha oli kostutettu ja lakaistu. Raikas
ilma tunki keuhkoihini, kaiken ja kaikkien yllä väikkyi kellertävä,
salaperäinen, vain itämaille ominainen valo.

Panin kapineeni kokoon. Sydämessäni uinuva lempeä vihollinen

heräsi:

»Haluatteko juoda kahvia kanssani?» sanoin tuntemattomalle.

Alhaalla me pakinoimme, tuprutellen tshibukeistamme valtavia

savupilviä. Hän uskoi minulle ensinnä huolensa, kertoen olevansa
ilman rahaa, työtön ja velkaantunut. Silloin minäkin ilmaisin hänelle
suruni ja sanoin:

»Minä olen hukannut paperini. Jos voitte sanoa minulle, kuinka

saan uudet tilalle, annan teille vaivoistanne turkkilaisen
kultamarkan».

Hänen kasvonsa kirkastuivat.

 »Se käy kyllä päinsä!» sanoi hän hiljaa. »Täällä on »yleinen
kirjuri», joka hankkii sellaisia, mutta hän vaatii paljon rahaa».

»Kuinka paljon?» huudahdin minä onnellisena.

»Neljä kultamarkkaa».

»Minä maksan ne! Ja te saatte lupaamani palkkion».

Tuntia myöhemmin kookas valkopartainen »kirjuri» vannoi päänsä

kautta viranomaisen edessä nähneensä minun syntyvän
Stambulissa sinä ja sinä armon vuonna, että nimeni oli »Stavro», ja
että siis olin »sulttaanin meidän Herramme, nöyrä alamainen».

Virkamies kuunteli hymyillen, tarttui sitten kynään, kirjoitti pitkän

sepustuksen kauniilla arabialaisella käsialalla, pani alle nimensä,
antoi vanhuksen tehdä samoin, painoi siihen hallitsijan sinetin ja
ojensi minulle tuon kallisarvoisen taikakalun.

»Hänelle on annettava juomarahaa», kuiskasi kirjuri korvaani.

Panin pöydälle kultamarkan.

»Se ei riitä», sanoi vanhus. Lisäsin vielä toisen, kääntyen aina

nurkkaan päin kemiriäni kaivelemaan. Ulkona maksoin tuon
syntymäni väärän todistajan. Jäätyäni yksin toverini kanssa, me
kiertelimme pitkin kaupunkia, söimme ja joimme.

Humaltuneina ja iloisina me molemmat palasimme illalla

asuntoomme, missä minä, vähät välittämättä lutikoista, nukahdin
kuin tukki, piilotettuani kuitenkin ensin kemirini pääni alle.

Herätessäni hämmästyin kovin huomatessani, että olin yksin

huoneessa. Mutta hämmästys muuttui muuksi, kun havaitsin
kemirini, tuon petollisen ja sydämettömän ystävän, myöskin
hylänneen minut, jättäen taskuuni vain kolme megdediä [1 megdedi
= 4 frangia 20 cent.] ja tuon kirotun taikakalun…

*****

Nyt ei enää ollut kysymys itkusta! Nyt oli edessä kuolema…

Vielä tänä päivänä tunnen rinnassani saman puristuksen, saman

tyhjyyden, joka tuona aamuna oli tappaa minut.

Paitahihasillani minä kuljeskelin huoneessa sinne tänne ja,

tietämättä itsekään syytä, kumarruin katsomaan ulos ikkunasta.
Samat ihmiset kuin eilenkin istuivat tupakoiden suihkukaivon
ympärillä. He näyttivät mielestäni ruumisarkkua vartioivilta
haudankaivajilta. Ja minä astuin koneellisesti eteenpäin, luullen
laskeutuvina alas portaita, ja syöksyin tyhjyyteen. Nousin heti
jaloilleni, mutta kasvoni vuotivat verta ja olin tukehtua. Kun isäntä
vieraineen juoksi luokseni, sain vain sanotuksi:

»Ke — ke — miri…»

Minulta kyseltiin yhteen ääneen jos jotakin. En yhä vieläkään

voinut sanoa muuta kuin: »Kemiri».

»Mitähän hän tarkoittanee kemirillään?»

He valelivat päätäni kylmällä vedellä, pesivät veren kasvoistani ja

pakoittivat minut nielemään hiukan viinaa.

»Puhu nyt suusi puhtaaksi!» huusi isäntä, ravistaen minua

olkapäästä.
 »Kemiri», vaikeroin minä lakkaamatta.

»Luulenpa, että toisessa vuoteessa nukkunut veijari on kiitokseksi

eilisestä kestitsemisestä varastanut hänen kemirinsä», päätteli hän.

Ja kun tahdoin kaiken aikaa vain nousta ja lähteä, pakoitti hän

minut istumaan tuolille, mihin lyyhistyin tuskani ristiinnaulitsemana,
käsivarret riipuksissa. Hän koetti lohduttaa minua sanomalla:

»No niin, sinua on kohdannut onnettomuus… Rahasi on

varastettu. Mutta sellaisen vuoksi ei maksa heittää henkeään! Sillä ei
pääse pitkälle… Paljonko sinulla oli rahaa?»

»Kemiri», toistelin minä yhä.

»Sepä se! Poika ei enää osaa sanoa muuta kuin »kemiri».

Tämän sanottuaan isäntä nousi huoneeseeni ja toi vaatteeni.

»Pue nyt päällesi!» sanoi hän.

Ikäänkuin olisin ollut halpautunut, minä annoin hänen pukea itseni

kiireestä kantapäähän. Tutkiessaan taskujani hän löysi
syntymätodistukseni ja rahani.

»Kas vaan!» huudahti hän, »etpä olekaan niin köyhä! Sinullahan

on kolme megdediä… Ja nimesi on Stavro. No niin, Stavro, kun on
noin paljon rahaa, ei kuole nälkään. Mitä työtä osaat tehdä?»
»Kemiri…»

»Kirottu kemiri!» kiljasi hän suuttuneena. Ja pannen kaikki jälleen

taskuihin! hän poistui, sanoen:
 »Kun kaikki käy ympäri, ei sinulla ollut tuossa kemirissä edes
kamelin hintaa, sillä silloin et olisi etsinyt yösijaa minun luotani».

Minulla oli kemirissäni paljon enemmän kuin kamelin hinta. Minulla

oli kahdeksankymmentäkolme turkkilaista kultamarkkaa, yhdeksän
jalokivisormusta ja kello! Ja tällainen omaisuus hallussani olin
todellakin etsinyt yösijaa hänen luotaan!…

Ei pidä lainkaan paikkaansa se väite, että inhimillinen olento

ymmärtää elämää. Hänen järkensä ei paljoonkaan pysty, eikä hänen
puhelahjansa tee häntä vähemmän typeräksi. Mutta kun on
kysymyksessä lähimmäisen onnettomuuden arvaaminen ja
ymmärtäminen, voittavat he tyhmyydessä eläimetkin.

Usein näemme kadulla miehen, jonka kasvot ovat kelmeät ja katse

epätoivoinen, tai vaikkapa itkevän naisen. Jos olisimme ylevämpiä
olentoja, pitäisi meidän pysähdyttää tuo mies tai tuo nainen ja tarjota
heille pikaista apua. Tällainen ylevyys kohottaisi minun mielestäni
ihmisen eläintä korkeammalle. Sellaista ei ole!

En enää tarkoin muista, — sillä siitä on jo viisikymmentä vuotta, —

kuinka nousin tuolilta ja lähdin majatalon pihasta, jonka poikki kuljin
ajatuksettomana, ja jatkoin matkaani läpi koko kaupungin. Mutta
tiedän vain, ettei ainoakaan käsi laskeutunut tämän synkkäkatseisen
nuorukaisen olkapäälle, joka kulki koneellisesti pitkin katuja; ei
ainoakaan ääni, eivät ainoatkaan ihmiskasvot ilmaisseet
mielenkiintoa häntä kohtaan. Ja tällaisessa tiedottomuuden tilassa
jouduin tuona kauniina huhtikuun aamuna Baptuma-nimiselle
puistokadulle Damaskuksessa.

Minut palauttivat tajuihini arabialaisen kuskin raivokkaat kiroukset,

hän kun oli vähällä ajaa ylitseni. Tunnustelin vyötäni, jossa ei enää
ollut kemiriä.

Sydämeni vapisi kuin pyydystetty lintu, ja kurkkuuni nousi pala,

joka salpasi henkeäni. Tämä käden liike synnytti minussa
hermoväristyksen. Joka kerta kun kosketin vyötäni, säpsähti
sydämeni, ja minun täytyi lakkaamatta vakuuttaa itselleni, että minua
oli todellakin kohdannut ilkityö, ja ettei minulla enää ollut kemiriäni.
Kun suuret surut kohtaavat herkkää sydäntä, on sen vaikea tottua
siihen ajatukseen, että onnettomuus on tapahtunut ja ettei mitään
enää ole tehtävissä.

Ohitseni kulki kansaa: onnellisia pareja, naisia lapsineen,

rauhallisia ja tyytyväisennäköisiä miehiä. He katsoivat minua
kasvoihin ja kulkivat ohi. He eivät nähneet mitään. He eivät
ymmärtäneet mitään. Ja minä olin kuolemaisillani. Minun oli yksin
kannettava onnettomuus, joka oli ylivoimainen iälleni, sydämelleni ja
kokemattomuudelleni.

Kuljin yhä eteenpäin. Tulin ulos metsästä. Syyrialainen maaseutu

tomuisine teineen ja beduiinihökkeleineen oli mielestäni eloton kuin
oma ruumiini. Katseeni ei löytänyt mitään kiinnekohtaa, ja kun käteni
sattui koskettamaan vyötäni, toistivat aivoni jälleen: »Minulla ei ole
enää kemiriä…» Ja taas puristi tuska kurkkuani.

Arabialainen lapsi ajoi hitaasti ohitseni istuen kahareisin aasin

selässä ja taluttaen nuorasta kamelia, jolla oli selässään kaksi
keinuvaa kantamusta. Tämä ruma eläin litteine käärmeensilmineen
peloitti minua. Etäämpänä tuli vastaani mustapartainen,
villinnäköinen beduiini, joka ajoi täyttä laukkaa ja pysähtyi kysymään
minulta jotakin arabiankielellä. En osannut vastata hänelle mitään.
Hän katosi näkyvistäni, jättäen jälkeensä miellyttävän vaikutuksen,
sillä hänen kaunis päänsä johti mieleeni Cosman.
 Pian saavuin kylään, missä asumukset olivat perin alkeellisia ja
missä miehet maassa istuen sorvasivat puita, käytellen paljaita
jalkojaan yhtä taitavasti kuin käsiäänkin. Mustiin kaapuihin puetut,
likaiset, hunnutetut naiset, — todellisia variksenpelättimiä, —
kantoivat päänsä päällä soikeita savimaljoja, ja tahraiset ja laihat
lapset kirkuivat leikkiessään kuin pienet paholaiset. Eräs mies otti
parhaillaan pieniä litteitä leipiä savesta kyhätystä uunista, joka oli
puolittain maan sisässä. Tunsin nenässäni paistetun leivän hajun.

Kylästä poistuessani huomasin koiran seuraavan uskollisesti

kintereilläni. Minä pysähdyin. Se pysähtyi, ja me katsoimme
toisiamme silmiin. Koira oli väriltään tummanharmaa, Suden
kokoinen, mutta tuolta raukalta puuttui Suden arvokkuus, ylpeä
käytös ja tyyneys. Se painoi nöyränä päänsä alas ja lyyhistyi
maahan pelosta. Sen silmien ilme oli epävarma, nöyrä, ja levoton.
Säälin sitä ja silitin sen päätä. Se nuoli kättäni. Sen kanssa ei ollut
vaikea tulla toimeen.

Palasin uunin luo ja ostin kahdella metelikiliä neljä pientä litteää

leipää. Se ahmaisi kaikki neljä pureskelematta. Ostin vielä toiset
neljä, panin ne taskuuni ja aloin jatkaa päämäärätöntä kulkuani. Se
seurasi minua kuten ennenkin.

Pieni, hiekkainen, aivan hedelmätön ja autio vuorenkukkula kohosi

edessäni. Pian saavuin sen luo ja aloin nousta rinnettä. Mutta
hengästyin pian ja istuuduin, koira vierelläni. Edessäni lepäävä
Damaskus, jonka kupoolit ja minareetit kohosivat laajojen
pengermien yli, näytti minusta suunnattomalta, valkean tomun
kattamalta hautausmaalta.

Luokseni ei kantautunut pienintäkään melua. Päässäni jyskyttivät

haavoitetun sydämeni kiihkeät lyönnit. Silmäni sumenivat. Damaskus
ja maailma häipyivät pois. Menneisyyteen katsellen minä näin
päivänselvänä lapsuuteni kodin. Tuon etäisen ajan suloinen elämä
levittäytyi suljettujen silmieni eteen. Elin uudelleen kaikki menneet
onnelliset päivät yksityiskohtiaan myöten, lapsuuteni hämärimmistä
muistoista alkaen aina kaameaan murhayöhön ja ryöstöön saakka.

Ja yhtäkkiä välähti aivoissani ajatus, että Kira ja minä saimme

onnettomuudellamme sovittaa sen, että olimme yllyttäneet rikokseen
ja olleet avullisina sen toimeenpanossa. Olimme toivoneet isän ja
veljen kuolemaa. Se oli kuolemansynti. Nyt Jumala rankaisi meitä,
Kiraa orjuudella, minua tuhoisalla vapaudella…

Kun avasin silmäni, niin kauhistuin. Taivas oli auringonlaskun

puolella veripunainen. Alhaalla riippuvat, maksoittuneen veren
väriset pilvet liikehtivät hitaasti, saaden mitä kummallisimpia, toinen
toistaan peloittavampia muotoja.

Pienen luolan edustalla, missä olin istua kyyröttänyt, minä

heittäydyin maahan, kätkien kasvot käsiini. Rukoilin kauan, pyytäen
anteeksi Jumalalta, isältäni, murhatun veljeni sielulta.

Ja yön varjot lankesivat katuvan nuorukaisen yli, jolla oli

onnettomuustoverinaan koira, jonka sattuma oli tuonut hänen
tielleen.

Rukous ja katumus tuo huojennusta uskoville sieluille. Sain rauhan

muutamaksi tunniksi. Mutta aamunkoitteessa on hiekka-aavikoilla
jäätävän kylmä. Kun aurinko kohosi Levantin taivaalle, puistatti
ankara vilu ruumistani, niin että pelkäsin kylmettymisestä saavani
kuoleman taudin. Sanoin itselleni: »Jos kuolen katuvaisena, antaa
Jumala minulle anteeksi eikä sieluni joudu iäiseen kadotukseen».
 Nousin ja lähdin paluumatkalle. Tiellä söin yhden leivistä ja annoin
muut kolme koiralle, joka oli nälkiintyneempi kuin minä.

Eikä aikaakaan, kun aurinko jo alkoi lämmittää selkääni, ja minä

tunsin hyväätekevän rauhan laskeutuvan rintaani. Saavuin kylään.
Se näytti minusta vähemmän rumalta. Täällä koira jätti minut. Tunsin
siitä ikävää, ja silitettyäni sen päätä, erosin siitä kuin ainakin
miellyttävästä seurasta, johon on tutustuttu lyhyellä matkalla.

Olin jälleen yksin, ja kadonneen kemirin muiston yhä ahdistaessa

mieltäni aloin taivaltaa Baptuman ja Damaskuksen metsäistä tietä.
Vastaani tuli pitkä kamelikaravaani, minua lainkaan peloittamatta.
Saavuin Baptuman puistokadulle hiukan ennen puoltapäivää
säteilevän sään vallitessa. Liikenteen vilkkaus hämmästytti minua.
Miehiä kauniissa turkkilaisessa asussa, nuoria ja viehättäviä naisia
— useimmilla vain kasvojen alaosa kevyen, valkean harson
peitossa, — liikehti ajaen tai jalkaisin puoleen ja toiseen. Kaikkialta
kuului äänekkäitä huudahduksia, iloista puheensorinaa, kaikkialla
helisi nauru, kuin olisi taikasauvalla kosketeltu kristallilaseihin.
Sointuvat äänet ja koristeelliset puvut hurmasivat minut. Muistin, että
oli perjantai, turkkilaisten sabatti. Naiset tervehtivät toisiaan sirosti ja
hillitysti, mutta miesten tunteenpurkaukset, turkkilaiset tervehdykset
ja loputtomat kädenpuristukset pakoittivat jalkamiehet usein
pysähtymään pitkäksi aikaa. Puhuttiin paljon turkinkieltä, vaikka
arabia olikin hallitsevana.

Seisoin kauan ja ihailin edestakaisin liikuskelevaa joukkoa. Sitten

vaunut ja ajopelit harvenivat. Minä jatkoin matkaani miettiväisenä,
rauhattomana, sillä sydämessäni taistelivat keskenään elämänhalu,
ilon kaipuu, onnettomuuteni ja häviöni. Pian olin yksin, yksin
suruineni. Vastaani ajoivat parihevosten vetämät kauniit vaunut.
Juuri kun ne olivat sivuuttamaisillaan minut, salpautui hengitykseni,
sydämeni lakkasi lyömästä.

Vaunuissa istui Kira…

Niin, vielä tällä hetkelläkin uskon, että se oli suloinen, rakastettu

sisareni. Se oli Kira sellaisena, jollaiseksi Nazim Effendi oli
purjealuksessaan hänet koristanut, odaliskin loisteliaassa puvussa,
muistuttaen kajuutan seinillä riippuvia kuvia.

Minä horjahdin, löin käteni yhteen ja huusin romaniankielellä:

»Kira! Kiralina!… Se olen minä, Dragomir!»

Nuori nainen hymyili läpinäkyvän huntunsa alla ja tervehti minua

hansikoidulla kädellään, mutta ajaja läimäytti piiskaansa ja hänen
vieressään istuva eunukki loi minuun salamoivan katseen, ja sitten
hevoset kiitivät kuin lentäen tiehensä.

Luulin kuolevani… Niin, se oli Kira, hän oli antanut minulle merkin!
Ja muitta mutkitta minä aloin harpata vaunujen perässä kuin strutsi,
sanoen itsekseni:

»Kaikkivaltias Jumala! Tuskin olen tunnustanut syntini ja katunut,

kun sinä armossasi jo lähetät luokseni kadonneen sisareni!…»

Ponnistuksistani huolimatta loittonivat vaunut silminnähtävästi.

Olin jo aivan hengästynyt ja pelkäsin kadottavani ne näkyvistä.
Onneksi näin niiden metsän loputtua suuntautuvan suoraan komeaa
huvilaa kohti, jonka suuri portti avautui selkoselälleen, niellen
ajoneuvot ja sulkeutuen jälleen.
 Huudahdin ilosta. Viimeiset voimani ponnistaen syöksyin porttia
vasten, ryskyttäen sitä raivokkaasti sekä käsin että jaloin. Heti
avautui ison portin vieressä oleva pienempi portti ja univormuun
puettu palvelija tuli esiin.

»Kira!» huusin minä hengästyneenä turkinkielellä, »sisareni…

tahdon puhutella…»

»Kuinka? Mitä tahdot?» kysyi palvelija samalla kielellä,

pysähdyttäen minut.

»Nainen… joka ajoi tänne vaunuissa… on… sisareni… Kira».

»Kira? Sinä olet hullu».

Olin todellakin hullu, sillä syöksyin palvelijan kimppuun ja työnnyin

hänen ohitseen pihaan. Mutta pitkälle en päässyt, ennenkuin
edessäni seisoi kaksi miestä, ja kuulin käheän, vanhan äänen
huutavan ikkunasta:

»Mitä tämä melu merkitsee? Pieskää hiukan tuota ghiauria

[turkinkieltä ja merkitsee kristitty], ja palvelijaa, joka päästi hänet
sisään!»

Minut laahattiin pois pihasta, heitettiin maahan, ja minua piestiin

nahkahihnalla, kunnes housuni ja pakarani olivat riekaleina. Sitten
pyövelit salpasivat portin, jättäen minut siihen makaamaan kivusta
puolikuolleena.

*****

Olen saapunut Golgatani huipulle. Tähän päättyvät myrskyisen

lapsuuteni kolme surullisinta vuotta… Sillä jos Jumala olikin ankara
minua kohtaan kieltäessään minulta Kiran, oli sittekin olemassa
Kaitselmus. Tämä Kaitselmus lähetti minulle ystävän.

Ponnistaen haavoitetun ruumiini viimeiset voimat, laahauduin

hädin tuskin tien toiselle puolen, missä vaivuin uupuneena maahan.
Tällöin minua lähestyi neljän- ja viidenkymmenen vuoden välillä
oleva, köyhästi puettu mies, kantaen toisessa kädessään salepin
valmistamiseen tarvittavia aineksia, toisessa koria kuppeineen, laski
kapineensa maahan, ja ristien käsivartensa huudahti kreikankielellä:

»Voi, poika-parka! Katselin voimattomana, kuinka sinua piestiin.

Miten lienetkään loukannut noita pakanoita, että he sinua noin
pahoinpitelivät?»

Katselin hänen vilpittömiä kasvojaan, pehmeää, harmahtavaa

partaansa, hyviä, surunvoittoisia silmiään uurteisen otsan alla, ja
huusin raivostuneena, nousten omia tunteitani vastaan:

»Menkää helvettiin ja jättäkää minut rauhaan!»

Ja purskahdin itkuun. Hänen hyvyytensä tulvahti yli äyräitten:

»Miksi toivotat minut helvettiin, lapseni?» sanoi hän. »Säälin sinua

sydämestäni ja tahtoisin auttaa sinua onnettomuudessasi».

»Jättäkää minut rauhaan, te ihmiset, säälinenne ja sydäminenne!

Olen saanut maistaa niitä kylliksi… Tahdon kuolla yksin!…»

»Oi, sinua onnetonta! Niin nuori ja jo elämään kyllästynyt! Mutta

juo kuitenkin tämä lämmin juoma. Se virkistää sinua hiukan».

Otin vastaan salep-kupin, tietämättä, mitä ajatella. Mikä sääntö,

mikä johtopäätös oli sovellutettava tähän lyhyeen kokemukseen, kun
niin monet ihmiset, jotka alussa olivat näyttäneet hyviltä ja avuliailta,
olivatkin päätyneet alhaisiksi ja rikollisiksi? Niin, kuusitoistavuotiaana
minä tunsin ihmissielun halpamaisuuden. Enkä kuitenkaan tietänyt
kaikkea.

En etenkään tietänyt, että Luojan työt ovat suunnattoman

monimutkaiset ja vaihtelevat, ja etteivät tuhannet kärsimämme
häväistykset oikeuta meitä sylkäisemään koko ihmisyyttä kasvoihin.
Jumala itsekin ymmärsi tämän, kun hän syntiseen ihmiskuntaan
vihastuneena päätti rangaista sitä, tuhoamatta sitä kuitenkaan
kokonaan, koskapa pelasti häviöstä hurskaan patriarkan ja hänen
perheensä. On totta, ettei vedenpaisumusta seurannut ihmiskunta
ollut edellistä parempi, mutta se ei ollut sen oma syy. Se johtui siitä,
että Jumala (niinkuin minä kuusitoistavuotiaana) tunsi huonosti
maailman eikä tietänyt, mitä teki.

Minä tiesin, minä, siitä päivästä lähtien, jolloin kohtalo lähetti

luokseni Barba Yanin, tuon sielultaan jumalaisen salepinmyyjän,
minä tiesin, että sen ihmisen on pidettävä itseään onnellisena, joka
on saanut tavata elämässään Barba Yanin kaltaisen olennon. Minä
olen tavannut vain yhden sellaisen. Mutta se riitti tekemään elämän
minulle siedettäväksi, saattoipa minut usein vielä siunaamaankin sitä
ja laulamaan sen ylistystä. Sillä yhden ihmisen hyvyys on
voimakkaampi kuin tuhansien pahuus. Paha työ kuolee tekijänsä
kanssa, mutta hyvyys säteilee vielä vanhurskaan poistuttuakin.

Niinkuin aurinko hajoittaa pilvet ja tuo maailmaan iloa, hävitti

Barba Yanikin sieluani kalvavan pahan ja teki sydämeni terveeksi.
En sallinut tämän tapahtua vastustelematta, mutta mikä sydän,
olkoonpa kuinka elämän murjoma tahansa, voi pitää puoliaan
kaikkivoipaa hyvyyttä vastaan.
 Minun täytyi antaa perään, ja kohtalon lähettämä salepinmyyjä sai
kuulla koko tarinani. Hänen lääkkeensä oli nopea kuin salama.

»Stavraki!» sanoi hän minulle, omaksuen varovasti väärän nimeni

ja antaen sille tuttavallisen muodon. »Ensinnäkin on sinun luovuttava
etsimästä sisartasi noin epäviisaalla tavalla. Et näy tietävän, että on
helpompi riistää hirvi tiikerin kidasta, kuin nainen haaremista. Ja jos
onnistut pitämään kurissa tämän sydämesi heikkouden, käy muu
helposti kuin tanssi. Sinulla on kolme megdediä taskussasi. Hyvä,
siinä on rahaa kyllin voidaksesi ostaa salepin valmistamisessa
tarvittavan ibrikin sekä kuppeja, toisin sanoen kojeet, jotka minulla
on tässä käsissäni ja joilla olen elättänyt itseni kaksikymmentä
vuotta. Ibrik toisella käsivarrellasi, kori toisella, sinä sitten kierrät
Barba Yanin rinnalla juhlista juhliin, markkinoilta markkinoille,
huudellen iloisesti: »Salepia! Salepia! Salepia!… Kas tässä on
salepinmyyjä!» Idän oiva maa avautuu sinulle suurena ja vapaana,
tosiaankin vapaana, sillä sanottakoonpa turkkilaisista maista mitä
tahansa, ei missään voi elää vapaampana. Mutta yhdellä ehdolla:
sinun on hävittävä, kadottava joukkoon, et saa millään tavoin
herättää huomiota, sinun on oltava kuuro ja mykkä… silloin, ja vain
silloin, voit näkymättömänä päästä kaikkialle. Hyvin teljetyt ovet eivät
avaudu väkipakolla».

Ibrik ja kori käsivarrellani minä jo seuraavana päivänä huutelin

valppaana Barba Yanin vierellä. »Salepia! Täällä myydään salepia!»
Ja nyt näin, millä tavoin petollinen ja sydämetön ystävä saadaan
palaamaan takaisin kemiriin, jonka se on jättänyt. Kolikoita sateli
joka puolelta, vapaus tuli kukkarooni, ja iltaisin tunsin, että ihminen
voi olla onnellinen, vaikka taskut eivät olekaan täynnä kultaa. Me
istuimme pengermällä vesipiippua poltellen, ja hyvyys, joka säteili
Barba Yanin koko olemuksesta, valtasi minut tykkänään. Olin hänelle
kiitollinen, rakastin häntä kuin hyvää isää ja ystävää. Asuin hänen
luonaan, tein työtä hänen rinnallaan, yhdessä me aterioimme,
yhdessä nautimme joutohetkistä, ja näin meistä tuli eroamattomat
toverit. Ennen pitkää yhdisti meidät toisiinsa luja ystävyys, mikä liitti
nuoren vesan varttuneeseen runkoon.

Barba Yani ehätti tyydyttämään uteliaisuuttani kohottaen

menneisyyttään peittävää verhoa. Tuo menneisyys ei ollut
moitteeton eikä katkeruutta vailla.

Toimiessaan opettajana eräässä pienessä kaupungissa Kreikassa

hän oli tehnyt itsensä syypääksi himorikokseen, mistä sai
rangaistukseksi kaksi vuotta vankeutta ja menetti virkansa.
Vankilasta päästyään hänen oli täytynyt lähteä kaupungista, harhailla
siellä täällä ja harjoittaa kaupustelua; hän oli kokenut
nurjamielisyyttä, saanut ystäviä, vuodattanut sydänvertaan. Toinen
rakkausseikkailu oli vähällä maksaa hänen henkensä. Silloin hän
siirtyi Vähään Aasiaan, missä vietti yksinäistä ja riippumatonta,
melkeinpä viisasta elämää.

Hän oli mies, joka osasi puhua ja vaieta, tehdä hyvää tulematta
itsetyytyväiseksi, ja häntä oli turha taivutella, milloin jokin seikka oli
vastoin hänen mieltään. Hän tunsi kaikki läheisen Idän murteet, ja
käytti joutoaikansa lukemiseen, kuljeskelemiseen ja vaatteittensa
pesuun. Hän ei pakoittanut minua mihinkään, mutta osoitti vain
minulle, mikä oli hyvää, hyödyllistä ja viisasta tehdä. Hän opetti
minut kirjoittamaan ja puhumaan kreikankieltä. Nähdessään, kuinka
uskollisesti liitin elämäni hänen elämäänsä, ei hänkään tinkinyt
ystävyydessään. Aluksi puhuttelin häntä »herraksi», mutta hän pyysi
minua sanomaan itseään nimellä »Barba». [Barba = setä;
kreikankielinen sana, jota käytetään puhuteltaessa tuttavallisesti
iäkästä miestä.] Unohdin kadonneen kemirini kalliine sisältöineen, ja
minusta tuli pian hänen oppilaansa, ainoa ystävänsä ja hänen
vanhojen päiviensä lohdutus.

Mutta sitä ennen oli minulla vielä jyrkkä rinne noustavana. Me

nousimme sen yhdessä.

Unohdin kadonneen kemirini, mutta en voinut tottua sisareni

menettämiseen. Rakastin Barba Yania, mutta Kiraa minä jumaloin.
Ja kun olin varma siitä, että hän asusti sen oven takana, missä olin
saanut selkäsaunan, neuvoi hirtehinen minua palaamaan sinne
takaisin.

Oli keskikesä, ja kolme kuukautta oli kulunut surullisesta retkestäni

Baptuman tiellä. Barba Yanin tietämättä minä kuljeskelin usein tuon
kirotun huvilan mailla, kiertelin ja kaartelin sitä kaukaa ja vakoilin
sitä. Ei mitään tulosta. Näin naisia ajavan vaunuissa, mutta Kiraa en
nähnyt.

Varovaisuuteni rohkaisemana minä päätin eräänä iltana olla

hiukan uskaliaampi. Hankin tikapuut, ja pimeän yön suosimana
asetin ne pihaa ympäröivää korkeaa kiviaitausta vasten. Tahdoin
päästä näkemään haaremin sisäpuolelle, missä tiesin naisten
liikuskelevan hunnuitta. Mutta vastassani oli kaikkialla vain
alaslasketut kierrekaihtimet. Olin itsepäinen ja aloin kiertää pitkin
aitausta, kunnes viimein näin valaistun ikkunan. Mutta näin vain ison,
upeasti sisustetun huoneen, jossa ei ollut ketään…

Jäin sykkivin sydämin odottamaan tikapuille, toivoen yhä saavani

nähdä naisväkeä.
 Yhtäkkiä tunsin tikapuiden allani liikahtavan ja olin putoamaisillani.
Pelosta jähmettyneenä tarrasin niihin kiinni parhaani mukaan, kun
äkkinäinen ja kiivas nykäisy pakoitti minut hellittämään otteeni.
Tikapuut kiskaistiin aitani ja minä putosin vartian syliin, joka sanaa
sanomatta alkoi iskeä minua nyrkeillään.

Minut sidottiin, heitettiin rattaille ja vietiin sen tien Damaskukseen,

missä minut teljettiin tutkintovankilaan.

Turkkilaiset tutkintovankilat olivat tuohon aikaan oikeita

unohduksen pesiä. Se onneton, joka joutui sinne niin raskauttavien
syiden perustalla kuin minä, ei koskaan tietänyt, milloin häntä
tutkittaisiin, ellei joku vaikutusvaltainen henkilö rientänyt lahjuksin
anomaan armoa valtaherroilta. Mutta hän ei kärsinyt niin paljon
vapautensa kadottamisesta kuin vankilaelämän kurjuudesta, etenkin
jos hän oli nuori mies.

Minun kopissani oli kymmenkunta vankia. Yhteinen,

maalaamattomista laudoista kokoonkyhätty lavitsa täytti
kolmeneljännestä huoneesta. Iso, puinen, kannella varustettu astia
eräässä huoneen nurkassa teki käymälän virkaa, levittäen
huoneeseen tukahduttavaa löyhkää. Syöpäläisiä, lutikoita ja rottia
vilisi rykmentittäin. Kukaan ei edes viitsinyt niitä tappaa, sillä olisihan
siihen tarvittu ihmisikä.

Mitä inhoittavimpia toimituksia harjoitettiin kaikkien nähden. Nämä

turkkilaiset, kreikkalaiset ja armenialaiset eivät enää olleet ihmisiä.
Inhimillinen alennustila oli täällä vertaansa vailla, sillä kaikista
luoduista olennoista vain ihminen yksin voi vajota näin syvälle.

Ja tähän maalliseen helvettiin, näiden hirviöiden keskelle, minä

jouduin. Mikä saalis minä olinkaan heille!
 Kukaan ei suojellut minua, kukaan ei puolustanut minua, enempää
muhamettilaiset kuin kristitytkään. Sensijaan he tappelivat
keskenään uudesta herkkupalasta, repivät toisiaan parrasta,
kynsivät toisensa verille. Aseistettuina he olisivat tappaneet toisensa.
Näin olin kokonaisen kuukauden alttiina mitä julmimmalle
häväistykselle…

Nyt en pahoittele, että minun oli kuljettava tuota tietä, sillä näin
opin tuntemaan ihmisolennon pohjiaan myöten. Jos säilyin hyvänä
kaikesta siitä huolimatta, mitä näin ja mitä kärsin, oli se vain siksi,
että tuottaisin kunniaa hänelle, joka on luonut Hyvyyden, luonut sen
niin harvinaiseksi ja antanut sen saada sijansa raakalaisten keskellä,
mikä on elämän ainoa oikeutus.

Olin kuin elävänä haudattu ja ajattelin kuolemaa. Kuulin

kerrottavan vangeista, jotka eivät enää jaksaneet kestää kidutusta,
vaan hirttäytyivät yöllä toisten nukkuessa nuoraan, minkä olivat
punoneet vaatteittensa riekaleista ja kiinnittäneet ikkunan
rautaristikkoon. Päätin noudattaa noiden marttyyrien esimerkkiä.

Mutta jokin sisäinen ääni kehoitti minua toivomaan. Tiesin, etten

enää ollut yksin maailmassa, kuten ennen. Sydämen mies, oiva
ystävä oli tuolla ulkopuolella. Hän oli köyhä ja vailla suojelijoita,
mutta hän oli hyvä ja viisas. Hän varmaankin ajatteli minua,
työskenteli minun vapauttamisekseni.

Olin oikeassa. Eräänä päivänä kopin ovi avautui, vartia astui

sisään ja hänen perässään Barba Yani. Mikä suunnaton ilo!… Vain
Kiran ilmestyminen olisi voinut saattaa minut yhtä onnelliseksi. Mutta
mikä suru samalla! Tänä kuukautena olivat miesparan hiukset
muuttuneet valkeiksi! Heittäydyin itkien hänen rintaansa vasten.