IY30CH19-Zychlinsky ARI 17 February 2012 13:38

ANNUAL
REVIEWS Further Neutrophil Function:
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Garret L. Hayes, Kathleen D. Metzler,
Annu. Rev. Immunol. 2012.30:459-489. Downloaded from www.annualreviews.org

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and Arturo Zychlinsky∗
Department of Cellular Microbiology, Max Planck Institute for Infection Biology,
by Harvard University on 09/10/13. For personal use only.

Charit´eplatz 1, 10117 Berlin, Germany; email: amulic@mpiib-berlin.mpg.de,
cazalet@mpiib-berlin.mpg.de, hayes@mpiib-berlin.mpg.de, metzler@mpiib-berlin.mpg.de,
zychlinsky@mpiib-berlin.mpg.de

Annu. Rev. Immunol. 2012. 30:459–89 Keywords
First published online as a Review in Advance on inflammation, antimicrobial, granule, phagocytosis, NET
January 3, 2012

The Annual Review of Immunology is online at Abstract
immunol.annualreviews.org
Neutrophils are the most abundant white blood cells in circulation,
This article’s doi:
and patients with congenital neutrophil deficiencies suffer from severe
10.1146/annurev-immunol-020711-074942
infections that are often fatal, underscoring the importance of these
Copyright  c 2012 by Annual Reviews.
cells in immune defense. In spite of neutrophils’ relevance in immunity,
All rights reserved
research on these cells has been hampered by their experimentally in-
0732-0582/12/0423-0459$20.00
tractable nature. Here, we present a survey of basic neutrophil biology,

All authors contributed equally to the work and with an emphasis on examples that highlight the function of neutrophils
are listed alphabetically.
not only as professional killers, but also as instructors of the immune
system in the context of infection and inflammatory disease. We focus
on emerging issues in the field of neutrophil biology, address questions
in this area that remain unanswered, and critically examine the experi-
mental basis for common assumptions found in neutrophil literature.

459

IY30CH19-Zychlinsky ARI 17 February 2012 13:38

INTRODUCTION early and enthusiastic evolutionary biologist in-
terested in the phagocytic capacity of cells.
In the late nineteenth century, Paul Ehrlich,
Metchnikoff demonstrated that injury of
dissatisfied with what he considered an in-
starfish embryos resulted in recruitment of
excusable disinterest in the white blood cell,
phagocytic cells to the site of injury (3). He
began to utilize newly developed cell-staining
theorized (correctly) that these cells migrate to
techniques to examine subpopulations of leuko-
injured sites and participate in microbe diges-
cytes. His experimentation led to a new appreci-
tion. Remarkably, this prescient view of neu-
ation for the heterogeneity of white blood cells
trophil action still aptly summarizes, more than
and to the discovery of several novel leukocyte
a century later, the basic role of neutrophils
subpopulations. Ehrlich named one of these
in immunity. The uniquely lobulated nucleus
newly discovered cell types, characterized by a
of the neutrophil also inspired Metchnikoff to
“polymorphous nucleus” and a tendency to re-
rename these cells: He called them polymor-
tain neutral dyes, the “neutrophil” (1) (see also
Annu. Rev. Immunol. 2012.30:459-489. Downloaded from www.annualreviews.org

phonuclear leukocytes (or PMNs), a title that
the sidebar, A Natural History of Neutrophils).
still enjoys frequent use and that is used inter-
The function of neutrophils was initially
changeably with neutrophil throughout this re-
shrouded in considerable mystery; their con-
view. Together with two other developmentally
by Harvard University on 09/10/13. For personal use only.

spicuous presence during infections led several
related cell types, the eosinophils and basophils
researchers to arrive hastily at a rather ironic
(also discovered by Ehrlich), PMNs form the
conclusion: They surmised that neutrophils
granulocyte family of white blood cells, a fam-
promote infection, serving as cellular shuttles
ily whose hallmark is the presence of “granules,”
for bacteria (2). Their actual function, that of
unique storage structures important in antimi-
antimicrobial actors in the immune response,
crobial functions (see section on Granules and
was eventually demonstrated conclusively by a
Degranulation, below).
contemporary of Ehrlich, Elie Metchnikoff, an
Neutrophils were discovered at the dawn
of the immunological sciences; consequently,
elucidation of their role in the immune re-
sponse has been an ongoing process stretching
A NATURAL HISTORY OF NEUTROPHILS over more than a century. We now know that
they are key components of the innate immune
Phagocytes are ancient cells that evolved to allow multicellular response and vital in immune function; unfor-
organisms to thrive in the face of constant competition with mi- tunately, their importance has often been over-
crobes for resources. Metchnikoff ’s seminal theory of cellular shadowed by breakthroughs in the study of the
immunity was based on comparative embryology and observa- adaptive immune response (4). Admittedly, this
tions of phagocytes in various simple organisms, including the mi- situation is exacerbated by neutrophils’ notori-
croscopic crustacean Daphnia. Remarkably, even the slime mold ous experimental intractability: They exhibit a
Dictyostelium discoideum has phagocytic cells that protect it from short life span and are terminally differentiated,
infection (200). The short-lived neutrophil with a lobulated nu- preventing growth in tissue culture. The stan-
cleus and granule-packed cytoplasm is a more recent evolutionary dard tools of molecular biology, such as trans-
adaptation. In insects, phagocytes are long lived and have round fection and RNA interference, are of little use
nuclei. They do, however, produce hydrogen peroxide and carry when applied to these cells, and immortalized
distinct classes of granules (201). Bony fish and frogs have bona “neutrophil-like” cell lines rarely reflect the
fide neutrophils that are functionally similar to mammalian ones functional diversification of neutrophils. Fur-
(202, 203). In both zebrafish and rodents, neutrophils are less thermore, neutrophil-like cells studied in the
abundant than in humans, comprising only 15–20% of immune isolation of a culture dish most certainly do not
cells. In chimpanzees, neutrophils account for more than 50% of mimic the complex biological reality in tissues
the differential blood count (204). or circulation. Conclusions from in vitro stud-
ies should, therefore, be carefully interpreted.

460 Amulic et al.

IY30CH19-Zychlinsky ARI 17 February 2012 13:38

Unfortunately, in vivo studies of neutrophil infection. Indeed, the number of neutrophils
function also raise concerns. Mouse neu- drastically increases during infection and some
trophils, the preferred model for in vivo diseases. Interestingly, neutrophils circulate
studies, differ in important aspects from their for only approximately 6–8 h and are among
human equivalents. This is perhaps best the shortest-lived cells in the human body.
exemplified by the differences in the respective Although the reason for this short life is unclear,
antimicrobial repertoires and the numbers of it may ensure neutrophil integrity; this hypoth-
PMNs in circulation (30% versus 70% in mice esis is bolstered by observations that apoptosis
and humans, respectively). prevents the release of noxious molecules.
Despite these difficulties, no picture of the Still, the question of why evolution opted for
immune response can be complete without eliminating neutrophils quickly as opposed
a comprehensive understanding of the neu- to reducing leakage of their dangerous cargo
trophil and its functions. The extensive nature remains an unanswered and intriguing mystery.
Annu. Rev. Immunol. 2012.30:459-489. Downloaded from www.annualreviews.org

of neutrophil research, however, precludes a Mature neutrophils emerge from the bone
comprehensive review of the subject matter. marrow intent on pursuing one simple, yet
In this review, we intend to provide a survey essential, question: Has host integrity been
of basic neutrophil biology and function, while compromised by potentially harmful invaders?
by Harvard University on 09/10/13. For personal use only.

emphasizing recent advances in neutrophil re- Should the answer prove to be “yes,” the
search and providing a critical assessment of neutrophil must swiftly enact a carefully
some current reports on PMN action. choreographed process to locate, attack, and
Our survey of the neutrophil begins in destroy the potential threat. At its disposal is
adult bone marrow where, under the in- an impressive arsenal of antimicrobial weapons
struction of growth factors and cytokines, that are deadly, indiscriminate, and brutish in
pluripotent hematopoietic cells differentiate their application. Although effective in their
into myeloblasts, a developmental cell type destructive capacity, these weapons can prove
committed to becoming granulocytes. As these to be just as dangerous to the host cells as to
precursor cells mature to neutrophils, they syn- their intended targets, the microbial invaders.
thesize proteins that are sorted into different Therefore, their deployment must be executed
granules (5). Traditionally, granules have been with exquisite precision and timing, at locations
subdivided into three different classes based where they are both contained and effective.
on their resident cargo molecules: azurophilic, How then does the neutrophil locate and
specific, and gelatinase granules. Although this identify infections? How does it transition
subdivision is practical, these designations are at the correct time and place from an in-
largely artificial. Granules are formed through a active cellular bystander to a fully activated
continuous process; vesicles bud from the Golgi microbial killing machine? This transition
apparatus and fuse, producing granular struc- process, during which the neutrophil inte-
tures. The content of these structures is dic- grates a complex barrage of environmental
tated by the transcriptional program active at cues and translates them into specific actions,
the time of their formation. As the maturing is known as neutrophil “activation.” As it
neutrophil sequentially alters its transcriptional pursues microbes, the neutrophil will enact an
profile, granule content changes, resulting in a impressive multitude of cellular mechanisms:
continuum of granule species with overlapping It will mobilize secretory vesicles and granules,
cargoes (6). identify chemotactic gradients and traverse
The release of neutrophils from the bone them through destruction and reorganization
marrow is tightly regulated in healthy in- of the actin skeleton, penetrate the endothelial
dividuals: Chemokines control the passage barrier and navigate a course through the
of PMNs into circulation and maintain a basement membrane, and begin transcription
pool of cells ready for release in case of of cytokines for recruitment of new immune

www.annualreviews.org • Neutrophil Functions 461

Syk. such as the bacterial-derived clustering of the β2 integrins on the surface of burst: a rapid increase lipopolysaccharide (LPS) and fMLP. endothelial cells (7. including Src migration by digesting the protein mesh 462 Amulic et al. in oxygen consumption upon as the classical chemoattractants and cytokines engage their endothelial ligands. 16). due to production of (IL)-1β. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 cells. and IL-17. 10). of laminins and collagen type IV. chemoattractants. a protein mesh consisting largely neutrophil at this early time point? The engage. it will seek the insulting pathogens and (PI3K). mediate cell adhesion acts as a monitor for host distress. neutrophil for its final chemotactic pursuit: The latory system. as oxidase side: the P-selectins. phosphoinositide 3-kinase site. and signaling endothelial cells near the inflammatory site. these Upon arrival at an endothelial cell junction. The β2 integrins then by Harvard University on 09/10/13. prepares the ICAMs (5). Once through the endothelial lining. resulting in selectin-mediated integrins and their endothelial partners and tethering of neutrophils to the vessel wall. the neutrophil must On the surface of neutrophils. lipodium where chemokine and phagocytic where flow dynamics and the constricted space receptors are concentrated. where the neutrophil stage for integrin activation and firm adhesion. members of neutrophil activation tumor necrosis factor (TNF)-α. are often the best-suited location chemotactic gradients. the cytoskeleton is are particularly amenable to increased random rebuilt and targeted toward movement along probing. As the neutrophil rolls well as direct cell-cell host-produced inflammatory signals are along the endothelium. negotiate a path through the endothelium into tively expressed proteins are critical for recog. β2 integrin family of proteins (LFA-1 and Integrins: Mac-1 proteins on the neutrophil). interaction with interaction and Annu.30:459-489. . For personal use only. the underlying tissue. 8). prompt endothelial cells to resulting in arrest of neutrophil rolling and ROS by the NADPH produce adhesion molecules on their luminal firm adhesion. As neutrophils traverse the circu. In a process dependent nition of the endothelial inflammatory signals: on β2 integrins and ICAMs. Downloaded from www. 2012.and E-selectins of complex interaction between (a) the neutrophil endothelial cells. Now firmly adhered. two constitu. This cascade initiates a number Selectins: transmembrane weapons. patrolling After selectin-mediated rolling. random contact with the endothelium. upon arriving at the infection family kinases. and the neutrophil commitment to microbial the neutrophil must navigate the basement killing commences. firm adhe- transmembrane sion is characterized by the arrest of neutrophil receptors that mediate NEUTROPHIL ACTIVATION attachment to the rolling in preparation for transendothelial extracellular matrix. these compounds stimulate the selectins. interleukin the ICAM-1 immunoglobulin superfamily. The initiation of these processes oc. E-selectins. neutrophils via binding to sugar vessels and vigilantly seeking out indications of enter a “firm adhesion” state mediated by the moieties an incipient inflammatory response. producing a leading-edge lamel- probe the vessel wall. cytokines.annualreviews. and initiation of the for neutrophils to encounter the stimulated neutrophil oxidative burst begins (17. and several well as continuing input from inflammatory members of the integrin superfamily. Rev. 18). and p38 mitogen-activated protein unleash its extensive arsenal of antimicrobial kinase (11–13). they continuously and randomly cell spreads. Upon site of transmigration is reached (19–21). a molecules engage the P. It is here migration through the endothelial junction that the complex activation cascade begins (13). (b) neutrophil surface proteins and various This is followed by a characteristic “rolling” of endothelial junction molecules results in trans- neutrophils along the endothelium. neutrophils the glycoprotein P-selectin glycoprotein crawl along the vessel wall until a preferred ligand-1 (PSGL-1) and L-selectin (9. 14). the chemoattractants and cytokines. of changes in neutrophil biology and sets the glycoproteins that curs in the bloodstream. the postcapillary venules.org abundant. bacterial products results in activation and Oxidative/respiratory These stimulants. Speculation ment of PSGL-1 and L-selectin on neutrophils abounds that granule proteases assist in this activates a variety of kinases. This integrin engagement. Immunol. as At inflammatory sites. as well the neutrophil (15. bacterial-derived and migration (13. Ultimately. What changes occur in the membrane.

flagellin (TLR5). 2012. the stimulation of FPR1 triggers this phenomenon is the strong priming effect the release of ATP. At low concentrations. stimulates L-selectin shedding and increased fMLP). prompt assembly of the oxidative burst ma. where no discernible gradient exists. executing tants further stimulates the oxidative response programs of phagocytosis. Rev. as is the case with the fMLP receptor FPR1 or IL-8 induces degranulation of neutrophils (27).org • Neutrophil Functions 463 . induction of the the end result of this signaling cacophony is oxidative burst (25. As the neutrophil nears unambiguous: The neutrophil begins to imple- its target. which initiate a sig. 23). pursuing host. point of high chemoattractant concentration. A notable example of Furthermore. degranulation. all but one the activation process is complex.. These worthy of note. oxidative burst. and the exact of these receptors (TLR3) are constitutively effects of priming. In addition. physiology at different concentrations. a the neutrophil follows chemotactic gradients phenomenon exemplified by one of the key Annu. The movement to ever-higher compounds will now be the primary dictators concentrations of chemoattractant is key in of neutrophil behavior and assume respon. and signal- expressed. continued activation by chemoattrac. whose autocrine action of LPS on the fMLP response (28). At the highest concentrations. neutrophil by a chemoattractant often results they are responsible for recognizing a number in endocytosis of the corresponding receptor. these chemoat.g. in parallel. In the interstitial space. During this process. Regardless. however. bacterial molecule (30. IL-8. the process of setting neutrophil www. the neutrophil finds itself in a its antimicrobial arsenal. expression of β2 integrins.g. That is. the chemokine receptors). alone they pathway (22. the fully in an antimicrobial attack state.. IL-8 pathogen-derived chemoattractants (e. another critical feature of the specific nonself patterns present on many mi. environment is awash in a soup of chemoat.e.30:459-489. including LPS (TLR4). response to other stimuli.. of pathogen-derived compounds. this process. For personal use only..org toward the invading microbes.annualreviews. The rich and varied input lipopeptides (TLR2). collectively thus leading to a desensitization of the neu- called pathogen-associated molecular patterns trophil to repeated stimulation with the same (PAMPs). Concomitantly. The complex signaling cascade leading to tractants and inflammatory stimulants.g. neutrophil-recruiting chemokines and ac- produced cytokines (e. slightly higher by Harvard University on 09/10/13. the neutrophil is now much different inflammatory milieu: Here. In neutrophils. sive experimental evidence for this is lacking. both final neutrophil activation has several facets host derived and of pathogenic origin. and their stimulation contributes ing are incompletely understood. 31). 26). tivators. Upon finally reaching a NETosis (i. ment its regime of microbial killing. IL-8) and. and received by a neutrophil during this final leg of DNA (TLR9). Stimulation of the of this family is the Toll-like receptors (TLRs). Downloaded from www. to further activation. Downstream molecules stimulate the oxidative response only mildly. tractants bind to their respective neutrophil concentrations result in initiation of the receptors (often G protein–coupled receptors. and and degranulation. stimulation process is the desensitization to pre- crobes (25). through activation of purinergic receptors is exposure of the neutrophil to LPS induces critical for the initiation of effective functional assembly of the NADPH oxidase machinery on responses in neutrophils (24). as individual chemoattractants sibility for initiating the concluding steps of may have very different effects on neutrophil neutrophil activation. but they dramatically enhance the subsequent chinery. e. Perhaps the best-known example viously encountered ligands. Immunol. conclu. the pattern-recognition activation of this machinery (29). desensitization. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 subsequent to degranulation. a hallmark of neutrophil activation. the Once the endothelial barrier has been neutrophil halts and begins the final release of traversed. the membrane. fMLP stimulation then induces a family of molecules. is activated through recognition of receptor priming. many chemoattractant molecules naling cascade dominated by the MAPK/ERK exert a “priming” effect. In this case. In contrast to receptors.annualreviews.

to complete activation and culminates in the The initiation of these microbicidal actions premiere killing functions of phagocytosis. a Capture b Rolling c Firm adhesion Neutrophil Integrin P-selectin and ICAM PSGL-1. overwhelming) view of neutrophil activation.30:459-489. and NETosis. 464 Amulic et al. more insightful to view neutrophil activation ever. and it begins to implement its antimicrobial agenda. The myriad its simplicity: (1) kill microbes. unanswered by the preceding exposition: What and signal cascades with varying effects and exactly is meant by the (admittedly ambiguous) outcomes. and (3) when in doubt. one could be (erroneously) led to believe that neutrophil activation refers only to NEUTROPHILS AND THE direct stimulation of the oxidative burst. It is. To fulfill this antimicrobial agenda. Rev. the neutrophil traverses through the endothelium and arrives at the site of inflammation. followed by (c) integrin-mediated firm adhesion. 2012. the neutrophil releases cytokines that recruit other immune cells.org In fact. journey through the activation process. priming steps. the selectins. (2) do no interactions that occur during a neutrophil’s harm to the host. therefore. Immunol. circulating of the literature presents the inexperienced neutrophils to their microbe-eliminating. indicates the final stage of the neutrophil’s degranulation. neutrophil is both effective and ruthless in fied view of a complex process. however. and formation of neutrophil extracellular traps (NETs). an oversimpli. release of granular antimicrobial molecules through degranulation. This is. parsed by the complex neutrophilic signaling trophils and the Elimination of Microbes. as this ELIMINATION OF MICROBES has been the canonical in vitro activation assay The basic instruction set of the activated by Harvard University on 09/10/13. Here. E-selectin L-selectin Phagocytosis Endothelial cell Degranulation Cytokine secretion NETs Figure 1 Neutrophil recruitment to sites of inflammation. (b) Selectin-mediated rolling along chemoattractant gradients then ensues. (a) Close to the inflammatory sites. Annu. a process that gradually leads below). a prominent question remains largely as a continuum of processes. How.annualreviews. which serve to capture circulating neutrophils and tether them to the endothelium. mechanisms. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 extracellular traps) (see the section on Neu. all focused on the realization of phrase “neutrophil activation”? A quick scan one goal: the transition of naive. . Among the processes employed are engulfment of microbes via receptor-mediated phagocytosis. Subsequently. The circulating neutrophil must recognize signs of inflammation and migrate to areas where its antimicrobial arsenal is needed for the elimination of infection. For personal use only. for decades. stimulated endothelial cells expose a class of molecules. see journey toward an inflammatory site must be rule 1. Downloaded from www. reader with a sometimes rather conflicting (and tissue-resident counterparts (Figure 1).

For personal use only. proteins BPI. and their contents seem determined by the timepoint during hematopoiesis at which they are produced (5). proteinase 3 (PR3).annualreviews. however. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 neutrophils possess an array of toxic weapons needs of neutrophils. and are the first formed during neutrophil maturation. Expect. with secretory vesicles being the first to fuse with the plasma membrane and the azurophilic granules demonstrating the least degranulation propensity. CD14. PR3. collagenase. 33). and cathepsin G (CG) (34). Gp91phox/p22phox. Gp91phox/p22phox. 2012. Neutrophil granules carry a rich variety of antimicrobials and signaling molecules. www. B12BP. Considerable overlap exists in the cargo of the different granules. CD11b. activation of immune exploit any and all weaknesses that microbes As mentioned above. Immunol. edly.3 μM in diameter. and their method of release is critical as peroxidase-positive or primary granules) are to understanding neutrophil function. MMP25. FPR. haptoglobin. Other stream and then correctly deploy them at the cargo of this granule class include the defensins. the largest. Downloaded from www. Gp91phox/p22phox. Granules are. there are three cells upon infection or might present during the course of infection. an enzyme of dangerous substances through the blood. Therefore. when uncontrolled it leads to An understanding of these weapons. alkaline β-glucuronidase SLPI. these cally tuned mechanics that address the unique granules are brimming with antimicrobial Primary Secondary Tertiary Secretory Granule type (azurophilic) (specific) (gelatinase) vesicles Stage of Myeloblast Promyelocyte Myelocyte Metamyelocyte Band cell formation PMN Degranulation propensity Characteristic Lysozyme Complement receptor 1 proteins Myeloperoxidase Lactoferrin FcγRIII Elastase Gelatinase Defensin Other Cathepsin G.30:459-489. fundamental types of granules in neutrophils injury. C1q-R. These antimicrobial weapons for dangerous substances. plasma proteins β2-microglobulin. CRISP3 Figure 2 Neutrophil granules. They are named for Granules and Degranulation their ability to take up the basic dye azure A and Annu. arginase-1. CD11b. Rev. critical in the oxidative burst (32. they are active and in- Inflammation: vary considerably in their methods of action dispensable participants in almost all neutrophil recruitment and and thus reflect the neutrophil’s attempt to activities during inflammation. and a number of serine proteases: evolved in neutrophils: the granule. Granules also differ in their ability to mobilize. and tertiary or gelatinase). bactericidal/permeability-increasing by Harvard University on 09/10/13. measuring approximately 0. their (Figure 2). structures called secretory vesicles are also considered to be a granule subset.org • Neutrophil Functions 465 . Azurophilic granules (also known tissue damage action. oroscomucoid. MMP25. azurocidin. They are typically divided into three types (primary or azurophilic. that are carefully regulated through controlled far more than just latent repository organelles mechanisms. CD10.annualreviews. As such. appropriate time. neutrophil elastase (NE). surprise that a specialty storage organelle has protein (BPI). pentraxin 3. Additionally. hCAP18. β2-microglobulin. these structures are replete with specifi. sialidase. CD11b. CRISP3 CD13. phosphatase.org The neutrophil must safely transport a plethora contain myeloperoxidase (MPO). secondary or specific. it comes as no lysozyme. heparanase. NGAL.

thus altering its molecular composition specific granule membrane (45). Downloaded from www. al. compartment. brane. 39.annualreviews. neutrophil maturation (5). 42). do not contain MPO. Consequently. Because granulation presents the astute observer with of this varying mobilization propensity. The second class of granules. and contain few antimicrobials. neutrophils are last population of granules formed during exposed to further activation signals that initiate by Harvard University on 09/10/13. hCAP-18. con- membrane-bound molecules employed during tributing to the antimicrobial activities of this neutrophil migration. 35). This fusion (6). In both cases. NADPH oxidase machinery. are Fusion of the secretory vesicles with the plasma also MPO-negative. the membrane of the granule as flavocytochrome b558. 2012.1 μM stimulation through selectins and chemoattrac- diameter). The as the FcγRIII receptor CD16 (5. permits assembly of the NADPH oxidase com- strate varying propensities for mobilization in plex and allows reactive oxygen species (ROS) response to inflammatory signals: Azurophilic production both inside the phagolysosome and granules are the most difficult to mobilize. These granules are formed after necessary for continued activation of the neu- azurophilic granules. are smaller (0. among others. This results in the transi- Annu. been conclusively demonstrated (43. For personal use only. secretory vesicles (37–41). These granules are also the ceed through the endothelium. resides in the brane. releasing their potent antimicrobials granules are mobilized and fuse with either the into the tissue. whose membranes are rich in key factors lactoferrin. such as gelatinase interaction with the endothelium. with a particular stage of neutrophil activation. As they pro- and leukolysin. are also leasing metalloproteases. 39). complement and fMLP receptors. 38. a fourth set mobilization of gelatinase granules. The different classes of granules demon.30:459-489. or fuse with the plasma mem- As a neutrophil proceeds through activation. thereby re- of structures. initiation of the oxidative burst and mobiliza- dominantly of plasma-derived proteins such as tion of the azurophilic and specific granules. a component of the becomes a permanent part of the target mem. mediated by β2 integrin number of metalloproteases. below). Degranulation of primary lowed by specific granules. In contrast to the classical basement membrane. Finally. but instead are formed through endocytosis At the inflammatory site. the β2 inte- range of antimicrobial compounds including grins. the secretory vesicles. fol. ternal environment. and are char. although this has not granules. each a tempting proposition: Could these granular 466 Amulic et al. are smaller than specific membrane exposes these components to the ex- granules. though regulation of intracellular calcium levels The release of granular proteins during de- appears to play a salient role (32. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 compounds and function as a primary reposi. and lysozyme (33. 44). inhospitable to invading pathogens. Rev. and secondary granules contributes to the and finally. they also contain a wide trophil. particular importance for the oxidative burst. The activity of these commonly considered part of the neutrophil proteases may help neutrophils traverse the granule family. gelatinase granules. their cargo consists pre. granule subset has been traditionally associated tory for the molecular weaponry of neutrophils. third class. as well NGAL. the specific (or After neutrophils contact the endothelium. The fusion of specific granules plasma membrane or the phagosome. releasing with the plasma or phagosomal membrane is of their contents into the respective environment. tants induces mobilization of secretory vesi- acterized by the presence of the glycoprotein cles. these do not bud from the Golgi. Immunol. The membrane of secretory vesicles These granules either fuse with the phagosome serves as a reservoir for a number of important (see section on Phagocytosis. including.org but they serve as a storage location for a tion to firm adhesion. complete acti- in the end stages of neutrophil maturation vation of the neutrophil ensues. outside of the cell. . The creation of an antimicrobial milieu at the in- underlying mechanisms for this differential flammatory site and produces an environment mobilization are not entirely understood. albumin. the gelatinase (tertiary) granules. secondary) granules. prompting (36).

IY30CH19-Zychlinsky ARI 17 February 2012 13:38 components also serve as signaling molecules whereas others may be redundant. including ditions that mimic an infection site.annualreviews. testing the relevance of antimicrobials in vivo cruitment. degranulation and its effects on neutrophil ac. Many of these antimicrobials were antimicrobials are thought to disrupt essential identified through biochemical fractionation of microbial functions. Therefore. Most and (c) proteins that deprive microorganisms evidence for in vivo degranulation relies on of essential nutrients. integrity. particularly chal- proteins may increase macrophage bacterial lenging. there by Harvard University on 09/10/13. much biochemical identification of neu- extracellular concentration of released granule trophil antimicrobials has been performed in proteins is insufficient to exert extensive micro. Here we present an observation of increased levels of extracellular overview of this rich field of investigation. most notably highly conserved throughout evolution (47). Because in vitro tests are often exe- cuted at high antimicrobial concentrations to obtain maximal microbial killing in the shortest Antimicrobial Proteins possible time. it is unclear whether this disrup- Neutrophils produce a plethora of peptides and tion reflects their mechanism of action under proteins that directly or indirectly kill microbes physiological conditions. Thus. Immunol. or production of energy. Mice. is known about antimicrobial concentrations nonetheless. Under artificial conditions. In ad- could be advantageous in situations in which the dition. cell damage. species with abundant neu- bicidal effects.org • Neutrophil Functions 467 . Rev. ablation of a single antimicrobial gene clearance by enhancing phagocytosis (46). some of them primarily through other means. A ity with a particular mutation. This is. evidence for clinical or biological this process truly relevant during the in vivo relevance of these molecules is still lacking. Annu. PR3 and azurocidin. The diversity of antimicrobials sug. granular proteins at inflammatory sites. mice lack the genes for some antimicro- By necessity. Even There are more than 800 antimicrobial so. and understandably so: Historically.org teins would instead operate as signaling and re. 2012. which are genetically tractable. been restrained by technical limitations. Indeed. Downloaded from www. This information. the granule pro. release of granular components could occur peptides described in nature. For personal use only. inflammatory response? The data here are There are three main types of antimicro- sparse. about the synergistic interactions of different www.annualreviews. neutrophil granule is essential. are few clinically relevant innate immune de- tivity have been acquired through biochemical ficiencies that directly link antimicrobial activ- approaches performed exclusively in vitro. such as DNA replication. microbial surfaces. showing in vivo relevance is chal. through formation of neutrophil extracellular These peptides are often charged. neutrophil extracts. Furthermore. can induce monocyte re. rabbits and humans. have neutrophils that function differently from cruitment factors (see section on Neutrophils those of other species. some. (b) enzymes. With the probably promotes their initial interaction with advent of intravital microscopy techniques. some (Table 1). antimicrobial mechanisms is to study their dence suggesting this does seem to be the case: function during concerted action and in con- Granule proteins from neutrophils. direct observation of the degranulation process many of these peptides disrupt the membrane in vivo may soon be realized. In such cases. Interestingly. achieved at inflammatory sites or in the phago- lenging. This may only subtly affect immune defense. the bials: (a) cationic peptides and proteins that possibilities for such an in vivo observation have bind to microbial membranes. as well as information gests that some of them evolved to act together. Alternatively. trophils. tioned. below). One of the for subsequent inflammatory cell recruitment? challenges in understanding the neutrophil’s Recent studies have provided experimental evi. most data on neutrophil bials identified in humans. and their in vitro activity transcription.30:459-489. a feature that traps. however. or cell lysis. Little is easily demonstrated in optimized conditions. with few pertinent question therefore presents itself: Is exceptions. as already men- in Immune Cell Cross Talk.

. The neutrophil cationic antimicrobial Neutrophils also contain a number of peptides include defensins and cathelicidins. Downloaded from www. and binds LPS avidly. Neutrophils mostly produce α-defensins. studied LL-37. Rev. but death then follows (51).annualreviews. full-length cationic antimicrobial proteins. is essential for designing appro. BPI binding to change under physiological conditions and LPS results in increased bacterial permeability increase their activity (48). A surprising num. HNP-2. peptides derived from them) as microbials matory sites. For personal use only. cell ber of functions are assigned to defensins.30:459-489. DNA activation of dendritic cells (DCs) (50). an essential bacterial nutrient  Binds to the lipid A part of LPS. Immunol. much like its structural ple disulfide bonds and whose structures may cousin the LPS binding protein.  Mechanism independent of a proteolytic activity by binding to the bacterial membrane Azurocidin Mechanism independent of a proteolytic activity by binding to the bacterial membrane Metal chelator proteins Lactoferrin  Alters bacterial growth by binding to iron. remains to be demonstrated in vivo (53). they may potentiate of action. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 Table 1 Mechanism of action of neutrophil antimicrobial proteins Antimicrobial peptide Antimicrobial mechanisma Cationic antimicrobial peptides α-defensins (HNP-1.org Proteinase 3 (PR3) Mechanism independent of a proteolytic activity by binding to the bacterial membrane Neutrophil elastase (NE). Cathelicidins.  Permeabilize membrane bilayers containing negatively charged HNP-3. a including BPI and histones. Interestingly. antimicrobials. causing a release of LPS from the cell wall and an increase in membrane permeability Calprotectin Alters bacterial growth by sequestering manganese and zinc a Only direct actions of neutrophil antimicrobial proteins on microbes are listed in the table. 2012. BPI is cationic protein family whose members possess multi.  Cleaves bacterial virulence factors and outer membrane cathepsin G (CG) proteins by Harvard University on 09/10/13. from larger proteins. and hydrolysis of bacterial phospholipids. and in addition to their priate in vitro conditions to probe mechanisms antimicrobial activity. Interestingly. RNA as well as protein biosynthesis  Inhibition of bacterial cell wall synthesis LL-37 Transmembrane pore-forming BPI Increase bacterial permeability and hydrolysis of bacterial phospholipids by binding to LPS Histones Unknown mechanism Proteolytic enzymes Lysozyme Degrades bacterial cell wall Annu. are extremely effective antimicrobials and inhibition of bacterial cell wall synthesis (49) were one of the first antimicrobials described was recently shown at low concentrations that (52). are proteolytically processed Given their dual role as an architectural 468 Amulic et al. HNP-4) phospholipids  Inhibit DNA. histones none have been validated in vivo. The significance of histones (and of the may be more similar to those present at inflam. including the well.

it to infections. Two of these chela.30:459-489. Superoxide can also react with nitric specificity (56). Interestingly. These serine proteases also play a antimicrobial actors in the phagosome (65). all caused by mutations destruction. 2012. antimicrobial activity: Neutrophils from The final class of neutrophil antimicrobials chronic granulomatous disease (CGD) patients consists of a number of proteins that chelate kill microbes poorly. their Reactive Oxygen Species in vivo significance is particularly difficult to Upon activation. indicating that their activity is 62–64). Upon degranulation into the mutations in humans. bacteria. Annu. Hypochlorous recombinant granulocyte macrophage colony. which sequesters zinc (60) and results in reactive oxygen cascade (66). Immunol. shown in mice deficient in this enzyme (54). but do not degrade. Rev. reactivity. Mice deficient in NE or CG is assumed to have direct antimicrobial effects are highly susceptible to bacterial and fungal in the phagosome. indicating the possibility of the oxide. perhaps as a result of their with amine groups. a theoretical infections (57. reducing molecular oxygen to superoxide. oxidant. NE cleaves by Harvard University on 09/10/13. result in neutropenia. forming hydrogen enterobacterial virulence factors with high peroxide. Chronic The second class of neutrophil antimi. and disease (CGD): proteolytic enzymes that participate in microbe permeability to membranes (62). This can be rescued by the administration of including hypohalous acids. It is mis. and NE. salient role in autoimmunity (see discussion in ROS are clearly important for neutrophil section on Autoimmunity. may be the most relevant cationicity. CGD patients can control catalase-negative tors are lactoferrin. many potential are tightly regulated intra. Another protein. below) (59). Superoxide. granulomatous crobials encompasses a broad assortment of cause they differ in their stability. Unexpectedly. Thus. first identified in milk. though not a strong deployed under specific conditions.annualreviews. NE strong oxidant. which is produced at high levels at coevolution of microbial virulence factors and inflammatory sites. Lysozyme destroys the bacterial rendering the ROS can modify and damage other molecules. collectively known as the serpro. their own hydrogen peroxide. which binds preferentially to iron. NADPH ox- “nutritional immunity” (61). is a member of the same family but lacks the hypochlorous acid produced would react protease activity. model of the phagosome suggests that most of cidin. it still kills with host proteins before reaching the bac- microbes. making these patients essential metals from microbes and possibly susceptible to many infections. MPO can react with hydrogen of this enzyme in mice. idase is also implicated in the regulation of www. NADPH oxidase wall. 58). but not genetic ablation phagosome. However. a antimicrobial effectors. this occurred independently of its The NADPH oxidase complex assembles susceptibility to enzymatic activity (55). azuro. as properties exploited by the host cell for signal. peroxide to produce various reactive species. all have antimicrobial activity independent produced when hypochlorous acid reacts of proteolysis. impact bacterial growth. nonfunctional. autoinflammation and begins the reactive oxygen cascade by CG. suggesting that these proteins may terium. Neutrophils also con.org • Neutrophil Functions 469 .org cidins) that exhibit differing specificities. however. to form peroxynitrite. thus providing protectin (also called S100A and many other a substrate for reactions downstream in the names). see References by serpins.annualreviews. leading to think of ROS as a single entity be. ing and antimicrobial action. on the phagosomal and plasma membranes infection and tain several serine proteases (including PR3. However. acid. a process called the respiratory burst. Downloaded from www. rapidly dismutates. is more reactive than su- patients still exhibit significant susceptibility peroxide and is antimicrobial in vitro. thought to be the major product of MPO stimulating factor (GM-CSF). and cal. Of further interest. This model predicts that chloramines. neutrophils produce ROS in demonstrate. these in the phagosome. making it an obvious antimicrobial. characterized by Surprisingly. For personal use only.and extracellularly reactions can occur (for details. which produce. They Downstream of superoxide. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 scaffold for DNA and as antimicrobials.

Immunol. In addition. (including phosphatases. hydroperoxyl radical. but this theory of deficient individuals suffer from frequent or se.annualreviews. which ROS owing to their short half-lives (72). for ROS detection that meets these criteria is by Harvard University on 09/10/13. these species are highly reactive. Because tween the neutrophil and the microorganism. there may be a broader include transcription profiling of superoxide reason for this discrepancy. Furthermore. other reactive species (e. Traditional probes for Annu. neutrophil granule After engulfment.org suggest that MPO’s products are not essential ROS do not meet these specifications. technical issues. One promising new approach ide. which does not require membrane extensions and caspases) by direct oxidation of cysteine or pseudopods (77). vere infections. by cellular redox ter characterized and includes two prototypical buffering systems and by limited diffusion of examples: FcγR-mediated phagocytosis. peroxynitrite) can still be produced in the the use of redox-sensitive fluorescent protein- neutrophil phagosome. The mild effects of MPO deficiency being used in vivo. 69). and partial MPO deficiency does not correlate with pathology (70). . through recognition thought to be too nonspecific to be involved in of PAMPs by pattern-recognition receptors. Some MPO. Studies of ROS are hampered by various and a few have been mistaken for CGD patients. metalloproteinases. Downloaded from www. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 inflammation. or signaling. (76). phagocytes.g. ization depend on the type of interaction be- ROS can modify host molecules. the probes often become radical of MPO. hydroperoxyl radical is based probes. Residual activity of MPO may be sufficient for antimicrobial activity: In Phagocytosis the case of CGD. most MPO-deficient individuals in should be specific. and capable of immunity. 2012. There is controversy around MPO-deficient individuals do not have striking which ions and which channel are responsible clinical manifestations (68. Epidemiological studies distinguishing receptor-mediated process during which a par- the degrees of MPO deficiency and their ticle is internalized by the cell membrane into correlation with clinical manifestations may be a vacuole called the phagosome. or hydrogen peroxide–sensitive genes as well gies can distinguish between individuals who as the detection of relatively stable products of are partially and completely MPO deficient. for charge compensation. 74). superoxide generation patients often suffer from autoinflammatory leads to an ionic influx into the phagosome to diseases (67). Other methods that can be used in vivo trations (65). this may activate gran- Paradoxically. protease activation is certainly intriguing (69). Rev. targetable to particular the developed world have apparently normal intracellular compartments. in for antimicrobial action. residues. hydrogen peroxide. specificity can be achieved opsonin mediated. such as roGFP and HyPer predicted to be present at antimicrobial concen. in the absence addition. However. a probe for ROS However. The latter mechanism is bet- on the submolecular level. tative matrix (75). (71). they are often Interaction can be direct. the mechanistic details of internal- In addition to direct antimicrobial action. and the reversible regulation of various targets complement receptor-mediated phagocytosis. acquir- vation of their inhibitors or by direct oxidation ing its lethal properties only after a drastic 470 Amulic et al. A relies on the formation of pseudopod extensions well-studied example of ROS in signaling is for engulfment of IgG-opsonized particles. especially with Candida species. Indeed.30:459-489. superox. It is an active. compensate for charge. which explains why CGD (73. Ideally. the nascent phagosome proteases can be regulated by oxidative inacti. even 1% of normal NADPH Phagocytosis is the major mechanism to re- oxidase activity leads to an improved prognosis move pathogens and cell debris. As with other necessary to understand the function of MPO. However. Modern technolo. although MPO is required ule proteases by releasing them from their pu- for neutrophil microbicidal activity in vitro. species (76).. reactive oxygen using mass spectrometry (76). is relatively benign to microorganisms. For personal use only.

to test the relevance pathogens.annualreviews. via a mechanism that also requires ROS (90). While the concentrations of antimicrobials (89). Neutrophil phago. Rev. not cells deficient have evolved strategies to survive inside neu.org toxic to most pathogens. can efficiently block gran. Ideally. an active form of cell death that especially in conditions of nutrient granules to the phagosome. such as Salmonella typhimurium and of NETs. Histone aureus confers antiphagocytic properties (81). in ROS production (92). The maintenance of this alkaline MPO are required for NET formation in re- pH is essential for the activation of the major sponse to chemical and biological stimuli (87. a “NETs knockout” organism should Streptococcus pyogenes. they are thought to somal pH regulation also differs significantly kill microbes by exposing them to high local from that observed in macrophages. the factors that are important for www. although this has not been confirmed NADPH oxidase to the phagosome so that in primary human neutrophils (95–97). The polysac. within the phagosome further emphasizes the Moreover. macrophages.30:459-489. ules to the nucleus and degrades histones. be generated to investigate its response to ule fusion with the phagosome (84. phagosome maturation happens upon fusion of NETosis. 85). In fact. and creating a more hospitable in the process. and it is sustained 90. 87). modulating phagosome cated in NET formation (93). ROS production. Francisella formation. NE translocates from the gran- intraphagosomal environment.annualreviews. neutrophil phagosomal pH is initially alkaline completely understood. NETs trap many types of mi- these two mechanisms create an environment crobes ex vivo and have been found in various Annu. serine proteases NE and CG. and jointly. other conducted ex vivo. Downloaded from www. These strategies include interfering oxidase. Simultaneously. a uing fusion of acidic granules. proteins (88). 91). Finally. but this has so far been shown only tularensis prevents triggering of the oxidative using a nonspecific inhibitor of autophagy (98). Nitric oxide donors can induce NETs via NADPH oxidase activity. Upon stimulation. All ac- the maturation process are described in more tivators of NET formation tested so far require detail in Reference 80. Macrophage phagocytosis follows an Neutrophil Extracellular Traps contents are degraded endocytic maturation pathway: In neutrophils. in which cellular trophils.org • Neutrophil Functions 471 . disease models in vivo. aureus may be an exception. citrullination may also play a role in NET for- Helicobacter pylori can disrupt targeting of mation. but further along maturation. Au- superoxide anions accumulate extracellularly tophagy is also thought to be required for NET rather than in the phagosome (82). some pharmacological inhibitors. Not all pathogens succumb to the hostile although those experiments were done using environment of the phagosome. and although these are certainly Autophagy: a process instructive. The mechanism of NET formation is not by Harvard University on 09/10/13. the extracellular space (86. The pathogens. essential differences exist in neu. whereby delivery leads to release of decondensed chromatin into scarcity and infection of antimicrobial molecules into the phagoso. Upstream of NADPH trophils. it is not possible to variety of mechanisms evolved by intracellular eliminate the main components of NETs— pathogens to resist killing and enable survival DNA and histones—from an infection model. macrophage phagosome gradually acidifies. Unfortunately. burst and also inhibits ROS production in The majority of research on NETs has been response to other stimuli (83). 2012. The fibrous mal lumen occurs. Immunol. lead- charide capsule expressed by Staphylococcus ing to chromatin decondensation (94). assembly structures termed NETs contain histones as of the NADPH oxidase on the phagosomal well as antimicrobial granular and cytoplasmic membrane allows ROS production. Key events of finding that awaits genetic confirmation. the Raf-MEK-ERK pathway is impli- with engulfment. as NADPH oxidase and of time (79). The reactive oxygen (78) and remains neutral for prolonged periods pathway is involved. For personal use only. neutrophils can undergo in lysosomes. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 maturation process. despite contin. S. Our understanding of importance of phagocytosis in the innate this process is largely based on studies in immune defense.

IY30CH19-Zychlinsky ARI 17 February 2012 13:38 NET formation. below).org patient’s neutrophils ex vivo (101). and ROS NETs. Downloaded from www. In addition. in the host. As they respond to infection or injury. and it may be that cytokine processing (112). NETs have also been observed in the airway fluids of cystic fibrosis patients. 111). and NE. Similarly. which restored NADPH oxidase of cytokines per cell. infection in a CGD patient was cleared after neutrophils typically produce lower amounts gene therapy. Macrophages Neutrophils participate in the communica. Here (107). and patrolling monocytes are among the initial tion networks that form the foundations of detectors of PAMPs and endogenous activators. such as during infections with large an appeal for immunological reinforcement. In addition to cytokines. timicrobial neutrophil functions. issuing instructions to practically the danger-associated molecular patterns (121). NETs can also have detrimental effects on primarily serves to recruit other neutrophils the host. On the other thermore. raising the we provide examples of how neutrophils possibility that NETs nucleate blood clots in interact with other cells to shape the immune the context of deep vein thrombosis (108). are associated such as hydrogen peroxide (119). neutrophils and their relatives in the mono- cyte/macrophage lineage coordinate their NEUTROPHILS IN IMMUNE activities. NETs are especially important under certain The initial neutrophil cytokine response is by Harvard University on 09/10/13.g. please see References autoantibodies.annualreviews. Because NETs expose self molecules (113). all other immune cells. the immune system has redundant mech. including on Autoimmunity. IL-8. The most abundantly produced cytokine. Immunol. lupus erythematosus (SLE). pathogens that are not readily phagocytosed. a persistent Aspergillus with other immune cells (e. these cells undergo a tran- characterized by thick. leading to alternating waves of re- CELL CROSS TALK cruitment of these two cell types. but they are so abundant activity. of signaling molecules (110. NET formation. Compared decreases in lung and digestive function 100). For now. bacteria that express DNases transcribe little after leaving the bone marrow. often against chromatin and 115. Although neutrophils the CFTR ion On the one hand. Platelets also bind to NETs.. transporter.30:459-489. they lead to autoimmunity: matory IL-1β and TNF-α induce other cells NETs have been implicated in systemic to produce neutrophil chemoattractants (114. . tions to launch the adaptive immune response. Rev. suggesting a causal link 472 Amulic et al. response (see Figure 3). neutrophil-secreted proteases can hand. such as NADPH oxidase. lipids (118). immunity. Fur- Annu. The secrete cytokines and chemokines critical in the influx of neutrophils is followed closely by the unfolding of the inflammatory response and in arrival of monocytes. neutrophils neutrophil components (102–106) (see section release other signaling mediators. macrophages). neutrophils of neutrophils to the inflammatory locus. where they may increase the viscosity of the sputum and Monocytes and Macrophages decrease lung function (109). are also critical for other an. which may point to evolutionary scriptional burst that results in the synthesis sticky mucus and pressure to avoid entrapment by NETs (99. the The cytokines released by PMNs are often Cystic fibrosis: caused by defects in evidence for the relevance of NETs is indirect. 116). an autoimmune 115) (for a comprehensive list of cytokines disease characterized by the formation of produced by neutrophils. Platelet-induced granule contents (117). as virulence factors disseminate more efficiently once activated. As one of the first cell and these cells work to summon large numbers types to arrive at sites of infection. conditions. establishing the correct environmental condi- MPO. modulate signaling networks in vivo through anisms to fight infection. neutrophil-derived proinflam- extracellularly. and NET-mediated at inflammatory sites that their contribution but not phagocytosis-mediated killing by the to total cytokine levels is significant (4). 2012. For personal use only. formed during sepsis. They also with hepatotoxicity due to tissue damage communicate via cell-cell contact (120). synthesized de novo.

in a mouse model of Leishmaniasis. activate neutrophils by secreting IFN-γ. Neutrophil Monocyte DC Blood Figure 3 Neutrophil communication with other immune cells. Rev. neutrophils impede DC function by inhibiting antigen presentation to CD4+ cells. Neutrophils interact with a variety of cell types. recruited by neutrophils (122). in turn. CCL3 (MIP-1α) (123). recruiting DCs to the production (128). Indeed. Monocyte recruitment is also affected indirectly by neutrophils: via upregulation of endothelial Dendritic Cells adhesion factors. In contrast.annualreviews.annualreviews. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 Tissue T cell Activation and CD8+ Lymph node IFN-γ differentiation DC T cell Crosspriming ROS? T cell Arginase? Neutrophil IFN-γ Activation DC DC NK cell IL-12 DC Antigen presentation CD4+ Macrophage Neutrophil T cell Th1 Activation Neutrophil Annu. neu. neutrophils. Resolution of Inflammation. triggered a massive and rapid infiltration of In addition to recruitment. The circuitous trophils recruit monocytes via several different nature of the cross talk of these two cell types mechanisms. CCL20 (MIP. neutrophils the appropriate removal of their postmortem use granule proteins to induce extravasation remains (see section on Neutrophils and of monocytes in vivo. Neutrophils also interact with the adaptive arm of the immune system: They can act as antigen-presenting cells by cross-presenting antigen to CD8+ T cells. behind these temporal dynamics. microbicidal activity (129). This was recently illustrated chemoattractants by other cell types. repress 3α). increase of transendothelial Neutrophils can also recruit and activate permeability.30:459-489. They are important both for recruitment of monocytes and dendritic cells (DCs) to infected tissues and for enhancement of macrophage and DC activity. which activates T cells. enhancement of production of DCs in vivo. For personal use only. and CG (125–127). they also secrete IL-12. neutrophils mod. These cells secrete the ulate monocyte and macrophage cytokine chemokine CCL3. where ulation of the activities of these chemokines subcutaneous inoculation of Leishmania major via proteolytic processing (reviewed in 128).org • Neutrophil Functions 473 . Downloaded from www. Immunol. and CCL19 (MIP-3β) (124).org Bacteria by Harvard University on 09/10/13. in the lymph nodes. 2012. as shown for LL-37. azurocidin (HBP/CAP37). neutrophils (130). and differentiated into macrophages. T cells. Additionally. DCs and natural killer (NK) cells colocalize and enhance each other’s activity via receptor-receptor interactions and soluble mediators. further neutrophil chemotaxis and ensure and perhaps more unexpectedly. They express classical monocyte becomes obvious during inflammation abate- chemoattractants such as CCL2 (MCP-1) ment: Monocytes. and mod. Finally. below). directly enhancing their site of inoculation and initiating a protective www.

30:459-489. The anti-Ly6G monoclonal antibody is stimulates IFN-γ production by NK cells and inflammation and more specific and hence a better reagent for this further activates neutrophils. implicated in type of experiment (133). human neutrophils. important in mice. this was dependent the anti-Gr1 antibody oplasma gondii (132). Recently.annualreviews. thought to belong to isolated compartments. 142). Additional in vitro interactions be- firmed using anti-Ly6G antibody depletion: In tween neutrophils and NK cells are extensively Mycobacterium tuberculosis infection. which depletes neutrophils but may also (via CD18-ICAM-1 interactions). Interestingly. interact in a m´enage a` trois involving both molecule-3-grabbing In this case. For personal use only. Neutrophils affect T cell macrophages and that these neutrophils inhibit function indirectly via DCs. The T helper 17 (Th17) cell subset secretes IL-17. reviewed in Reference 138. and DCs intercellular adhesion leading to local secretion of TNF-α (120). 144). IY30CH19-Zychlinsky ARI 17 February 2012 13:38 Th1 response (131). Downloaded from www. Natural Killer Cells a key cytokine in the control of neutrophil Studies of interactions between neutrophil and dynamics. and their interpretation is neutrophils can induce the maturation of DCs frustratingly difficult owing to the question- in vitro through specific receptor-receptor able purity of cell preparations. and TNF-α 474 Amulic et al. ficking of DCs to lymph nodes and activation of CD4+ T cells were both dependent on PMNs. the reduced levels of cytokine cytokine signaling and direct cell-cell contact nonintegrin production foster specificity. as only proximal (137. In sharp contrast to the above findings. Cytokine communication in swarming activity in response to parasitic occurs in both directions: For instance. tact (139). prompting T helper cells that result in depletion of many other cell types in the DCs to produce IL-12p70. The functions and mechanistic which is secreted by T cells. A similar with Legionella pneumophila triggered produc- receptor 1 (Gr1): activation model was earlier proposed for Tox. The crucial role of neutrophils also activate NK cells by direct con- autoimmunity neutrophils in DC activation was recently con. G-CSF. NK for neutrophils) and and induce proliferation of T cells (120. 132). Furthermore.org be interpreted cautiously because they are on the basis of in vitro data. Neutrophil-activated DCs on both PMN-derived IL-18 and DC-derived RB6-8C5 reacts with both Ly6G (specific produce the proinflammatory cytokine IL-12 IL-12 (137). their interactions with T cells (135). infection of mice Granulocyte DCs receive the maturation signal. In this scheme. NK cells. some of these experiments should and a positive feedback loop has been proposed many immune cell Annu. increases phagocytic capacity (145). . Immunol. neu- types) based on the injection of the anti-Gr1 antibody trophils interact with a specific subset of DCs. In one report. ble that neutrophils have site-specific effects on PMNs secrete IL-12. produce IL-17. Th17 cells: subset of (RB6). which may be crucial for DCs and can be stimulatory at peripheral sites Th1 cell differentiation (141. It is possi. IFN-γ. a separate study using an immunization model neutrophils and T cells shape and impact showed that neutrophils recruited to lymph each other’s functions. Rev. Neutrophils express several T cell chemoattractants (116) exhibit fascinating and somewhat enigmatic be. timely traf. tion of IFN-γ by NK cells. and DCs colocalize at inflammatory sites. Ly6C (present on However. prolongs neu- details of these swarms are unknown and trophil life span. activated performed in vitro. 2012. which acts by upregulating expres- natural killer (NK) cells have historically been sion of CXCL8 (IL-8). Simultaneously. Similarly. Previously (134). as well as B cell development and maturation havior in the lymph nodes. but can also influence T cell function directly. infection (136). where they engage factors (143. They also and inhibitory in the lymph nodes. this study demonstrated that Lymphocytes DCs presented bacterial antigens when they A surprising finding in recent years is the exten- ingested infected neutrophils just as efficiently sive cross talk between cells located at opposite as they did via direct uptake of Mycobacterium ends of the immune spectrum. as outlined above. it DC-SIGN: dendritic cell–specific interactions between Mac-1 and DC-SIGN. cells. was shown that neutrophils. both qualitatively and nodes compete for antigen with DCs and quantitatively (140). induces gene expression. 138). and represent questions of immense interest. which in turn by Harvard University on 09/10/13.

Using mice in which profes. 2012. and is only now starting to be unraveled. type of inflammatory Neutrophils potentially have suppressive ef. NEUTROPHILS AND but its expression can be restored in mature RESOLUTION OF cells by inflammatory signals such as G-CSF or INFLAMMATION GM-CSF. Given that neutrophils are terminally diseases. in cell-death program. In line with these findings. Interestingly. neutrophil modulation of differentiated. below). and stromal cells Although some collateral damage to host (146). a finding that could have caspases (for an extensive review. removes apoptotic cells from inflamed sites. Proposed to have prosurvival effects. such interactions in vivo is still missing. mouse neutrophils that migrate to the lymph all of which prolong neutrophil survival. it also produces signals that abro- Neutrophils also appear capable of expressing gate further neutrophil recruitment. also been well characterized. and tissue erosion in the colon and rectum which inhibits T cell responses in vitro (147).org T cell responses by cross-presenting exogenous antigens in vivo. This process is essential for maintenance of and (b) hydrogen peroxide–mediated suppres. detrimental effects on inflamed tissues. 151). as proposed in a cancer model (119) (see “nonresolving inflammation. one such protein is survivin. neutrophil appendix infiltrates. www.” a problematic section on Cancer. also harmful to host cells. see Reference implications for susceptibility to autoimmune 155). Once neutrophils have executed by Harvard University on 09/10/13. and Bid). endothelial. Resolution of inflammation is an active process bowel disease fects on T cells via two proposed mechanisms: that limits further leukocyte infiltration and characterized by ulcers (a) L-arginine depletion by release of arginase. In humans. Bax. However. especially in light of the finding that by environmental conditions such as hypoxia. Apoptosis and Clearance sional antigen-presenting cells do not express Apoptosis is a central aspect of inflammation functional MHC class I. ptotic (Mcl-1 and A1) or proapoptotic proteins suggesting concomitant variations in the ability (Bad. Bak. leads to sion. macrophages to adopt an anti-inflammatory and they can present antigen to CD4+ T cells phenotype. it is unexpected that molecules adaptive immunity seems to be highly complex controlling cell proliferation regulate survival.30:459-489. Immunol. serious. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 by epithelial. the functional Neutrophil death is influenced by inflamma- significance for protective immunity remains tory mediators such as GM-CSF and LPS and unclear. Downloaded from www. These networks there are large variations in the ability of also control the expression of known antiapo- donors to express MHC class II (149. Phagocy- MHC class II and costimulatory molecules tosis of apoptotic neutrophils also reprograms under inflammatory conditions (149–151). in vitro (152–154). Collectively.org • Neutrophil Functions 475 . they die via a built- induce differentiation of cytotoxic T cells. However. Beauvillain et al. survivin The lethal cargo of neutrophils is not only is also highly expressed in neutrophils at sites destructive toward invading microbes. if impeded. and they also activate to activate T cells. with ulcerative colitis (156).annualreviews. but of inflammation. these Th17-associated tissues is inevitable during infection. tissue homeostasis and.annualreviews. present. and intestines of patients deployment must be tightly controlled. Direct evidence of condition that contributes to many diseases. It is expressed more highly in immature neutrophils than in mature ones. such as cystic fibrosis sputum. Thus. not only does These striking findings imply that neutrophils apoptosis reduce the number of neutrophils have characteristics of antigen-presenting cells. For personal use only. Therefore. The node have a negative effect on CD4 responses signaling networks that regulate survival have in an immunization system (135). neutrophils influence CD8+ Annu. (148) resolution. neu- cytokines increase granulopoeisis as well as the trophils must be removed before they have Ulcerative colitis: a recruitment and life span of neutrophils. showed that antigen-pulsed neutrophils can their antimicrobial agenda. Rev.

leukocyte recruitment flammation is the proper removal of apoptotic resolvins. Of these lipids. Thus. Rev. . nose. Interestingly. These are the most completely understood. This relies on the release of “find-me” effects. and (157). proinflammatory resistance of neutrophils to apoptosis (158). How do lipid mediators contribute to functions including Equally important for the resolution of in. This mechanism com- in the composition of secreted lipids. damage of vital organs replication. resolvins. in the arachidonic acid evant in Wegener’s granulomatosis and sepsis. shedding of L-selectin and the upregulation of genase and lipoxygenase pathways. detail elsewhere (118). This mechanism is rel. adhesion. such as trophil proteases. They showed that neutrophil exposure to these lipids increases expression Lipid Mediator Class Switch of CCR5 on the surface of late apoptotic neu- Soluble mediators play a crucial role in the trophils. lipoxins can inhibit the and are synthesized through the cyclooxy. microor- synthesized by PCNA is targeted for proteosomal degradation.annualreviews. inflammation also attributed to proliferating cell nuclear thus exists. such as lipoxins. 170). endothelial cells. results in secondary (BLT1) (161–167). recruitment and activation while inhibiting tract phagocytes. in synthesis of mediators with proresolving pathway.org which correlates with an increase in caspase-3 that can be used by neutrophils for resolvin 5-lipoxygenase. (168) also pro- necrosis and release of products that generate posed an interesting mechanism of action for proinflammatory signals (Figure 4). by Harvard University on 09/10/13. All three lipid mediators reduce neutrophil inflammatory cytokines tumor growth factor recruitment. governed by temporally regulated leads to reduced blood antigen (PCNA). lipoxins are prostaglandins and leukotrienes. lipoxins. cyclin-dependent kinases func. tissue destruction. Downloaded from www. tion as prosurvival factors in neutrophils. More recently. promoting monocyte/macrophage signals at early stages of cell death. leading to efficient sequestration of the resolution of inflammation. it associates The different biosynthesis pathways of prore- Prostaglandins and with procaspases in the cytosol and is thought solving lipid mediators have been reviewed in leukotrienes: lipids to prevent their activation. This factor usually resides expression of different lipoxygenases and the flow. The inhibitory effect signals are required for specific recognition of extends to all essential steps of neutrophil apoptotic cells. the termination of inflammation? Lipoxins. neutrophils synthesize in which chemokines are inactivated by neu- proinflammatory lipid mediators.30:459-489. distinct “eat me” neutrophil functions. a process that involves the lipoxin- (TGF)-β and IL-10 (155). thereby reducing adhesion of neu- neutrophils interact with various cell types in trophils to endothelial cells (169. Ingestion of apoptotic cells by responses: migration. which at. chemoattractants CCL3 and CCL5. and in the nucleus. and lungs. and leukocytes) and par- Pharmacological inhibition of these cell cycle ticipate in the transcellular biosynthesis of lipid Wegener’s granulomatosis: regulators induce caspase-dependent apoptosis mediators with anti-inflammatory and prore- vasculitis affecting the and block life-span extension by survival factors solving activities. have where stabilization of PCNA is associated with functions at the site of infection (159. where it is involved in DNA mobilization of different fatty acid substrates. 2012. 160). prosurvival effects were protectins. but in neutrophils. For personal use only. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 Similarly. At early plements other anti-inflammatory processes stages of inflammation. microbes also likely participate respectively. and caspase-8 activities. macrophages drives the production of the anti. In neutrophils. ganisms are also a source of lipid precursors cyclooxygenases and Annu. fibroblasts. resolvins. Likewise. synthesis. During β2 integrins in response to proinflammatory the later stages of the inflammatory response. platelets. Immunol. Finally. their vicinity (epithelial cells. During apoptosis. A major lipid mediator class switch kidneys. Ariel et al. In addition to derived from arachidonate precursor molecules neutrophil recruitment. The successful progression unavailable to recruit neutrophils to inflamed of inflammation appears to hinge on a shift sites (168) (Figure 4). and protectins exert cell-type specific cells. by contrast. 476 Amulic et al. and protectins in clearing in- flammatory sites. stimuli. Failure to clear these A4 receptor and the leukotriene B4 receptor apoptotic cells. and activation. The se- a particularly prominent role is assumed by questration of these chemokines means they are lipid mediators.

and anti-inflammatory lipid mediators such as lipoxins. Immunol. Proresolving lipid mediators also promote the expression of CCR5 on the surface of apoptotic neutrophils. driving the production of the anti-inflammatory cytokines tumor growth factor (TGF)-β and IL-10 and prostaglandin-E2 (PGE-2). inflammation leads to tion of the anti-inflammatory cytokines TGF-β COPD is a major cause of death in indus. Rev.30:459-489. which also recruit neutrophils. Instead. trialized nations. and protectins are produced. engulfment of apoptotic (COPD): lung disease obstructive pulmonary disease (COPD). microbes trigger the production of proinflammatory lipid mediators.annualreviews. flammation often directly correlates with poor during macrophage activation.g. 2012. which drive the lipid mediator class switch. For personal use only. In the early stages of inflammation. interaction of neutrophils with platelets induces the production of lipoxins. chronic pulmonary disease inflammatory lipids. Monocytes differentiated into macrophages ingest apoptotic neutrophils. Chemokines Chemokine clearance Microorganisms ? TNF-α NETotic IL-6 neutrophil Macrophage Figure 4 From inflammation to homeostasis: neutrophil apoptosis and lipid mediator class switching in the resolution of inflammation. As inflammation progresses.org • Neutrophil Functions 477 . e.annualreviews. including cystic fibrosis. a switch occurs. tobacco smoking. where smoking is a prime www.org Macrophage Apoptotic neutrophil Chemokines CCR5 IL-10 Leukotrienes TGF-β Prostaglandins PGE-2 by Harvard University on 09/10/13. secreting proinflammatory cytokines and chemokines that alert the immune system and promote neutrophil recruitment. and how this could impact inflammation resolution. and neutrophils is not accompanied by the release of caused by noxious rheumatoid arthritis (RA). produc. such as leukotrienes and prostaglandins. Anti-inflammatory lipid mediators initiate the resolution of inflammation by blocking neutrophil and promoting monocyte recruitment. clinical outcome. Nonresolved Inflammation NF-κB activation. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 Initiation Resolution of inflammation Leukotrienes Prostaglandins TNF-α Lipoxins Resolvins Protectins IL-10 TGF-β of inflammation Platelets Monocyte Lipoxins Neutrophil Lipoxin Resolvins Protectins Annu. In the presence of anti- pathologies. lipid mediators promote mains result in chronic inflammation. resident macrophages initiate an inflammatory response. resolvins. At the site of infection.. These nonphlogistic (noninflammatory) phagocyto- conditions lead to the accumulation of cyto- sis of apoptotic neutrophils by monocytes toxic substances and are associated with severe Chronic obstructive and macrophages. lung obstruction and IL-10 is increased (163. as typically occurs particles or gas. Notably. is currently unknown. A timeline of the inflammation process from initiation to resolution is summarized in the upper part of the figure. Downloaded from www. and IL-8 expression (170). The contribution of macrophages to the clearance of NETotic neutrophils. The failure of neutrophils to apoptose or mal- In addition to directly impacting neu- functions in the removal of their apoptotic re- trophil functions. Chemokine clearance upon phagocytosis of apoptotic neutrophils by macrophages further contributes to the reduction of neutrophil infiltration and the return to tissue homeostasis. providing a means of scavenging chemokines. The severity of in- proinflammatory mediators. 171). lipoxins also impact neutrophil activation by Disorders Associated with inhibiting ROS and peroxynitrite production.

IY30CH19-Zychlinsky ARI 17 February 2012 13:38 instigator of this disease. also demonstrate the cross talk between neu. a potent neutrophil chemoat- deformity tractant and proinflammatory agent. the recruitment of a mediators. Since then.org conferring the enzyme with anti-inflammatory reviews. thus tion. Neutrophils Leukotriene B4 can act in an autocrine manner do. this. It is. to the changing chemoattractant environment Rheumatoid arthritis (RA): chronic tion. 182). 2012. This in human disease remains to be established. of leukotriene B4 by neutrophils in joints convincing evidence for PMN-mediated DNA is essential for disease development (174). neutrophil action can also properties. and second wave of neutrophils is independent of matrix-degrading proteases (reviewed in Ref- this leukotriene B4–BLT1 pathway. including cytokines. The relevance of this model that affects many leukotriene A4 hydrolase (LTA4H). category. Downloaded from www. clinical studies indicate that neutrophil IL-1β into the joint. Neutrophils Cancer are the most abundant leukocytes present in the The link between cancer and inflammation synovial fluid of RA patients. In addition. immune complexes are essential for a “protumor” and “antihost” effect of these stimulating infiltrating neutrophils to deliver cells. A chronic neutrophil the previous section. At this erence 180). In one model. some cancers cells and sustains neutrophil recruitment (175. NEUTROPHILS IN DISEASE LTA4H is an aminopeptidase that inactivates Neutrophils are prominent players in the innate a specific neutrophil chemoattractant. antibody depletion of neutrophils reduces and chemokines that orchestrate neutrophil tumor growth (184). Immunol. via the neutrophil receptor BLT1 to promote They are abundant in tumors and influence the recruitment of a first wave of neutrophils tumor development through several secreted into the joint. Although mice (173–175). synthesis this has been demonstrated in vitro (178.annualreviews. a subject addressed in several prominent Annu. enhancement of metastasis (186). The majority of findings support stage. and 478 Amulic et al. This in turn induces the infiltration of tumors is associated with poorer production of chemokines by synovial tissue prognosis (181. seem to actively recruit neutrophils through 176). Another prime example of a disease linked to nonresolving inflammation is RA. Rev. ROS. First. For personal use only. However. promoting the accumulation of by Harvard University on 09/10/13. joints. the immune response and the clearance of infec- proline-glycine-proline tripeptide (PGP). certain malignant cancers both leukotriene B4 and PGP. Interestingly. Indeed. One of the key molecules controlling or if different subsets of neutrophils are suc- inflammatory disease the inflammatory response in the lung is cessively involved. Second. however.30:459-489. The protumor function recruitment in chronic inflammation. . and their role in was noted as early as 1863 by Rudolf Virchow pathogenesis has been demonstrated in several (177). This in turn are also prime examples of illnesses in which promotes neutrophil recruitment and fuels neutrophils play a salient role. chronic lung inflammation (172). tissues and organs but enzyme has two opposing activities. They of neutrophils operates at multiple levels. These models primarily used neutrophil-derived ROS have the potential to neutrophil depletion or adoptive transfer of initiate tumor formation by genotoxic stress wild-type neutrophils in leukotriene-deficient and induction of genomic instability. it has been proposed that animal models. These studies exemplify the complex production of IL-8 (183). tively inhibits only the aminopeptidase activity A host of autoimmune disorders belong to this of LTA4H. 179). Later. tobacco smoke selec- support disease progression in other illnesses. including production of angiogenic factors trophils and other immune cells discussed in (185). cytokines. unknown infiltration in the lungs of COPD patients whether all neutrophils are capable of adapting promotes tissue damage and organ dysfunc. In agreement with regulation cascades involving lipids. its although the clinical similarities between this primarily synovial hydrolase activity converts leukotriene A4 into mouse model and human RA are encouraging. however. impact cancer progression. severe inflammation causes leukotriene B4. mutagenesis in vivo is still lacking.

ANCA accelerate ROS-dependent accumulation of these precursors. Early some autoimmune and infectious diseases. MDSCs inhibit T cell proliferation by proteins: secreted by Moreover.org neutrophils underwent a complete reversal in their effect on CD8+ T cells.org • Neutrophil Functions 479 . Immunol. 191. given that release of to secrete the anti-inflammatory cytokine IL. whose growth MDSCs are strong producers of ROS. For personal use only. In Although neutrophils had long been suspected www. Inter- inhibition of TGF-β signaling led tumor. autoantibodies directed against anti- plasticity in their responses. Downloaded from www.annualreviews. Likewise. 192). In cancer and cycle of tissue damage and inflammation. Neutrophil talk with invariant NKT cells could counter products act as both targets and mediators of this response. fering with the release of MDSCs or using drug associated neutrophils to assume a heightened interventions to polarize neutrophil responses proinflammatory state. cycle culminating in organ damage (194. evidence currently supports the existence of Wegener’s granulomatosis is consistently as- different populations. leading to vironment. Differential Autoimmunity neutrophil responses were also demonstrated in Deregulated neutrophil cell death and/or by Harvard University on 09/10/13. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 suppression of the antitumor immune response human renal cell carcinoma. NOS activities. concentration in the tumor microenvironment could. vation or autoantibody titers (196). In this instance. 25% or more during inflammation cytotoxic arsenal at tumor cells. Furthermore. These alternatively activated sent novel therapeutic strategies against cancer. MDSCs are found in the bone degranulation and release of chemoattractants marrow. serving to activate rather than suppress these cells. promoting individuals. a melanoma study. called myeloid-derived ANCA bind MPO and PR3 expressed on the suppressor cells (MDSCs) (189).30:459-489. In healthy surface of activated neutrophils. which they usually help to fuel. affecting multiple tissues and organs. Cross trophils can trigger organ damage. which also inhibited T cell responses. of monocytic and granulocytic cells (189). MD. face markers as do activated neutrophils (190. it is likely that neutrophil sociated with the presence of ANCA. Thus. which act as neutrophil apoptosis. 187). 2012. MDSCs have (119. Rev. Although little gens present in the cytoplasm of neutrophils. the extent of organ damage in patients than previously thought. Further- responses are more flexible and less stereotyped more. tibodies produced in SLE are predominantly although they consist of variable proportions either ANCA or directed against chromatin. and in SLE is another chronic autoimmune disease mice. in an inflammatory en- differentiation is partially blocked. Annu. Acute-phase trophils and confirmed their tumorigenic role. they are typically detected using the neu. increased clearance often accompanies autoimmune syn- systemic levels of the acute-phase protein dromes (193–195) and may play a major role serum amyloid A (SAA-1) induced neutrophils in disease pathogenesis. under cer. investigation of gets of antineutrophil cytoplasmic antibodies neutrophils in cancer has revealed considerable (ANCA). suggesting a feed-forward powerful suppressors of T cell functions. 195). mature myeloid cells. both of which use this amino plasma changes by tain circumstances. causing a reduction in in the tumor microenvironment could repre- tumor growth. with Wegener’s granulomatosis correlates with Another major mechanism of tumor escape the PMN infiltrate rather than with traditional from immune control has recently been autoimmunity parameters such as T cell acti- attributed to a heterogeneous category of im. SCs have characteristics of neutrophils. be induced to target their acid as a substrate (189. which together lead to a vicious neutrophils and monocytes. identical morphology and express the same sur- Fridlender and colleagues (187) depleted neu. restoring a proinflammatory autoimmunity.annualreviews. proteolytic and cytotoxic molecules from neu- 10. where they differentiate into mature and ROS. Using the anti-Ly6G antibody. the study showed that neutrophils in limiting L-arginine availability via arginase and liver. Pharmacological suppresses T cell responses (119. reports also suggest that. MPO and PR3 are the main tar- activation status (188). 191). Autoan- trophil surface markers CD11b+ and Gr-1+ . 192).

studies have reported the presence of a particu. PMNs also characteristics of immature neutrophils with network with many other immune cells and nonsegmented nuclei and higher expression help regulate the initiation of specific T and of MPO. pursuing new therapies. their role in SLE patho. These microbe exterminators of the immune system. Rev. trolled neutrophil responses can exacerbate and An increased capacity to form NETs and a even cause autoimmune and inflammatory dis- heightened cytotoxicity toward endothelial eases. antiribonucleoprotein. mation. 106). which degrades NETs in will better define neutrophils’ role in defense vitro. 36% exhibited can potentially provide insights into several either elevated titers of autoantibodies directed unique aspects of basic cell biology. a central cytokine in SLE genesis remained elusive. ponents or if they contribute to autoreactive B vessels It was proposed that TNF-α and IFN-α cell activation. exciting times await the know that NETs induce plasmacytoid DCs humble neutrophil. their timely removal may deploying their armamentaria? How do they be an essential mechanism for maintaining kill microbes? How specific are their instruc- tissue homeostasis. Should anti-PR3. Many challenges remain in understand- cells could bestow them with pathogenic ing neutrophil function: Is there specialization properties in lupus (105). They are endowed with antimicrobial lar subset of neutrophils in PBMC preparations mechanisms that make them the preeminent from pediatric and adult SLE patients. . 104. role of NETs may provide critical insights into Thus. 2012. Their strik- against NET components or inhibitors of ingly short life spans make them excellent mod- DNase I. Notably. NE. above) (197. understanding the or anti-LL-37 autoantibodies (103. Moreover. high levels of inflammatory cytokines in the role of microbial infections as a trigger of autoimmune patients are believed to sensitize autoimmunity. a familial form of SLE is linked and disease and will provide a rational path for to a mutation in DNase I (199). neutrophils to NETosis. In short.org role for NETs in autoimmune pathology was Neutrophils are specialized phagocytes that obtained when NETs were identified in renal arose as an evolutionary adaptation in verte- and/or skin biopsies from patients with SLE brates to coordinate and execute one of the most and small vessel vasculitis (103–106). For personal use only. However. nucleus is a feat of higher-order nuclear Can it be that NETs play a general role architecture that is just beginning to yield in modulation of autoimmune responses? We its secrets. whereas autoanti- bodies may trigger a switch from apoptosis to NETosis. impaired reliance on ROS as biochemical effectors may NET degradation correlates with development reveal novel ways for relaying intracellular of lupus nephritis.30:459-489. Furthermore. neutrophils do not be related to the MDSCs discussed previously always act in ways beneficial to the host: Uncon- (see section on Cancer. 480 Amulic et al. The recent discovery pathogenesis (103. both of which may protect NETs els for investigating cell death. low-density granulocytes display phenotypic In addition to this important role. Additional evidence suggesting a CONCLUDING REMARKS Annu. whereas their from degradation. Immunol. it remains to of a link between SLE and NET formation be determined if DCs can present NET com- Vasculitis: inflammation of blood has helped to shed light on this quandary.annualreviews. 104). It is also possible that NETs are prime cells for NET formation in response to involved in other autoimmune diseases. anti-HNP. among PMNs? Are they more plastic than we Because NETs appear to be formed during suspect? How do they make decisions before autoimmune disease. and they may B cell immunity. this prove to be the case. Human serum contains the tions to other cells? Answering these questions nuclease DNase I. Most notably. However. to produce IFN-α. one of the most severe signals. and defensin-3. neutrophils in a cohort of SLE patients. Downloaded from www. The uniquely lobulated neutrophil manifestations of SLE (102). 198). Several fundamental physiological responses: inflam- by Harvard University on 09/10/13. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 to be causative agents.

Ann.P. E-selectin engages PSGL-1 and CD44 through a common signaling pathway to induce integrin αLβ2-mediated slow leukocyte rolling. Blood 88:3259–87 10. 277:20887–94 17. Klopocki AG.annualreviews. Wang D. Thompson DA. Yago T. Clin. Nature 379:266–69 8. 1846. Chew TW. Cowland JB. von Andrian UH. Kiosses WB. Microscopic observation on the perforation of capillaries by the corpuscles of blood and the origin of mucus and pus globules. Methodologische Beitr¨age zur Physiologie und Pathologie der verschiedenen Formen Annu. Downloaded from www. Cell Biol. Klin. Finger EB. Getting to the site of inflammation: the leukocyte adhesion cascade updated. 6:173–82 5. Xu J. Immunity 33:657–70 6. Piccio L. J. For personal use only. 1:553–60 2.30:459-489. memberships. Han J. Kunkel EJ. Cell Biol. Mueller H. Butcher EC. Biol. is supported by an EMBO Long-Term Fellowship. Blood 89:3503–21 7. 2000. Tyrosine kinase Btk regulates E-selectin-mediated integrin activation and neutrophil recruitment by controlling phospholipase C (PLC) γ2 and PI3Kγ pathways. Nat. 173:6403– 8 www. J. Chem. Nourshargh S. Stadtmann A. Goldfinger LE. Shao B. Mag. 136:717–27 9. Immunol. G. 2007. Sci.annualreviews. A fragment of paxillin binds the α4 integrin cytoplasmic domain (tail) and selectively inhibits α4-mediated cell migration. Blood 116:485–94 12. Rev. Hedrick J. 162:731–41 18. Wang F. J. J. 1998. Van Aken H. Puri KD. Siani MA. Chemokines trigger immediate β2 integrin affinity and mobility changes: differential regulation and roles in lymphocyte arrest under flow. Z. Springer TA. Cell 114:201–14 19. et al. Salgia R. IY30CH19-Zychlinsky ARI 17 February 2012 13:38 DISCLOSURE STATEMENT The authors are not aware of any affiliations. Ginsberg MH. Ley K. Divergent signals and cytoskeletal assemblies regulate self-organizing polarity in neutrophils. is an Alexander von Humboldt Foundation Scholar. Majeed M. Giagulli C. Herzmark P. 2003. Borregaard N. Schenkel AR.org der Leukocyten. Neutrophils. Pasteur 7:348–57 by Harvard University on 09/10/13. 2010. Yao L. Howe AK. Blood 115:3118–27 13. 4. Miner JJ. Investig. McEver RP. Van Keymeulen A. Kim JY. Nat. Campbell JJ. Waller A. 2004. 1996. Selectins and their ligands: current concepts and controversies. et al. Immunol. Edinburgh Philos. 7:678–89 14. or financial holdings that might be perceived as affecting the objectivity of this review. Nathan C. Lawrence MB. 2010. Muller WA. et al. J. Zlotnik A. Rose DM. Straight A. Cummings RD. Slepak M. Alon R. et al. Kansas GS.org • Neutrophil Functions 481 . 1880. 2:271–80 3. Chemokines and the arrest of lymphocytes rolling under flow conditions. Lecon sur la pathologie comparee de inflammation. Inst. from marrow to microbes. Lawrence MB. Platelet endothelial cell adhesion molecule deficiency or blockade significantly reduces leukocyte emigration in a majority of mouse strains. Kiosses WB. 1996. ACKNOWLEDGMENTS We thank Diane Schad for assistance with graphic design and Cornelia Heinz for administrative help. and B. Immunol. Threshold levels of fluid shear promote leukocyte adhesion through selectins (CD62L. Hirsch E. Lond. 1997.H. Ehrlich P. 2003. Borregaard N. 2006. 2002. 1893. 2010. 2012. Rev. 100:S97–103 11.E). Liu S. LITERATURE CITED 1. 1997. Med. J. Metchnikoff E. Immunol. Laudanna C. Rev. Science 279:381–84 15.A. Cybulsky MI. et al. Ley K. Constantin G. Spatial restriction of α4 inte- grin phosphorylation regulates lamellipodial stability and α4β1-dependent cell migration. Kansas GS. Adhesion through L-selectin requires a threshold hydrodynamic shear. Role of PSGL-1 binding to selectins in leukocyte recruitment. 1997. Neutrophils and immunity: challenges and opportunities. funding. Granules of the human neutrophilic polymorphonuclear leukocyte. Immunity 13:759–69 16.

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and Kristin A. IY30-Frontmatter ARI 17 February 2012 11:21 Annual Review of Immunology Contents Volume 30. 2012 Decisions About Dendritic Cells: Past. Horton p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p39 Sphingosine-1-Phosphate and Lymphocyte Egress from Lymphoid Organs Jason G. Nathanael McCurley. Immunol. Douek p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 149 The Response to and Repair of RAG-Mediated DNA Double-Strand Breaks Beth A.30:459-489. Rao. Cyster and Susan R. Jameson. Stephen C. Hogquist p p p p p p p p p p p p p p p p p p p p p p p95 Adaptive Immunity to Fungi Marcel Wuthrich. Masanori Kasahara. Powell. 2012. Jr. ¨ George S. Kristen N. Rev. Cooper p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 203 Immune Regulatory Function of B Cells Claudia Mauri and Anneleen Bosma p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 221 Lung Dendritic Cells in Respiratory Viral Infection and Asthma: From Protection to Immunopathology Bart N.annualreviews. Helmink and Barry P. and Future Ralph M. Schwab p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p69 Selection of Self-Reactive T Cells in the Thymus Gretta L. Lambrecht and Hamida Hammad p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 243 Tolerance of Infections Janelle S. O’Connell. and David Baltimore p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 295 ix .. Emily B. Present. and Max D. by Harvard University on 09/10/13. For personal use only. Heikamp. Sleckman p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 175 VLR-Based Adaptive Immunity Thomas Boehm. and Maureen R. Deepe. Michael Schorpp.org Fritz Melchers p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p23 Regulation of Immune Responses by mTOR Jonathan D. Schneider p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 271 microRNA Regulation of Inflammatory Responses Ryan M. Ayres and David S. Yoichi Sutoh. Steinman p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 1 The Basel Institute for Immunology Annu. Brenchley and Daniel C. Pollizzi. Downloaded from www. Stritesky. and Bruce Klein p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 115 Microbial Translocation Across the GI Tract Jason M. Dinesh S.

and Alexander Y. Hayes. IY30-Frontmatter ARI 17 February 2012 11:21 Reflex Principles of Immunological Homeostasis Ulf Andersson and Kevin J.30:459-489. Ryan M. Xavier p p p p p p p 611 Innate Lymphoid Cells: Emerging Insights in Development. Nolte. Young. Signaling by Myeloid C-Type Lectin Receptors in Immunity and Homeostasis David Sancho and Caetano Reis e Sousa p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 491 Regulatory T Cells: Mechanisms of Differentiation and Function Steven Z. Kiyoshi Hirahara. and Hamid Mattoo p p p p p p p p p p p p p 357 Monogenic Autoimmunity Mickie H. and Cornelis Murre p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 337 Siglecs and Immune Regulation Shiv Pillai. Staudt p p p p p p p p p p p p p p p p p p p p p p p p p p p 565 Autophagy and the Immune System Petric Kuballa. Kathleen D. Rev. and Function Hergen Spits and Tom Cupedo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 647 Cancer and Inflammation: An Old Intuition with Rapidly Evolving New Concepts Giorgio Trinchieri p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 677 Transcriptional and Epigenetic Control of T Helper Cell Specification: Molecular Mechanisms Underlying Commitment and Plasticity Yuka Kanno. O’Shea p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 707 Induced CD4+ Foxp3+ Regulatory T Cells in Immune Tolerance Angelina M. Lafaille p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 733 The Microbiome in Infectious Disease and Inflammation Kenya Honda and Dan R. Nussenzweig p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 429 Annu. Bilate and Juan J. For personal use only. Metzler. Annaiah Cariappa. Adam B.annualreviews. Lineage Relationships. and Louis M. Ilka Arun Netravali. Li-Fan Lu. Christel Cazalet. Shaffer III. Rudensky p p p p p p p p p p p p p p p p p p p p p p p p p p 531 Pathogenesis of Human B Cell Lymphomas Arthur L. Victora and Michel C. Immunol. and Ramnik J. Downloaded from www. Anderson p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 393 Germinal Centers Gabriel D. Tracey p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 313 Chromatin Topology and the Regulation of Antigen Receptor Assembly Claudia Bossen. 2012.org Neutrophil Function: From Mechanisms to Disease Borko Amulic. Golnaz Vahedi. Robert Mansson. Garret L. Whitney M. Castoreno. Josefowicz. Kentner Singleton. and John J. Littman p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 759 x Contents . and Arturo Zychlinsky p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 459 by Harvard University on 09/10/13. Cheng and Mark S.