Multiple Sclerosis (1998) 4, 16 ± 21

 1998 Stockton Press All rights reserved 1352 ± 4585/98 $12.00

Myelin basic protein in cerebrospinal ¯uid and other body ¯uids

John N Whitaker
Department of Neurology, University of Alabama at Birmingham and the Neurology and Research Services of the
Birmingham Veterans Medical Center, Birmingham, Alabama, 35294-0007, USA.

Myelin basic protein (MBP) or a fragment thereof may enter cerebrospinal ¯uid (CSF) and other body ¯uids in an etiologically nonspeci®c fashion
to provide information about the status of central nervous system (CNS) myelin damage. MBP immunochemically detected is referred to as MBP-
like material (MBPLM). The clinical utility of the assay for MBPLM in CSF is to document the presence, continuation, or resolution of CNS myelin
injury. The analysis of CSF for MBPLM is subject to many variables, among which are the antisera and the form of the assay utilized. The
dominant epitope of CSF MBPLM is in the decapeptide of 80 ± 89 from the intact MBP molecule of 170 residues. Normally, CSF has no detected
MBPLM. Following an acute relapse of MS, MBPLM rises quickly in the range of ng/ml and rapidly declines and disappears. The presence of
MBPLM in CSF in chronic and progressive phases of the disease is unusual, but it may sometimes be detected in low levels, depending on the
assay used for detection. The level of CSF MBPLM is related to both the mass of CNS myelin damage and how recently it occurred. The level of
CSF MBPLM rarely is elevated in optic neuritis. The level of CSF MBPLM is unrelated to CSF protein level, level of IgG, presence of oligoclonal
bands or pleocytosis. CSF MBPLM has the potential of serving as a marker of therapeutic effectiveness in MS and does have predictive value for
response to glucocorticoids given for worsening of disease. The detection of MBPLM in body ¯uids other than CSF would be of great value because
of the resulting improved feasibility for objectively monitoring the natural history of MS and response to therapy. Studies on blood have yet to
produce a valid assay of MBPLM. Urinary MBPLM, though different in its features from that in CSF, may provide a correlate, not with acute
demyelination in MS as is the case for CSF, but with progression of disease.
Keywords: cerebrospinal ¯uid; urine; myelin basic protein; multiple sclerosis; myelin

Introduction
One of the most critical needs for improving the
management of multiple sclerosis (MS) is an objective
and feasible test, or surrogate marker, for monitoring
disease activity or disease status.1,2 Such a test would
permit the determination of success or failure of
experimental treatment as soon as possible and later
to use such treatment accurately. The need for such a
procedure is evident, but the approach to be taken is
less clear. The current options are neuro-imaging or
quantitation of molecules in body ¯uids arising from
either the primary demyelination or in¯ammation in
the central nervous system (CNS). In spite of the
enormous power of cranial MRI in providing informa-
tion about multifocality and disease activity in MS,
especially where unsuspected,3,4 its uncertain role in
predicting changes in patient functioning,5 its limited
assessment of the spinal cord where so many small
and disabling lesions are located, its expense and the
impracticality of repeated neuro-imaging all lessen its
feasibility as a clinical monitor. Associated with the
breakdown of the CNS myelin sheath in MS,
components of CNS myelin, speci®cally MBP or its
peptides and, possibly, peptides of other myelin
proteins may enter CSF, urine, and blood. MBPLM in
CSF is a reliable indicator of myelin damage in MS.6,7
While other substances, particularly cytokines,8 solu-
ble cytokine receptors9,10 and shed adhesion mole-
cules11 have been sought as better markers of disease
Correspondence: JN Whitaker
Presented in part at the European Charcot Foundation Sym-
posium at the 2nd Congress of the European Federation of
Neurological Societies, Rome, Italy, October 31, 1996
activity or pattern in MS, none has replaced a CNS
myelin constituent as a laboratory test for CNS myelin
damage. Furthermore, the interpretation of levels of
cytokines, cytokine receptors and adhesion molecules
is dif®cult because of their potential origins from
blood, in¯ammatory cells or activated glia.
Selected features of MBP
MBP accounts for 30% of CNS myelin proteins,12 is
encoded by a single gene of seven exons located on
chromosome 1813 and is normally expressed only by
oligodendrocytes and Schwann cells.14,15 Alternate
splicing of the MBP transcript gives rise to at least
four isoforms of human MBP with the major ones
having molecular weights of 21.5 kd (encoded by all
seven exons) and 18.5 kd (encoded by exons 1 and 3 ±
7).15 The 18.5 kd isoform containing 170 amino acid
residues dominates in adult human CNS myelin. In
addition to multiple isoforms, MBP may have a
number of post-translational modi®cations,16 such as
amino-terminus acylation,17 selected phosphorylation
of serine and threonine,18,19 deamidation of glutamine,
loss of C-terminal arginine19 ± 21 and methylation of the
arginine at residue 10722 that result in charged isomers.
Several recent observations have demonstrated
further complexities in the structure of MBP. First is
its gene structure. The human MBP gene had
previously been reported to be 45 kb23 or 32 ± 34 kb24
with a cDNA of 2.2 kb. The murine, and presumably
the human, MBP gene actually comprises exons 5 ± 11
of a more complex gene, designated as Golli (Gene of
oligodendrocyte lineage)-mbp.25 The 105 kb Golli-mbp
gene contains 11 exons and a 5' portion of 73 kb. The

amino portion of MBP encoded by exon 1 of MBP, or
exon 5 of the Golli-mbp gene, is likely to be
synthesized from Golli-mbp transcripts.25 The Golli-
mbp gene is located, at least in part, in lymphoid
tissue.25,26 The Golli-mbp can be recognized by human
T cells reactive with MBP.27 Second is the amino-
terminus acylation which may include fatty acids of
up to eight or more carbons.28 Third is the citrullina-
tion of six of the 19 arginine residues present in MBP
which results in a less basic MBP.29 Citrullinated MBP
is referred to as C8. MBP-C8 is formed early in
myelinogenesis,30 is relatively well preserved in MS
brain tissue and is immunogenic for humoral31 and
cellular32 responses.
MBP has multiple independent epitopes throughout
its 170 residues,33,34 and a multideterminant model is
likely34. Certain regions of MBP appear to have special
relevance to MS: one of these is residues 80 ± 100
(Table 1). There is evidence for intramolecular folding
and b-sheet structure in this region.35 Antibodies to
MBP in CSF36,37 and brain extracts38 frequently react
with this MBP region. Moreover, MBP peptide 87 ± 99
contains the encephalitogenic epitopes for the SJL
mice39 ± 41 and a minor I-E restricted encephalitogenic
epitope in the Lewis rat.42 This is also a prominent
region recognized by MBP-sensitive T cells in CSF43 or
lines44 derived from MS patients. Antibodies in CSF of
MS patients recognize a similar epitope of MBP.45 The
®ne speci®cities of epitopes bound by polyclonal46 and
monoclonal47 antibodies to MBP implicate three or
more epitopes, two of which are cryptic, in MBP
peptide 80 ± 89. Surprisingly, mAbs reactive with
either MBP peptide 80 ± 89 and acetyl-1-9 express a
cross-reactive idiotope present on their kappa light
chains.48
MBPLM in CSF (Table 2)
The presence of material that was cross-reactive with
antibodies to MBP6 or a MBP peptide7 and appearing
in CSF after acute CNS myelin damage was ®rst noted
20 years ago. A number of subsequent reports49 have
con®rmed these initial observations. MBPLM in CSF is
not disease speci®c but appears during active demye-
lination or more generalized CNS tissue injury
accompanied by myelin damage. Patients with stable
or chronic progressive phases of MS and those with
demyelinating diseases of the peripheral nervous
system rarely or never have MBPLM in CSF.50 CSF
Table 1 Important roles for MBP peptide 80 ± 100
. Encephalitogenic determinants
SJL mouse39 ± 41
Lewis rat (I ± E)42
. Region of dominant epitope
CSF antibody to MBP in MS45
MS brain antibody to MBP45
MS T cell clone recognition44
Cross-reactive with Golli-MBP27
CSF MBPLM51
Urine MBPLM58,80
MBP in cerebrospinal fluid
JN Whitaker
17
MBPLM in MS becomes undetectable 10 ± 14 days
following CNS myelin damage.7,50 CSF MBPLM is
larger in size in patients with strokes51 and head
injury52 than MS in relapse and follows a different
temporal pro®le.50 Patients with optic neuritis rarely
have an elevation of CSF MBPLM.53
Antisera to MBP are highly variable as to whether
they will detect CSF MBPLM at all and to what level.54
All current detection systems for CSF MBPLM use RIA
and polyclonal reagents. A suf®ciently sensitive ELISA
or mAb-based assay for CSF MBPLM has not yet been
developed. As the result of a series of investigations of
human CSF,7,46,50,55,56 the immunochemical features of
MBP and its peptides33,55,57 and the correct primary
sequence of human MBP,58,59 there is compelling
evidence that MBPLM in CSF contains an epitope
present in the residues of Thr-Gln-Asp-Glu-Asn-Pro-
Val-Val-His-Phe of MBP 80 ± 89 that is noncryptic and
exposed in intact MBP. The additional presence of
some or all of the residues of Phe-Lys-Asn-Ile at
positions 90 ± 93 seems likely in order to promote the
formation of a noncryptic epitope in MBP peptide 80 ±
89.60,61 This probability led to the recent use of human
MBP peptide 78 ± 93 which was the ®rst MBP peptide
to successfully stimulate the formation of an antibody
reactive with CSF MBPLM.62 Studied at neutral pH,
CSF MBPLM from MS patients has a size of greater
than 30 kd.63 At pH 3 in aqueous solutions it is much
smaller, possibly in the 1 ± 3 kd range.51,63 The larger
size of CSF MBPLM at neutral pH could be due to the
formation of complexes with antibody to MBP.51,64
However, the size of the CSF MBPLM of under
100 kd63 makes this less likely.
Metabolic fate of MBPLM in CSF
Little information exists on the metabolic fate of
MBPLM in CSF except that it is rapidly cleared.7
Similar observations are derived from experiments
with rats.65 The MBPLM must at some point enter
blood, but the validity of its presence in blood in
humans has been dif®cult to ascertain because of
technical problems in its detection due to the presence
of interfering substances,66 to binding by serum
proteins67,68 and to proteinases69 which may degrade
radioligand or MBPLM. In the rabbit, MBPLM can be
detected in plasma after subcutaneous injection in
Table 2 Features of MBPLM in CSF
. Assay reagents critical
No nonimmune validation yet
Dominant epitope is MBP peptide 80 ± 89
Conformation and MBP peptide 78 ± 93
. Disease-nonspeci®c index of acute CNS
myelin damage
. Unrelated to other CSF measurements
. Documents recent demyelination
. Predictor of response of MS to steroids
. Bound to larger molecule (430 KD) pH 7
. May be bound as immune complexes
. Range of sizes pH 5
 MBP in cerebrospinal fluid
JN Whitaker
18
adjuvant and during the immune response to it.70 correlate with acute CNS myelin injury in MS as does
When infused into rabbits, human MBP peptide 45 ± the CSF MBPLM, it could be an indicator or predictor
89 is rapidly cleared (half-life of 50 ± 60 min) from of transition to a chronic progressive phase. In the
blood and catabolized by the kidney.71 Human and Phase 3 trial of interferon beta-1b (IFNb) in relapsing-
animal kidney both contain proteinases capable of remitting MS,5,79 urine specimens were analyzed from
degrading MBP peptide 45 ± 89 and generating smaller a cohort of study patients undergoing serial cranial
peptides.72,73 MRI at 6 week or yearly intervals. Those patients
converting from a relapsing-remitting to a secondary
progressive course were signi®cantly more likely to
MBPLM in urine have an elevated level of MBPLM.80 Furthermore, the
The detection of MBPLM in urine has required the elevations of MBPLM correlated with the number and
recognition of a cryptic epitope in MBP peptide 80 ± volume of lesions seen on T2-weighted cranial MRI's
89,60,61,74 the correct primary sequence of residues of (Table 5).
Glu-Asn rather than Gln-Asp at positions 83 and 84 of To summarize (Table 3), the MBPLM in CSF must
MBP58 and the availability of a highly selected contain the same noncryptic (i.e., exposed) epitope
antiserum to measure it.61 This has been a dif®cult found in human MBP peptide 80 ± 89 and in intact
and time-consuming process.75 The clinical utility of MBP. While the precise chemical nature of CSF
MBPLM has been limited by the lack of information on MBPLM has not been determined, it seems likely that
the structure of urinary MBPLM. It has been possible in MS it is noncovalently attached to another
to prepare a mAb (Fusion 41) that reacted with urinary molecule(s). The urinary MBPLM is immunochemi-
MBPLM.62 This success was a result of the use of MS- cally expressed as a different and cryptic (i.e.,
patient urine in the screening step to select fusion
products. Immunochemical characterization of F41 has
revealed, unexpectedly, that it recognized a cryptic
MBP epitope in peptide 80 ± 85. The results indicate Table 3 MBPLM in body ¯uids
that MBP peptide 80 ± 89 has two cryptic, or
unexposed, epitopes, one in residues 83 ± 89 recog- CSF Urine
nized by polyclonal antibody R11061 and a second in
Size 430 kd 51 kd
residues 80 ± 85 recognized by mAb F41. Epitope 80 ± 89 84 ± 89
cryptic No Yes
Correlation of the level of MBPLM in CSF and urine Presence
Normal No Yes
in MS patients receiving glucocorticoid and immuno- :: MS Acute Chronic
suppressive treatment
CSF MBPLM56 levels were measured in MS patients,
who were experiencing a clinical worsening, given a
regimen of intravenous methylprednisolone followed
by a tapering dose of oral prednisone. Those MS Table 4 Urinary MBPLM-MS correlates78
patients with an elevated level of CSF MBPLM were
more likely to respond to the treatment protocol MBPLM/creatinine
indicating that tests for CSF MBPLM can be used not NG/MG
only as a monitor of CNS myelin damage6,7 but also as
Controls 87.2 (38.5)
a predictor of a bene®cial response.76 The degree of MS RR 100.8 (73.3)
disability, as determined by the Kurtzke score,77 and MS SP 357.7 (15.7)
the CNS site of new clinical de®cit accompanying the
exacerbation did not affect the therapeutic response. No correlation with active disease clinically, on MRI or by
Thus, even though not feasible for serial sampling, CSF
CSF can furnish predictive information about response
to certain treatment.
A study of MS patients has recently been completed
in which the level of urinary MBPLM was correlated Table 5 Correlations of urinary MBPLM and disease status
with clinical ®ndings, levels of CSF MBPLM and of multiple sclerosis80
changes on serial cranial MRI.78 Successful completion
MBPLM/creatinine
of this large study required revalidation of the RIA, Clinical EDSS P50.01
including a change in RIA standard, and studies in a RR ? SP P50.005
clinical research unit and of normals and patients with Cranial MRI lesion number P50.05
other neurological diseases. Urinary MBPLM does not Volume P50.005
parallel acute myelin damage but appears to re¯ect an New (7) P50.005
ongoing process, possibly linked to attempted efforts at Confounding in¯uence of creatinine
remyelination. This is consistent with the highest MBPLM only RR ? RR 129+81
values of MBPLM in the urines of patients with (P = 0.0042) RR ? SP 198+60
secondary chronic progressive MS (Tables 3 and 4). MBPLM/creatinine P = 0.075
Although the level of urinary MBPLM does not

unexposed in the intact molecule) epitope from the
same general region of the MBP peptide 80 ± 89. The
urinary MBPLM is not attached to another molecule
but may represent a small MBP peptide released in a
chemically modi®ed form or modi®ed subsequently
during its catabolism and clearance. Not only do the
MBPLMs in CSF and urine differ in size and epitope,
they also re¯ect different clinical events in the patient
with MS. CSF MBPLM appears and disappears in a
predictable way after acute CNS myelin damage6,7 and
correlates with changes on cranial MRI.81 In contrast,
urinary MBPLM is found in normal urine, ¯uctuates
widely with no apparent relationship to acute clinical
change, diet or renal function, shows no relationship
to changes on cranial MRI or to CSF MBPLM but
correlates best with the clinical pattern of chronic
progressive MS.
Acknowledgements
Ms Linda Brent and Mr R David Kachelhofer provided
excellent assistance in the preparation of the manu-
script. The research summarized from the author's
laboratory was supported by the National Institutes of
Health (NS 23240), the National Multiple Sclerosis
Society (RG 2667-A-7), the Thompson Research Fund
for Multiple Sclerosis, Berlex Laboratories, and the
Research Program of the Veterans Administration.
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