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Polyclonally triggered B cells in the peripheral blood and bone marrow of normal individuals and in patients with systemic lupus erythematosus and primary sjgren's syndrome.

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ARTHRITIS
577
&
RHEUMATISM
POLYCLONALLY TRIGGERED B CELLS IN
THE PERIPHERAL BLOOD AND BONE MARROW OF
NORMAL INDIVIDUALS AND IN PATIENTS WITH
SYSTEMIC LUPUS ERYTHEMATOSUS AND
PRIMARY SJOGREN’S SYNDROME
ANTHONY S. FAUCI and HARALAMPOS M. MOUTSOPOULOS
Numbers of B cells spontaneously secreting Ig
(IgC, IgA, and IgM) were determined by a plaque-forming cell (PFC) assay simultaneously in the peripheral
blood and bone marrow of normal individuals, patients
with systemic lupus erythematosus (SLE), and patients
with primary Sjogren’s syndrome. Normal individuals
had 382 (f89) PFC per 106 mononuclear cells in peripheral blood. Patients with either active or inactive Sjcigren’s syndrome had normal numbers of spontaneous
Ig-secreting cells in peripheral blood (P > 0.2). Conversely, patients with inactive as well as active SLE had
markedly increased spontaneous PFC (P < 0.05 and
P c 0.001, respectively). Patients with active SLE had
significantly greater PFC than patients with inactive
SLE: 3,984 (+ 960)versus 1,605 (+ 527) PFC per 106
mononuclear cells (P < 0.05). The lack of increased
numbers of activated B cells in the blood of patients
with Sjogren’s syndrome was not explained by a preferential sequestration of activated B cells in the bone marrow. However, of particular interest was the finding that
the bone marrow served not only as a major source of
virgin B cells but as a lymphoid organ of either in situ
- . . - .-- .
From the Laboratory of Immunoregulation. National Institute of Allergy and Infectious Diseases, National Institutes of Health.
Bethesda, Maryland 20205 and the Department of Medicine, University of Ioannina Medical School, loannina, Greece.
Address reprint requests to Anthony F. Fauci, MD, Building
10. Room 118-13, National Institutes of Health, Bethesda, M D 20205.
Submitted for publication May 7, 1980 accepted in revised
form December 8, 1980.
Arthritis and Rheumatism, Vol. 24, No. 4 (April 1981)
activation of B cells or sequestration for activated B
cells. Normal individuals had approximately a 20-fold
relative increase of activated B cells per 106 mononuclear cells in the bone marrow compared to peripheral
blood, while patients with inactive and active SLE both
had a 35-fold relative increase in activated B cells in
bone marrow compared to peripheral blood. The potential relevance of circulating activated B cells and their
sequestration in lymphoid organs is discussed concerning the discrepancy in this regard between Sjogren’s syndrome and SLE, and our understanding of the
significance of polyclonal B cell activation in the pathogenesis of these diseases.
Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by bone marrow-derived (B) cell hyperreactivity with hypergammaglobulinemia and autoantibody production (1-5). In
addition, abnormalities of thymus-derived (T) cell subsets, particularly a deficiency of suppressor T cells, have
been reported in a number of studies (reviewed in reference 5). This association of hyperactive B cells and irnmunoregulatory T cell abnormalities is strongly analogous to the murine models of SLE (6,7). It has been
unclear whether the B cell hyperreactivity in SLE is independent from and actually may precede the T cell abnormalities or whether the B cell hyperreactivity results
from deficient suppressor cell activity. Of interest in this
regard is the fact that certain studies in the murine mod-
FAUCI AND MOUTSOPOULOS
578
els of SLE have demonstrated that the B cell abnormalities may be primary, even present from birth, and
antedating detectable T cell abnormalities (8- 13).
It has been demonstrated that peripheral blood
lymphocytes from patients with active SLE spontaneously secrete polyclonal antibody against multiple noncross-reacting antigenic determinants (14- 17). This is
similar to observations in the murine models (18,19).
There are certain similarities and overlap between SLE and Sjogren’s syndrome. Sjogren’s syndrome is also characterized by B cell hyperreactivity including hypergammaglobulinemia, non-organ specific
autoantibodies such as rheumatoid factor,. and antibodies to extractable nuclear antigens (20-23). In addition, circulating IgG immune complexes have been
demonstrated in certain patients with Sjogren’s syndrome (24). However, although patients with Sjogren’s
syndrome have reversible abnormalities in certain T cell
subsets such as T, cells, abnormalities of suppressor cell
function were not demonstrated (25). Interestingly, despite the clinical manifestations of B cell hyperreactivity
in Sjogren’s syndrome, polyclonally triggered B cells
were not demonstrated in the peripheral blood of these
patients (25). Since activated and antibody-secreting
cells not found in the peripheral blood may be sequestered in other organs, it was possible that the triggered B
cells in Sjogren’s syndrome were not circulating, but
were sequestered. The bone marrow contains large
numbers of B cells in both animals and humans (26-29).
Since the bone marrow can serve as both a source of B
cells in humans and a potential site of B cell sequestration, the present study examined the relative levels of B
cell activation simultaneously in the peripheral blood
and bone marrow of patients with SLE compared to patients with Sjogren’s syndrome in order to determine: l)
the relationship, if any, between polyclonally activated
B cells in the blood and bone marrow and 2) the possibility that activated B cells are sequestered in the bone
marrow of patients with Sjogren’s syndrome.
MATERIALS AND METHODS
Patients. Twelve patients with SLE and 15 with primary SjBgren’s syndrome were studied. Nine normal subjects
were studied as controls. At least I normal subject was studied
simultaneously on every occasion that patients (usually 3 per
experiment) were studied. The diagnosis of SLE was made according to the criteria of the American Rheumatism Association (30). The diagnosis of Sjogren’s syndrome was based on
the presence of at least two of the following clinical findings:
xerostomia (both decreased parotid flow rate and abnormal
parotid scintigraphy), keratoconjuflctivitis sicca by slit-lamp
examination, and recurrent parotid-gland enlargement. In all
patients, the diagnosis was confirmed by lip biopsy. All of the
Table 1. Clinical profile of patients with systemic lupus erythematosus
Laboratory datat
Patient
1
2
3
4
5
6
7
8
9
10
11
12
Clinical description
Clinical activity.
c 3 , jig%
Anti-DNA, %
Serositis; arthritis
Serositis; proliferative
glomerulonephritis; myositis;
arthritis
Serositis; Sjbgten’s syndrome
Fever; pancytopenia; proliferative
focal glomerulonephritis
Proliferative diffuse
glomerulonephritis
Serositis
Membranous glomerulonephritis;
nephrotic syndrome
Leukopenia; discoid lupus;
glomerulonephritis
Proliferative glomerulonephritis
Glomerulonephritis; discoid lupus;
serositis; arthritis; 1 month
status post splenectomy for
severe thrombocytopenia
Coombs positive anemia; erythema
multiforme
Arthritis; myositis; cutaneous
eruption; proliferative focal
glomerulonephritis
Inactive
Inactive
100
94
43
75
Inactive
Active
92
74
21
36
Inactive
92
20
Inactive
Active
107
90
32
15
Inactive
100
0
Inactive
Active
135
86
0
17
Active
90
70
Active
48
76
* Clinical activity as determined at the time the study was performed (see Materials and Methods).
t Normal values for C3 are 84-175
pg% and for anti-DNA 0-25%.
B CELL ACTIVATION IN SLE AND SJOGREN’S SYNDROME
579
Table 2. Clinical profile of patients with Sjogren’s syndrome
Laboratory datat
Patient
1
2
3
4
5
6
7
8
Clinical description
Vasculitis
Vasculitis; interstitial nephritis
Glandular
Pseudolymphoma
Hypergammaglobulinemicpurpura
Glandular
Hypergammaglobulinemic
purpura; interstitial nephritis
Chronic persistent hepatitis;
hypergammaglobulinemic
Clinical
activity’
Anti-DNA,
RF
C3,pg%
%
1:8,192
1:32
< I : 16
1 : 1,024
1 :32,768
1 : 16,384
1 : 1,024
150
149
166
89
114
110
74
0
0
0
8
2
0
4
Active
1 :32,768
110
25
Active
1 : 16,384
107
0
Inactive
1:512
-
120
115
0
0
1 :124
5
2
0
5
Inactive
Inactive
Inactive
Active
Active
Inactive
Active
purpura
9
10
11
12
13
14
15
Pseudolymphoma, recently
evolved into lymphoma
Glandular
Hypergammaglobulinemic
purpura
Glandular
Glandular
Hypergammaglobulinemic
purpura
Raynaud’s phenomenon
Active
Inactive
Inactive
Active
1:8,192
133
123
120
Active
1:512
142
1:512
* Clinical activity as determined at the time the study was performed (see Materials and Methods).
t RF = rheumatoid factor as determined bv the bentonite flocculation technique. Normal values: RF
=
< 1 : 16; C3 = 84-175 pg%; anti-DNA = 0125%.
patients had primary Sjogren’s syndrome, i.e., without other
associated connective tissue disease (21,3 1).
The clinical profiles of the patients with SLE and Sjogren’s syndrome are listed in Tables 1 and 2, respectively. SLE
was considered active when clinical findings such as arthritis,
nephritis, serositis, myositis, cerebritis, or widespread cutaneous lesions were associated with either low levels of the third
component of complement (C3) (32) or high levels of antibody to DNA (33). Five patients with Sjogren’s syndrome had
disease limited to the exocrine glands (glandular Sjogren’s
syndrome) and for purposes of the study will be considered as
having “inactive” disease (Table 2). Ten patients with Sjogren’s syndrome had extraglandular involvement such as interstitial nephritis, vasculitis, and pseudolymphoma (extraglandular Sjogren’s syndrome). These patients will be considered as having active disease except for patients no. 1 and 2
(Table 2) whose extraglandular manifestations were inactive
at the time of the study.
Bone marrow aspirates and peripheral blood suspensions. Bone marrow aspirates of 2 to 3 ml were obtained in
heparinized plastic syringes from the posterior superior iliac
crest as previously described in detail (27-29). The relative
proportion of peripheral blood lymphocytes contaminating
bone marrow lymphoid cells in such aspirates is minimal and
has previously been discussed in detail (27-29). Immediately
prior to obtaining the bone marrow aspirate, 60 ml of heparinized venous blood was drawn and with each individual, bone
marrow and blood specimens were subsequently assayed simultaneously.
Preparation of cell suspensions. Purified mononuclear
cell suspensions were obtained from peripheral blood samples
by the standard Hypaque-Ficoll gradient centrifugation
method. Bone marrow specimens were handled as follows:
Immediately upon aspiration, the specimens were expressed
from the syringe into 50 ml of cold RPMI-1640 media (Microbiological Associates, Walkersville, MD) contained in a conical plastic centrifuge tube. The specimen was vigorously vortexed and washed into a button 3 times. The red blood cells
(RBC) were then lysed by hypotonic lysis and the leukocyte
button resuspended, counted in a Coulter Counter (Model Fn,
Coulter Electronics, Hialeah, FL), and a cytocentrifuge preparation was made and stained with Wright’s stain. In this manner, the percentage of mononuclear cells in the specimen
could be determined even though the entire leukocyte suspension was subsequently assayed.
In previous studies on human bone marrow lymphocytes (27-29), purified mononuclear cells were obtained from
the bone marrow aspirates by a sucrose density gradient centrifugation method. In those studies, only mononuclear cells
were assayed and not the entire bone marrow leukocyte suspension. This was done because of concern over the possibility that the other leukocyte elements would affect the in
vitro responses of the bone marrow lymphocytes after several
days in culture. Since in the present study we were examining
spontaneous, unstimulated immunoglobulin (Ig) production
at 0 hours or immediately upon obtaining the specimens without a previous in vitro culture period, it was not necessary to
remove the other leukocytes, provided responses were corrected for the proportion of mononuclear cells in suspension.
In addition, we were concerned that the sucrose gradient technique might preferentially select certain B cell subsets out of
the mononuclear cell layer. In this regard, in the first series of
experiments, bone marrow specimens from individuals were
divided into two aliquots. In one aliquot, the RBC were lysed
and the entire leukocyte suspension was assayed in the other,
the cells were fractionated over sucrose gradients to obtain
purified mononuclear cells. In four of four experiments, there
was no difference in the number of spontaneous Ig-secreting
FAUCI AND MOUTSOPOULOS
5 80
cells per lo6 mononuclear cells between the two aliquots.
Thus, because of the relative ease of preparation, the former
approach was taken in subsequent experiments.
Detection of spontaneous Ig-secreting cells. The number of cells spontaneously secreting Ig of all classes was determined by a modification of a previously described plaqueforming cell (PFC) assay (34). Briefly, sheep RBC (SRBC)
targets were coated with staphylococcal protein A (SPA) by
the chromic chloride method (34). PFC were assayed by a
modification of our originally described ultra-thin layer hemolysis-in-gel technique (35,36). Briefly, 60 X 15 mm plastic
petri dishes were precoated with agarose in the following
manner: 1,400 mg agarose (Accurate Chemical, Hicksville,
NY) was added to 100 ml of sterile distilled H,O and boiled
until dissolved. This was then added to 100 ml of 2 X RPMI
which had been pre-warmed to 56°C. The mixture was kept at
56"C, and 4 ml of the agarose mixture was added to the petri
dishes. The dish was swirled quickly to cover the bottom of
the plate, cooled, and stored at 4°C until used.
Rabbit anti-human polyvalent IgG (Cappel Labs,
Dowington, PA) was diluted in RPMI-1640 to 150, 1:100, and
1:250. Guinea pig complement (absorbed with SRBC) was diluted 1:40 in veronol-buffered saline.
For the plaqueing procedure, mononuclear cells were
brought up in RPMI at an appropriate dilution to give approximately 50 to 100 PFC per plate. One-tenth milliliter of
cells together with 0.85 ml of agarose and 0.06 ml of a 25% solution of SPA-coated SRBC were added to a 12 x 75 mm tube
heated in a 47°C heating block. The tube was vortexed and
the contents of the entire tube poured into the agarose-precoated petri dish, swirled, cooled, and then kept in a 37°C incubator for 2 hours. One milliliter of media (control) or the
developing antiserum in various dilutions was then layered o n
the plate and incubated for an additional 2 hours. RPMI or
the developing antisera was removed, and 1.0 ml of guinea pig
complement was added. Incubation was continued for 1 hour
longer at 37"C, and PFC (which were visible macroscopically)
were read under a dissecting microscope. Data were expressed
as PFC per lo6 mononuclear cells that were plated. For bone
marrow specimens, a simple conversion factor was employed
to determine the number of PFC per 10' mononuclear cells
based on the proportion of mononuclear cells in the entire
suspension that was assayed. For peripheral blood, purified
mononuclear cells had been obtained by the Hypaque-Ficoll
method, and so essentially 100% mononuclear cells were assayed, negating the need for a conversion factor.
RESULTS
Peripheral blood lymphocytes and bone marrow
mononuclear cells. Table 3 lists the absolute numbers of
circulating lymphocytes in the peripheral blood of the
various patient groups compared to normal controls. In
addition, the relative proportions of mononuclear cells
in the bone marrow aspirates of the different groups are
listed. It should be pointed out that although patients
with active or inactive Sjogren's syndrome and active or
inactive SLE were, as groups, all significantly lymphopenic compared to normal controls, the patients with
Sjogren's syndrome, whether active or inactive, had percentages of mononuclear cells in their bone marrows
which were comparable to controls (Table 3). Conversely, patients with SLE had markedly reduced proportions of mononuclear cells in their bone marrow aspirates. This was especially true among the group with
inactive SLE ( P < 0.01, Student's t-test). The proportion
of bone marrow mononuclear cells in the group with active SLE was 5.5 (+ l.O)% compared to 13 (-+ 2.6)% in
normal controls. This , however, did not reach statistical
significance, perhaps due to the smaller sample of patients in this group.
It does not appear that corticosteroid therapy
was responsible for these bone marrow findings since
within the SLE group, all of whom were receiving variable doses of prednisone on a daily or alternate day
regimen, there was no correlation between magnitude of
dose and dose regimen of prednisone and proportions of
mononuclear cells in the bone marrow. Furthermore, of
the 2 patients with Sjogren's syndrome receiving corticosteroids, one had inactive disease (patient no. 2) and
the other had active disease (patient no. 7). Both patients were receiving 10 mg of prednisone on alternate
days. Patient no. 2 (inactive) had 18% and patient no. 7
(active) had 1% mononuclear cells in the bone marrow
aspirate. Of note is the fact that within the group with
the lowest proportion of bone marrow mononuclear
Table 3. Mononuclear cells in the peripheral blood and bone marrow aspirates of patients with systemic
lupus erythematosus (SLE) and Sjogren's syndrome and normal controls
Peripheral blood
lymphocytes,
cells per mm3*
Subjects (number)
~
I
_
2,389 (rt 247)
-
cells in bone
marrow aspirate
___
13 (k 2 6)
1,521 (rt 172)
t o 02
14 (rt I 8)
l,314(rt 112)
1,031 (rt 244)
1,295 (rt 361)
<o 01
t o 01
10 (f2 2)
3 ( + I 1)
<O 05
55(+10)
_
P value?
P value?
- .-
-___-___I___
Controls (10)
Inactive Sjogren's
syndrome (7)
Active Sjogren's
syndrome (8)
Inactive SLE (7)
Active SLE (5)
* Data represent mean
% mononuclear
(-f
___--
-_
-
>o 2
>o 2
<o 01
__
I _ _ _
SEM)
1. P value represents comparison of each patient data point with normal controls
-0 1
B CELL ACTIVATION IN SLE AND SJOGREN'S SYNDROME
cells (inactive SLE), 4 of the 7 patients were receiving
cytotoxic agents (patients no. 5 and 9 receiving bolus cyclophosphamide intravenously every 3 months; patients
no. 2 and 8 receiving cyclophosphamide and azathioprine orally 25 to 50 mg per day) in addition to prednisone. None of the 5 patients with active SLE were receiving cytotoxic agents at the time of the study.
Thus, despite peripheral lymphocytopenia in
each group, patients with Sjogren's syndrome had normal proportions of bone marrow mononuclear cells,
while patients with SLE had markedly reduced proportions of mononuclear cells.
Ig-secreting cells in peripheral blood. The relative numbers of peripheral blood B cells in patients and
controls spontaneously secreting Ig as measured by
PFC responses are illustrated in Figure 1. Patients with
inactive and active Sjogren's syndrome had numbers of
circulating B cells spontaneously secreting Ig which
were the same as normal controls (P> 0.2 for both patient groups). Conversely, patients with both inactive
and active SLE had markedly increased numbers of
cells spontaneously secreting Ig (P < 0.05 and P <
0.00 1, respectively). Furthermore, patients with active
SLE had significantly more PFC in peripheral blood
than did patients with inactive SLE (P< 0.05).
A
A
A
A
assn
-
*
hORMALS'INACTIVE' ACTIVE
SJOGRENS
SYNDROME
&
-..AJACTlVl
SLE
Figure 1. Spontaneous Ig-secreting cells in the peripheral blood of
normal individuals and patients with Sjogren's syndrome and SLE.
Each symbol represents an individual subject; horizontal bars represent mean responses.
'"oi1 1,
,
130 120 140
u)
Y
0
2
2
110-
2
y1
100-
z
P0
50
n
$
2n
90-
80-
60
70
50
-
30 -
40
20
-
I
A
0
A
1
.
1
SJOGREN,S
SYNDROME
SLE
Figure 2. Spontaneous Ig-secreting cells in the bone marrow of normal individuals and patients with Sjogren's syndrome and SLE. Bone
marrows were assayed at the same time as the peripheral blood in the
same individuals. Each symbol represents an individual subject; horizontal bars represent mean responses.
Ig-secreting cells in bone marrow. The relative
numbers of mononuclear cells spontaneously secreting
Ig in the bone marrow of patients and controls are illustrated in Figure 2. Bone marrow aspirates were technically unsuccessful in one patient with active Sjogren's
syndrome and one patient with active SLE, hence the
discrepancy in numbers of bone marrow aspirates (Figure 2) in these groups compared to peripheral blood
(Figure 1).
In certain experiments, IgG, IgA, and IgM PFC
were determined in addition to total Ig PFC, and it was
consistently found that the increase in PFC was predominantly, but not exclusively, of the IgG class. This
was also true for PFC detected in the peripheral blood.
The mean (+ SEM) PFC response of bone marrow
mononuclear cells in controls was 7,217 (+ 2,163) PFC
per lo6 mononuclear cells. Thus in normal subjects, the
bone marrow contains a 20-fold greater number of
spontaneous Ig-secreting cells per 1O6 mononuclear cells
compared to peripheral blood with 382 (+ 89) PFC per
lo6 mononuclear cells (Figure 1).
Similar to peripheral blood, patients with inactive and active Sjogren's syndrome had numbers of
bone marrow mononuclear cells spontaneously secreting Ig which were the same as controls (P > 0.2 for
582
FAUCI AND MOUTSOPOULOS
both patient groups). Again, similar to peripheral blood,
bone marrow mononuclear cells from patients with inactive and active SLE had markedly increased numbers
of cells spontaneously secreting Ig (P < 0.01 and P c
0.05, respectively). Although the mean PFC response of
patients with active SLE is apparently greater than that
of patients with inactive disease, the difference is not
statistically significant. This may be due to the wide
range of responses together with the small sample of patients.
Thus, although patients with SLE had
markedly reduced relative proportions of mononuclear
cells in bone marrow aspirates (Table 3), among those
bone marrow mononuclear cells that were present were
markedly increased numbers of activated B cells spontaneously secreting Ig. Furthermore, this spontaneously
secreted Ig was truly polyclonal in nature, since in several experiments PFC simultaneously assayed against a
number of non-cross-reacting antigenic determinants
such as SRBC, trinitrophenyl hapten (TNP), and keyhole limpet hemocyanin (KLH) were all markedly increased above normal, paralleling the increased total Ig
PFC (Table 4). In addition, the polyclonal nature of the
B cell activation in the bone marrow of patients with
SLE is shown in Table 5.
DISCUSSION
The present study demonstrates several interesting and potentially important observations regarding
the presence of spontaneous Ig-secreting cells in the peripheral blood versus the bone marrow of normal individuals as well as patients with either Sjogren’s syndrome or SLE, both of which are characterized
clinically by B cell hyperreactivity. With regard to normal individuals, we have previously reported in a large
number of normal subjects that the mean number of
Table 4. Polyclonally activated B cells in the peripheral blood of
normal individuals, patients with active Sjogren’s syndrome, and
patients with SLE
Direct PFC per lo6 lymphocytes*
Subject
(number)
AntiSRBC
Controls (5)
Active Sjogren’s
syndrome (5)
Active
SLE (5)
1.2 (& 0.6)
4 (f 1.5)
2 ( * 1)
5.2 (& 1.4)
17 (f3.9)
11 (k2.7)
135 (*42)
11 1 (f3 1)
110 (& 28)
AntiTNP
AntiKLH
* Data represent the mean (*SEM) PFC per lo6 lymphocytes measured immediately upon separation of mononuclear cells from peripheral blood, i.e., spontaneous PFC.
Table 5. Polyclonally activated B cells in the bone marrow of
patients with active SLE
Direct PFC*
Patient
no.
Indirect
total
IgPFC*
AntiSRBC
AntiTNP
AntiKLH
4
7
10
390,000
108,000
39,000
20,Ooo
8,900
2,700
14,500
11,600
6,300
22,700
7,600
4,200
* Data represent the PFC per lo6 mononuclear cells measured immediately upon obtaining cells from bone marrow aspirates, i.e.,
spontaneous PFC.
spontaneous Ig-secreting cells in peripheral blood measured by the SPA-PFC assay was 468 PFC per lo6
mononuclear cells (34). This is consistent with the value
of 382 (f69) PFC per lo6 mononuclear cells reported
here. However, of particular importance was the almost
20-fold increase in PFC in simultaneously assayed bone
marrow from the same normal individuals. We (27-29)
and others (26,37) have previously stressed that in humans, the bone marrow is an important source of lymphocytes, particularly B cells. However, the increased
numbers of Ig-secreting cells in bone marrow cannot be
explained merely by the presence of a greater proportion of B cells, since the proportion of cells with detectable B cell surface markers relative to total mononuclear cells is approximately the same in human
peripheral blood and bone marrow (27,37).
B cells spontaneously secreting Ig immediately
upon assay in vitro are believed to have been activated
in vivo (reviewed in reference 38). Therefore, the increased relative proportion of B cells spontaneously secreting Ig in the bone marrow suggests that the bone
marrow may function in part as an organ of sequestration for activated B cells as well as an organ of de novo
production of virgin B cells. Another possibility is that
B cells are activated in situ in the bone marrow and
only subsequently recirculate. Both concepts are further
strengthened by the present data with both inactive and
active SLE patients. Both had significantly increased
numbers of activated Ig-secreting cells in peripheral
blood (Figure 1). However, both groups had a 35-fold
relative increase in activated Ig-secreting cells in the
bone marrow compared to peripheral blood. This striking increase was seen despite the fact that bone marrows
in both these groups were relatively depleted of total
mononuclear cells.
With regard to bone marrow sequestration of activated B cells, we sought to determine if our previous
failure to demonstrate increased numbers of spontaneous Ig-secreting cells in the peripheral blood of Sjo-
B CELL ACTIVATION IN SLE AND SJOGREN’S SYNDROME
gren’s syndrome patients (25) was due to the possibility
that activated B cells were sequestered in other lymphoid organs such as the bone marrow. This was not
found to be the case since patients with either active or
inactive Sjogren’s syndrome had normal numbers of Igsecreting cells in both the peripheral blood and bone
marrow. This may be explained by the possibility that
activated B cells in Sjogren’s syndrome may be sequestered in other organs such as the spleen or even the involved exocrine glands. In this regard, it has been previously demonstrated that lymphoid tissue infiltrating
accessory salivary glands in lip biopsies of patients with
Sjogren’s syndrome synthesizes Ig in much larger
amounts than found in normal salivary glands taken
from the corresponding site (39). Furthermore, we have
recently demonstrated that lip biopsies from patients
with Sjogren’s syndrome contain increased numbers of
plasmacytoid cells, and these cells have a shift toward
production of IgG compared to a predominance of IgA
production seen in normal controls, as determined by
staining for intracytoplasmic Ig (Fauci AS et al, unpublished observations). The lack of circulating activated B
cells in Sjogren’s syndrome and their presence in the involved salivary glands may merely indicate that the activated cells do not freely enter the circulation in large
numbers.
It is unclear why activated B cells can be detected in the circulation of SLE patients or even in other
disease states where this phenomenon may be as yet
unappreciated. It should be pointed out that in both experimental animals (40’41) and humans (42,43) who
have been parenterally immunized with antigen, cells
synthesizing a specific antibody are present in the circulation for limited periods of time following immunization. We have noted an interesting phenomenon following immunization and boosting of normal volunteers
with KLH. B cells secreting KLH specific antibody were
noted in the circulation for brief periods of time following primary and booster immunization. However, even
larger numbers of B cells spontaneously secreting polyclonal Ig, predominantly IgG, were noted together with
the B cells secreting anti-KLH antibody (Fauci AS, unpublished observations). Thus, an antigen specific immunization resulted in the appearance in the circulation
not only of B cells secreting specific antibody, but B
cells secreting polyclonal antibody.
In the present study, circulating B cells in patients with SLE were secreting polyclonal Ig, particularly of the IgG class, in agreement with other studies
(16,17). The relationship of this phenomenon to the
presence of polyclonally activated circulating B cells in
583
SLE and their absence in Sjogren’s syndrome may have
potential importance in appreciating the nature, origin,
and duration of the stimulus to B cells in each of these
disorders.
ACKNOWLEDGMENTS
The authors gratefully acknowledge the expert technical assistance of Ms Gail Whalen, Ms Cynthia Burch, and
Mrs. Patricia Andrysiak. We also thank Mrs. Cynthia Earp
and Mrs. Joan Barnhart for expert secretarial and editorial assistance.
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