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The Journal of Immunology, 2007, 178: 557-565.
Copyright © 2007 by The American Association of Immunologists, Inc.

B Cell Clonal Expansion and Somatic Hypermutation of Ig Variable Heavy Chain Genes in the Synovial Membrane of Patients with Osteoarthritis1

Reng-Rong Da*, Yufen Qin*, Dominique Baeten{dagger} and Yiping Zhang2,*

* Department of Neurology, University of California Irvine, Irvine, CA 92697; and {dagger} Division of Clinical Immunology and Rheumatology, Academic Medical Center, University of Amsterdam, The Netherlands


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
Inflammatory mediators have been explored as possible factors in the initiation and/or progression of osteoarthritis (OA). This study shows that synovial infiltration by B lymphocytes is present in almost half of the knee OA cases. The degree of B lymphocyte infiltration is associated with more pronounced synovial inflammation and with the presence of plasma cells and lymphoid follicles in more severe cases. To examine whether these B cells are merely bystanders or could be involved in the pathogenesis of OA, we analyzed the Ig H chain variable region (VH) genes of B cells recovered from the synovial membrane of five OA patients with marked B cell infiltration. Sequence analysis of CDR3 regions of rearranged VDJ genes revealed clonal or oligoclonal B cell expansions in all cases. Expanded B cell clones in four of five OA patients showed clustered somatic mutations, occurring mainly in the CDRs and with a high replacement-to-silent ratio (>2.9), indicating that these cells are postgerminal center B cells that had been positively selected through their Ag receptor. These data demonstrate the presence in inflamed knee OA synovium of clonally expanded, Ag-driven B cells that may contribute to the development or progression of the disease.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
Osteoarthritis (OA)3 is generally considered as a noninflammatory disease, but some degree of inflammation is often found in symptomatic OA cases (1, 2, 3). At macroscopic inspection, half of the patients with knee OA depict synovial abnormalities, which were shown to be associated with more severe chondropathy (4). Histological analysis of synovial biopsies confirmed mild to moderate synovial inflammation in OA, with an infiltration of the synovial sublining by mononuclear cells, including T and B lymphocytes (5, 6, 7, 8, 9, 10, 11). Autoimmunity toward cartilage degradation products is an attractive hypothesis for the genesis of this inflammatory synovial reaction (12), although it is still openly disputed whether these responses represent primary or secondary immune reactions. Accumulating evidence for T and/or B cell autoimmunity against cartilage Ags in OA includes: 1) cartilage Ags, such as collagens and proteoglycans, are released from the damaged joints in patients with OA (13, 14); 2) T cell responses to cartilage matrix proteins are higher in OA patients than in normal healthy donors (15); 3) analysis of DNA restriction enzyme patterns of T lymphocytes from OA synovium shows oligoclonal TCR beta-chain gene rearrangements, suggesting a response by a limited number of T cell clones (16); and 4) autoantibodies against both native and denatured cartilage matrix components are often found in osteoarthritic articular cartilage (17, 18), especially in generalized, nodal OA (19).

The latest observation suggests that Ag-specific B lymphocytes and plasma cells may participate in the pathogenesis of OA. However, direct evidence for this hypothesis is still lacking. The few studies of Ig variable H chain (VH) gene usage by synovial B cells in OA yielded conflicting results with regard to clonal expansion and somatic hypermutation (20, 21, 22). Moreover, these studies used surgical samples obtained in end-stage disease and it remains thus unclear in how far the obtained results are relevant for the pathogenesis of the disease.

Using a unique set of arthroscopic synovial biopsies, we demonstrate in this study that almost half of the OA synovial tissues (ST) depict a mild to moderate synovial infiltration with B lymphocytes. The degree of B cell infiltration is associated with more pronounced local inflammation and with the presence of plasma cells and lymphoid follicles in the more severe cases. We provide direct evidence for clonal and oligoclonal expansion of these synovial B cells. These clonally expanded B cell clones are mainly T cell-dependent, postgerminal center (GC) B cells with Ag-driven somatic hypermutations. Our results suggest that these B cells may contribute to the pathogenesis of knee osteoarthritis by promoting, enhancing, or perpetuating synovial inflammation in a subgroup of patients.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 Disclosures
 References
 
Patients and specimens

This study included 41 patients with knee OA fulfilling the American College of Rheumatology (Atlanta, GA) criteria (23) and presenting consecutively with pain and/or hydrops of the affected knee joint. None of the patients had an underlying inflammatory arthritis such as rheumatoid arthritis (RA), psoriatic arthritis, spondyloarthritis, or crystal-induced arthritis. Accordingly, none of the patients had arthritis-related autoantibodies such as rheumatoid factor and anticitrullinated protein Abs, erosive changes on x-ray, or crystals in synovial fluid (SF) analysis. All but three patients were being treated with nonsteroidal anti-inflammatory drugs; none of the patients were being treated with corticosteroids or disease-modifying antirheumatic drugs. There was no bias for inclusion in terms of disease stage or severity. The demographic, clinical, and biological data of the patients are detailed in Table I. ST biopsies were obtained by needle arthroscopy of the involved knee joint as described previously (24). All patients gave their written informed consent before inclusion in the study, which was approved by the local ethics committees.


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  		Table I. Histopathological analysis of OA synovitisa

 
Synovial histopathology

In each patient, eight synovial biopsies were used for histological analysis on paraffin sections and eight biopsies for immunohistochemistry on frozen sections. Paraffin-embedded biopsies were stained with H&E for histological analysis (6), including the synovial lining layer thickness, the degree of vascularity, the global infiltration by mononuclear cells. The lymphocyte infiltrate was further studied by immunohistochemistry on frozen sections with mAbs directed toward CD3 (T cells), CD20 (B cells), and CD138 (plasma cells), all three from DakoCytomation as extensively described previously (6). Lymphoid follicles were defined as dense aggregates of lymphocytes with a radial cell count of 10 or more (25). Follicular dendritic cells (clone CNA42; DakoCytomation) and high endothelial venules (anti-CD62L; BD Pharmingen) were analyzed as typical features of GCs. Parallel sections were stained with irrelevant origin-, isotype-, and concentration-matched Ab as negative control. Tonsils were used as positive control. Stained sections were coded and analyzed by two independent observers, who were blinded to the clinical data. We used a four-point scale, semiquantitative scoring methodology (zero as lowest score and three as highest score), as extensively described and validated previously (6).

PCR amplification of biopsy synovial B cells

Total RNA was extracted from additional synovial biopsies (n = 5; six biopsies in each patient) from OA patients with pronounced B cell infiltration using an RNeasy kit (Qiagen). None of the patients were being treated with corticosteroids or disease-modifying antirheumatic drugs. A mucosal B cell lymphoma and peripheral blood cells from a healthy individual were used as positive and negative control, respectively, for the B cell clonality analysis. PCR was conducted for 40 cycles under standard conditions (denaturation, 1 min at 94°C; annealing, 2 min at 52–56°C; and extension, 1 min at 72°C). Aliquots of the PCR product were analyzed by electrophoresis in a 2% agarose gel (Sigma-Aldrich) containing ethidium bromide. Briefly, the first-strand cDNA was synthesized using oligo(dT) as primer and avian myeloblastosis virus reverse transcriptase. Ig VH and VDJ genes were amplified via PCR (26) in a final volume of 50 µl of reaction buffer (50 mM Tris-HCl (pH 9.0) at 25°C,20 mM (NH4)2SO4, and 3.0 mM MgCl2) containing 2 U of recombinant Taq polymerase, and 20- pmol primers (27, 28) under standard conditions (denaturation, 1 min at 94°C; annealing, 2 min at 52–56°C; and extension, 1 min at 72°C). Aliquots of the PCR product were analyzed by electrophoresis in a 2% agarose gel (Sigma-Aldrich) containing ethidium bromide. When PCR products showed negative results, nested PCR was performed.

Sequencing Ig VH genes

PCR products were recovered and ligated into the pGEM T vector (Promega) and transfected into Escherichia coli DH5a according to Hanahan (29). According to power analysis, at least eight white colonies were picked at random in each case and grown overnight in 3 ml of Luria-Bertani medium. The dsDNA template from the colonies containing VH gene inserts was confirmed by EcoRI digestion and then sequenced using a high-efficiency DNA sequencer (MCLAB).

Assignment of mutations

Mutations identified by comparing each sequence with germline sequences (30) (international ImMunoGeneTics database at http://imgt cines.fr/) were defined on the basis of nucleotide changes in the VH segment. Variabilities at the joining sites of the VH, DH, and JH gene segments were not classified as mutations because they might result either from the insertion of N regions or from mutation. Two nucleotide exchanges in a single codon were scored as one replacement mutation. The human Ig H chain CDR3 region and clonality were determined by Ig VDJ junction analysis with software provided by ImMunoGeneTics/JunctionAnalysis (31).

Statistics

For analysis of the clinical and histological parameters, the OA samples were classified according to the degree of B cell infiltration and compared with the unpaired Mann-Whitney U test; the data are represented as median (interquartile range). For clonality analysis, the positive control showed identical B cell clones in >80% of the randomly selected sequences and the negative control 0%. Sample size estimation with nQuery software (Statistical Solutions) indicated that in this situation analysis of seven sequences of each sample would provide 85% power by the Fisher’s exact test of equal portions with a p value of 0.05, two-sided significance. Therefore, more than seven single Ig VH nucleotide sequences (8–16 clones) of each ST sample were analyzed.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 Disclosures
 References
 
B lymphocyte infiltration in active OA synovitis

We analyzed 41 OA ST samples for B lymphocyte infiltration (CD20). In 22 cases, no B cell infiltration was observed, whereas 13 samples depicted mild B cell infiltration and 6 cases moderate to strong B cell infiltration (Table I). There were no differences in demographic or clinical features between these groups, including disease duration and signs of systemic inflammation. Reflecting local inflammation, SF effusion was found in 13 of 22, 10 of 13, and 6 of 6 cases, respectively, with a concomitant increase in the number of white blood cells in the SF (Table I). Histological analysis showed no difference in synovial lining layer hyperplasia and degree of vascularity in the three groups, but the global inflammatory infiltration was significantly increased in the group with moderate to strong B cell infiltration (Table I). Accordingly, T lymphocyte infiltration (CD3) was significantly increased in this group. Plasma cells (CD138) were found in all samples with moderate to strong B cell infiltration, but in only one of the other samples. Similarly, lymphoid follicles were found in all samples with moderate to strong B cell infiltration but in none of the other samples. However, these follicles did not show follicular dendritic cells and high endothelial venules as observed in genuine GCs. Collectively, these data indicate that OA synovitis is characterized by B cell infiltration in almost half of the cases and that pronounced B cell infiltration is associated with more severe local inflammation with the presence of plasma cells and synovial lymphoid follicles (Fig. 1).


Figure 1
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  		FIGURE 1. Histology study of OA synovial tissue. Representative micrographs of ST obtained from OA knee joints with clinical inflammation as evidenced by joint effusion (left panels) and from OA knee joints without clinical inflammation (right panels). Upper panels, Mild to moderate inflammatory infiltration of the sublining layer was seen in both cohorts, but follicular organization was only seen in the group with clinical inflammation. Immunohistochemical analysis demonstrate that T lymphocytes (CD3) are seen in both groups, whereas the infiltration by B lymphocytes (CD20) and plasma cells (CD138) is more pronounced in the inflammatory OA group. The immunohistochemical stainings confirm the follicular organization of the lymphoid infiltration in the more severe cases of inflammatory synovitis in OA.

 
Ig H chain CDR3 region and clonality analysis. Because our histological observations suggested an association between synovial B cell infiltration and inflammation in the OA joint, five cases with pronounced B cell infiltration were selected for further clonality analysis by RT-PCR and sequencing. A B cell lymphoma sample and normal peripheral blood cells were used as monoclonal and polyclonal controls, respectively. The PCR primers were designed to amplify all VH gene segments (27, 28). RT-PCR showed that Ig VH genes were amplified in all of the specimens and the controls. No VH gene was amplified from the H2O control.

Nucleotide sequences of the VH gene CDR3 region revealed dominant clonal or oligoclonal expansion in all five OA samples. The expanded clones derived from each of these specimens were identical in the VH-N-D-N-JH regions, with similar sequences and nucleotide lengths and a defined VH, D, and JH fragment usage (Fig. 2). In case 5, only one dominant B cell clonal expansion was found (VH1 3*01, D4 4*01, and JH6*02). The other OA synovial samples showed two expanded B cell clones. The sequences from B cell lymphoma showed eight of eight identical CDR3 sequences, whereas the sequences from peripheral blood B cells showed variability at the joining sites of the CDR3 genes and had no identical CDR3 rearrangements (Fig. 2). Based on a comparison of the expressed D segments to those of published germline D segments (30), the combination of VH, DH, and JH segments of the clonally expanded B cells from OA synovium showed D-D fusion, gene replacement, and the addition of nucleotide sequences termed N regions (Fig. 2). The VH family and gene segment use of the clonal B cells were quite heterogeneous, including the VH 1, VH 3, and VH 5 families (Fig. 2). The deduced peptide sequences of the VDJ junctions of the clonally expanded B cells are shown in Fig. 3. When compared, the nucleotide sequences of dominant B cell clone(s) from each of five cases each possessed an individual pattern of VH, DH, and JH combination.


Figure 2
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  		FIGURE 2. B cell clonality analysis in OA patients. Eight to 16 complete Ig VH genes from each specimen were randomly selected and sequenced. The nucleotide sequences in the CDR3 region, grouped into VH, NDHN, and JH regions, are shown. The name of the germline VH, DH, and JH genes showing maximum homology to the sequences are also shown. CDR3 gene sequences of peripheral blood B cells from a normal donor served as polyclonal B cell control. As positive control we used randomly selected B cell clones from a biopsy specimen from a patient with mucosa B cell lymphoma. The bold letters with underline indicates the offspring nucleotide variations and dashes indicate the identity to the reference sequence. Os, Offspring.

 

Figure 3
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  		FIGURE 3. VH gene CDR3 region-encoded peptides of OA synovial B cells. The deduced amino acid sequences of the CDR3 region are shown in the VH, NDN, and JH regions. The bold letters with underline indicates the amino acid change resulting from offspring replacement nucleotide mutation and dashes indicate the identity to the reference sequence. Os, Offspring.

 
Somatic hypermutation

The clonal and oligoclonal expansion of synovial B cells in OA synovium raised the question of the origin and characteristics of these cells. To elucidate these issues, the nucleotide mutations along the sequences of clonally expanded Ig VH genes were analyzed. Table II shows the nucleotide substitutions when compared with the closest known germline VH genes (National Center for Biotechnology Information at http://www.ncbi.nlm.nih.gov/igblast). These VH gene sequences expressed by clonally or oligoclonally expanded B cells from OA ST showed various degrees of similarity (88–97%) with the appropriate germline VH gene sequence. The total nucleotide substitutions, replacement mutations, and the ratio of replacement and silent mutations (R:S) of framework and CDRs are summarized in Table II. The dominant VH gene sequence from case 3 displayed the highest degree of similarity (97%) with the germline VH5-51*03 gene (30); nine substitutions were found in its VH gene, five in the CDR regions. and four in the framework regions (FR). One of the VH sequences (VH1-69) of case 1 showed the lowest degree of similarity (88%) with the germline sequence; it contained 34 nt substitutions (Table II).


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  		Table II. VH family usage and differences in nucleotide sequences in comparison with the nearest germline VH genesa

 
Compared with the germline, the majority of the VH genes analyzed in OA STs contained substitutions distributed in a pattern characteristic of Ag-driven affinity maturation, i.e., the somatic mutations were highly concentrated in the CDRs or FRs, with a clustering of replacement mutations in the CDRs rather than in FRs. The CDR region R:S value of expanded B cell clones in cases 2, 3, 4, and 5 (case 2, clone VH4-31 and clone VH3-11, both with a R:S = 6; case 3, cloneVH3-7, 4.0; case 4, clone VH3–33, 3.0 and clone VH1-2, 6.0; and case 5, 8.0) were all significantly higher than the theoretical R:S value of ~2.9, which was calculated for somatic mutations occurring randomly in a gene encoding a protein whose structure need not be preserved (31). These patterns are consistent with the notion of an Ag-driven selection with high-affinity Ag binding sites. Three expanded B cell clones (case 1, clone VH5-51 R:S = 1.0 and clone VH1-69 R:S = 1.4; and case 3, clone VH5-51 R:S = 1.5) had R:S values that are much lower than the theoretical value, suggesting the participation of non-Ag-driven B cell clones.

Intraclonal nucleotide variation

It was interesting that clone VH3-11 of case 2 had further nucleotide variations. Although they had the same length of nucleotides in the CDR3 region, in comparison to offspring 1–4, there was one replacement nucleotide substitution (from g to c, see Fig. 2, case 2) in the VDJ regions (offspring 5, 6, and 7) and results in an amino acid change from serine to threonine in the CDR3 peptide sequence (offspring 5 and 6) and from phenylalanine to leucine (offspring 7) (Figs. 3 and 4). The alignment of expanded VH sequences to germline VH3-11 sequences also showed two intraclonal silent mutations in the FR3 region (g to a and c to t; Fig. 4) with no amino acid substitution (offspring 5–7; Fig. 4). Moreover, offspring 7 had an additional two mutations, one in CDR1 (g to a), which caused an amino acid substitution of methionine to isoleucine, and one silent mutation in framework 2 (Fig. 4). In this study, these offspring nucleotide variations were limited in case 2, clone VH3-11. No similar nucleotide variations were observed in the other eight VH clonally expanded B cells (Figs. 2 and 3).


Figure 4
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  		FIGURE 4. Nucleotide variations and deduced amino acid sequences of offspring clones from case 2 clone VH3-11. A, The nucleotide sequences are shown in comparison with germline sequences. B, The deduced amino acids of each sequence are also shown. Conventionally defined FRs and CDRs are indicated. Dashes indicate the identity to the reference sequence. The bold letters indicate the offspring (Os) silent mutations. The bold letters with underline indicate the replacement mutations.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
Disclosures
 References
 
In recent years, the analysis of the Ig V gene repertoire had a great impact on the understanding of B cell responses in organ-specific autoimmune diseases, such as multiple sclerosis (32) and RA (33). Autoreactive B cells and Ab specificities have been widely studied in peripheral blood, SF, ST, and bone marrow of RA patients (33), whereas such data are scarce in other types of chronic arthritis including OA. To gain insights into the possible role of B cells in OA, we analyzed the presence and Ig VH gene usage of B cells in OA synovitis. In this study, we show that B lymphocytes are infiltrating the synovial membrane in clinically inflamed knee OA joints, with the presence of lymphoid follicles and plasma cells in severe cases (1); B cell clonal and oligoclonal expansions are common in inflamed OA synovium (2); most of the VH genes are highly mutated in the CDR regions and indicate the presence of Ag-driven, post-GC memory B cells, although some Ag-independent B cell clones may also infiltrate into OA ST (3); and the expression of intraclonal offspring mutations in OA synovial B cells indicate an in situ proliferation of these clone(s) under the pressure of local Ag stimulation (4).

OA is generally considered as a disease of articular cartilage, with a primary failure of chondrocyte and extracellular matrix homeostasis induced by a variety of genetic, metabolic, and biochemical factors. However, increasing evidence points toward inflammation of the synovial membrane in OA, ranging from minimal lining cell layer hyperplasia to severe acute and chronic synovitis (10, 34, 35). Early OA with active joint effusion can also depict some systemic signs of inflammation such as increased C-reactive protein levels (36). In the joint, the infiltrating leukocytes and the resident synovial fibroblasts can release inflammatory mediators, in particular matrix metalloproteinases and IL-1, that contribute to the degradation of the extracellular cartilage matrix and the suppression of chondrocyte anabolism (37). These data indicate that, although probably not the primary mechanisms initiating the disease, inflammation may contribute to the progression and/or perpetuation of OA.

Although this inflammatory component of OA is now well recognized, recently accumulated evidence supports the involvement of autoimmune mechanisms in this process. The synovial infiltration in knee OA is characterized by the presence of both T and B lymphocytes (22, 38). Cellular autoimmunity against cartilage link protein and proteoglycan aggrecan have been observed (15) in OA patients. Moreover, autoantibodies against cartilage and bone proteins such as collagen type II (17, 39), the cartilage intermediate layer protein (40), YKL-39 (41), and osteopontin (42), which have been demonstrated in the serum of RA patients, have also been detected in OA patients. A recent comprehensive proteomic surveillance has revealed that some Ags are recognized predominantly in OA rather than in RA (4). In animal models, immunization with cartilage proteins such as collagen type II, human cartilage glycoprotein-39 (44), cartilage link protein (45), and proteoglycans (46, 47) induce destructive arthritis whereas cartilage intermediate layer protein (40) and YKL-39 (42) cause mild synovitis but no cartilage or bone destruction. Whether the immune reaction against these or other cartilage autoantigens play a role in triggering and/or perpetuating synovitis and cartilage damage in OA remains to be demonstrated.

In the present study, we demonstrated that mild to moderate synovial infiltration with B lymphocytes is present in nearly half of the OA STs, independently of systemic inflammation or disease duration. Interestingly, the samples with B cell infiltration were characterized by more pronounced local inflammation in terms of SF effusion and synovial inflammatory cell infiltration. Moreover, samples containing high numbers of B cells had the most severe inflammation with the presence of plasma cells and lymphoid follicles, which is consistent with data obtained from surgical sample sets (5, 11, 25, 48). The specific microarchitecture of the lymphoid infiltrate is of particular importance because ectopic lymphoid organogenesis in the synovial membrane has been suggested to promote autoimmunity in other forms of chronic arthritis (49, 50).

We provided further evidence for a possible role of these B cells in OA by demonstrating their clonal expansion by Ig VH gene analysis. In other words, the B cell homing and/or expansion in the inflamed OA synovial membrane is a selective rather than a random event. The first impression of clonality of OA synovial B cells came from the relative sharp bands of PCR products (data not shown). presumably due to the identical length of CDR3 sequences of most VH CDR3 fragments usage, as shown in an earlier study (22). We show here that the synovial B cells of all five OA samples each had a clonal and or dual clonal rearrangement of the CDR3 region. Each expanded B cell clone from a given individual was identical in the sequence and nucleotide length of the VH-N-D-N-JH regions (Fig. 2) as well as in the deduced CDR3 peptide sequence (Fig. 3). In comparing all the expanded B cell clones, no apparent preferential use of any particular VH, DH, and JH gene segment set was seen.

Compared with the germline, the VH genes analyzed in OA STs contained substitutions distributed in a pattern characteristic of Ag-driven affinity maturation, i.e., the somatic mutations were highly concentrated in the CDRs or FRs, with a clustering of replacement mutations in the CDRs rather than in FRs. The R:S values in CDR regions of most expanded B cell clones in OA synovium were significantly higher than the theoretical R:S value of ~2.9. This pattern suggests an Ag-driven selection toward high-affinity Ag binding sites. In contrast to a previous study demonstrating somatic hypermutation but no clonality or lympoid follicle formation in OA synovium (20), our observations are coherent and consistent with the notion that most of the clonally expanded OA synovial B cells have undergone Ag-driven, T cell-dependent somatic hypermutation and affinity maturation in the GCs of secondary lymphoid tissue before recruitment into the ST or, alternatively, directly in the ectopic lymphoid follicles that we detected in a subset of the OA synovia. However, we failed to demonstrate the presence of follicular dendritic cells and high endothelial venules in the cases of follicular OA synovitis. The presence of fully functional GCs in OA synovium therefore needs to be further analyzed in larger cohorts before we can lend any support to this alternative hypothesis.

The nucleotide variations in seven offspring in clone VH3-11 (Fig. 4) of case 2 are of great interest because it may suggest a consistent Ag stimulation in the lesion. Practically, these mutations may be attributable to infidelity of Taq polymerase, which has been reported to result in an error rate of 1/5000–1/9000 errors per base polymerized (51, 52). However, the degree of sequence variation present in case 2 clone VH3-11 was 88 of 2541 nt and cannot, therefore, be totally accounted for by Taq infidelity. However, this observation only occurs in one of five case studies and the most nucleotide variations are limited in FR3 and CDR3 regions. Therefore, whether these intraclonal nucleotide variations of VDJ genes expressed have biological and pathological consequences in OA development need more sophisticated studies.

Two limitations of the present study should be carefully considered for the clinical translation of the proposed hypothesis. First, it is unknown whether the described B cell alterations are limited to a subset of manifest knee OA as studied here or whether the potential involvement of B cells is more widely applicable to different forms of OA. Indeed, OA is a heterogeneous disease with different subtypes according to disease localization, metabolic and genetic factors, stage, and severity. Second, knee OA is a multifactorial disease including a variety of mechanical and metabolic pathways. The interaction of B cell-mediated inflammation with these complex disease mechanisms as well as the relative contribution to drive, enhance, or rather perpetuate the disease remains to be clarified.

The findings of the present study lend credence to the hypothesis that osteoarthritic Ags may drive chronic B cell autoimmunity, which could in turn positively or negatively modulate the disease process (8, 53). The Abs produced by the plasma cells found in follicular OA synovitis or other B cell effector functions may further aggravate synovial inflammation and cartilage damage. Collectively, the presented data lead to the new hypothesis that Ag-driven and presumably autoreactive B cells may be involved in the pathogenesis of inflammatory OA and warrant further studies of the precise underlying molecular pathways as well as of the clinical and pathological implications.


   Disclosures
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
References
 
The authors have no financial conflict of interest.


   Footnotes
 
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

1 This study was supported by a National Institutes of Health Grant RO1 AI 54911. Back

2 Address correspondence and reprint requests to Dr. Yiping Zhang, Department of Neurology, University of California Irvine, 100 Irvine Hall, Irvine, CA 92627. E-mail address: zhangy{at}uci.edu Back

3 Abbreviations used in this paper: OA, osteoarthritis; VH, variable H chain; ST, synovial tissue; GC, germinal center; R:S, replacement and silent mutation; FR, framework region; RA, rheumatoid arthritis; SF, synovial fluid. Back

Received for publication April 14, 2006. Accepted for publication October 10, 2006.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 

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