Search tips
Search criteria 


Logo of molmedLink to Publisher's site
Mol Med. 1998 April; 4(4): 240–257.
PMCID: PMC2230361

Clonally-related immunoglobulin VH domains and nonrandom use of DH gene segments in rheumatoid arthritis synovium.


BACKGROUND: Synovia of patients with long-standing rheumatoid arthritis (RA) are typically infiltrated with B lymphocytes and plasma cells that secrete large amounts of immunoglobulin. The CDR3 of an immunoglobulin heavy chain is composed of the VH-DH-JH join, with interposed N region addition, and thus defines clonal relatedness. Furthermore, the CDR3 lies at the center of the antigen binding site, so its length and composition influence antigen binding. We sought definitive evidence of an antigen-driven B cell response (i.e., clones derived from the same VH, DH, and JH gene segments with shared somatic mutations) in RA synovial mRNA transcripts, and to characterize CDR3 intervals at the target of inflammation in this autoimmune disease. MATERIALS AND METHODS: We screened a cDNA library generated from unselected cells from the knee joint of a 62-year-old white female with long-standing RA. This technique does not have the potential bias of selecting for antibodies that express a particular reactivity such as rheumatoid factor. C gamma recombinants were sequenced and progenitor VH, DH, and JH gene segments were assigned and somatic mutations determined by comparison to germline sequences. Analyses of DH reading frame utilization and hydropathy characteristics of CDR3s were performed. RESULTS: Two of 67 recombinants were derived from the same VH (V3-11) and JH gene segments, demonstrated shared mutations, and contained nearly identical VH-DH-JH joins, including N region addition. Three other recombinants contained identical sequence throughout the variable domain. We also found preferential utilization of a limited number of VH and DH gene segments and marked preference for a DH reading frame encoding predominantly hydrophilic residues. CONCLUSIONS: Analysis of expressed heavy chain variable domains strongly supports the hypothesis that the B cell response in RA synovium is at least in part antigen driven and oligoclonal.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (2.8M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Images in this article

Click on the image to see a larger version.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Zvaifler NJ. The immunopathology of joint inflammation in rheumatoid arthritis. Adv Immunol. 1973;16(0):265–336. [PubMed]
  • Tonegawa S. Somatic generation of antibody diversity. Nature. 1983 Apr 14;302(5909):575–581. [PubMed]
  • Shin EK, Matsuda F, Nagaoka H, Fukita Y, Imai T, Yokoyama K, Soeda E, Honjo T. Physical map of the 3' region of the human immunoglobulin heavy chain locus: clustering of autoantibody-related variable segments in one haplotype. EMBO J. 1991 Dec;10(12):3641–3645. [PubMed]
  • Cook GP, Tomlinson IM, Walter G, Riethman H, Carter NP, Buluwela L, Winter G, Rabbitts TH. A map of the human immunoglobulin VH locus completed by analysis of the telomeric region of chromosome 14q. Nat Genet. 1994 Jun;7(2):162–168. [PubMed]
  • Matsuda F, Shin EK, Nagaoka H, Matsumura R, Haino M, Fukita Y, Taka-ishi S, Imai T, Riley JH, Anand R, et al. Structure and physical map of 64 variable segments in the 3'0.8-megabase region of the human immunoglobulin heavy-chain locus. Nat Genet. 1993 Jan;3(1):88–94. [PubMed]
  • Kodaira M, Kinashi T, Umemura I, Matsuda F, Noma T, Ono Y, Honjo T. Organization and evolution of variable region genes of the human immunoglobulin heavy chain. J Mol Biol. 1986 Aug 20;190(4):529–541. [PubMed]
  • Lee KH, Matsuda F, Kinashi T, Kodaira M, Honjo T. A novel family of variable region genes of the human immunoglobulin heavy chain. J Mol Biol. 1987 Jun 20;195(4):761–768. [PubMed]
  • Shen A, Humphries C, Tucker P, Blattner F. Human heavy-chain variable region gene family nonrandomly rearranged in familial chronic lymphocytic leukemia. Proc Natl Acad Sci U S A. 1987 Dec;84(23):8563–8567. [PubMed]
  • Berman JE, Mellis SJ, Pollock R, Smith CL, Suh H, Heinke B, Kowal C, Surti U, Chess L, Cantor CR, et al. Content and organization of the human Ig VH locus: definition of three new VH families and linkage to the Ig CH locus. EMBO J. 1988 Mar;7(3):727–738. [PubMed]
  • van Dijk KW, Mortari F, Kirkham PM, Schroeder HW, Jr, Milner EC. The human immunoglobulin VH7 gene family consists of a small, polymorphic group of six to eight gene segments dispersed throughout the VH locus. Eur J Immunol. 1993 Apr;23(4):832–839. [PubMed]
  • Ichihara Y, Matsuoka H, Kurosawa Y. Organization of human immunoglobulin heavy chain diversity gene loci. EMBO J. 1988 Dec 20;7(13):4141–4150. [PubMed]
  • Buluwela L, Albertson DG, Sherrington P, Rabbitts PH, Spurr N, Rabbitts TH. The use of chromosomal translocations to study human immunoglobulin gene organization: mapping DH segments within 35 kb of the C mu gene and identification of a new DH locus. EMBO J. 1988 Jul;7(7):2003–2010. [PubMed]
  • Siebenlist U, Ravetch JV, Korsmeyer S, Waldmann T, Leder P. Human immunoglobulin D segments encoded in tandem multigenic families. Nature. 1981 Dec 17;294(5842):631–635. [PubMed]
  • Corbett SJ, Tomlinson IM, Sonnhammer EL, Buck D, Winter G. Sequence of the human immunoglobulin diversity (D) segment locus: a systematic analysis provides no evidence for the use of DIR segments, inverted D segments, "minor" D segments or D-D recombination. J Mol Biol. 1997 Jul 25;270(4):587–597. [PubMed]
  • Ravetch JV, Siebenlist U, Korsmeyer S, Waldmann T, Leder P. Structure of the human immunoglobulin mu locus: characterization of embryonic and rearranged J and D genes. Cell. 1981 Dec;27(3 Pt 2):583–591. [PubMed]
  • Sanz I. Multiple mechanisms participate in the generation of diversity of human H chain CDR3 regions. J Immunol. 1991 Sep 1;147(5):1720–1729. [PubMed]
  • Desiderio SV, Yancopoulos GD, Paskind M, Thomas E, Boss MA, Landau N, Alt FW, Baltimore D. Insertion of N regions into heavy-chain genes is correlated with expression of terminal deoxytransferase in B cells. Nature. 1984 Oct 25;311(5988):752–755. [PubMed]
  • Shlomchik MJ, Marshak-Rothstein A, Wolfowicz CB, Rothstein TL, Weigert MG. The role of clonal selection and somatic mutation in autoimmunity. Nature. 328(6133):805–811. [PubMed]
  • Bridges SL, Jr, Lee SK, Koopman WJ, Schroeder HW., Jr Analysis of immunoglobulin gamma heavy chain expression in synovial tissue of a patient with rheumatoid arthritis. Arthritis Rheum. 1993 May;36(5):631–641. [PubMed]
  • Lee SK, Bridges SL, Jr, Koopman WJ, Schroeder HW., Jr The immunoglobulin kappa light chain repertoire expressed in the synovium of a patient with rheumatoid arthritis. Arthritis Rheum. 1992 Aug;35(8):905–913. [PubMed]
  • Sanger F, Nicklen S, Coulson AR. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. [PubMed]
  • Elgavish RA, Schroeder HW., Jr SAW: a graphical user interface for the analysis of immunoglobulin variable domain sequences. Biotechniques. 1993 Dec;15(6):1066–1071. [PubMed]
  • Mortari F, Wang JY, Schroeder HW., Jr Human cord blood antibody repertoire. Mixed population of VH gene segments and CDR3 distribution in the expressed C alpha and C gamma repertoires. J Immunol. 1993 Feb 15;150(4):1348–1357. [PubMed]
  • Brezinschek HP, Brezinschek RI, Lipsky PE. Analysis of the heavy chain repertoire of human peripheral B cells using single-cell polymerase chain reaction. J Immunol. 1995 Jul 1;155(1):190–202. [PubMed]
  • Gu H, Förster I, Rajewsky K. Sequence homologies, N sequence insertion and JH gene utilization in VHDJH joining: implications for the joining mechanism and the ontogenetic timing of Ly1 B cell and B-CLL progenitor generation. EMBO J. 1990 Jul;9(7):2133–2140. [PubMed]
  • Schroeder HW, Jr, Mortari F, Shiokawa S, Kirkham PM, Elgavish RA, Bertrand FE., 3rd Developmental regulation of the human antibody repertoire. Ann N Y Acad Sci. 1995 Sep 29;764:242–260. [PubMed]
  • Kyte J, Doolittle RF. A simple method for displaying the hydropathic character of a protein. J Mol Biol. 1982 May 5;157(1):105–132. [PubMed]
  • Schroeder HW, Jr, Hillson JL, Perlmutter RM. Early restriction of the human antibody repertoire. Science. 1987 Nov 6;238(4828):791–793. [PubMed]
  • Schroeder HW, Jr, Wang JY. Preferential utilization of conserved immunoglobulin heavy chain variable gene segments during human fetal life. Proc Natl Acad Sci U S A. 1990 Aug;87(16):6146–6150. [PubMed]
  • Schroeder HW, Jr, Dighiero G. The pathogenesis of chronic lymphocytic leukemia: analysis of the antibody repertoire. Immunol Today. 1994 Jun;15(6):288–294. [PubMed]
  • Soto-Gil RW, Olee T, Klink BK, Kenny TP, Robbins DL, Carson DA, Chen PP. A systematic approach to defining the germline gene counterparts of a mutated autoantibody from a patient with rheumatoid arthritis. Arthritis Rheum. 1992 Mar;35(3):356–363. [PubMed]
  • Dersimonian H, Schwartz RS, Barrett KJ, Stollar BD. Relationship of human variable region heavy chain germ-line genes to genes encoding anti-DNA autoantibodies. J Immunol. 1987 Oct 1;139(7):2496–2501. [PubMed]
  • Cairns E, Kwong PC, Misener V, Ip P, Bell DA, Siminovitch KA. Analysis of variable region genes encoding a human anti-DNA antibody of normal origin. Implications for the molecular basis of human autoimmune responses. J Immunol. 1989 Jul 15;143(2):685–691. [PubMed]
  • Lu EW, Deftos M, Tighe H, Carson DA, Chen PP. Generation and characterization of two monoclonal self-associating IgG rheumatoid factors from a rheumatoid synovium. Arthritis Rheum. 1992 Jan;35(1):101–105. [PubMed]
  • Ebeling SB, Schutte ME, Akkermans-Koolhaas KE, Bloem AC, Gmelig-Meyling FH, Logtenberg T. Expression of members of the immunoglobulin VH3 gene families is not restricted at the level of individual genes in human chronic lymphocytic leukemia. Int Immunol. 1992 Mar;4(3):313–320. [PubMed]
  • Stüber F, Lee SK, Bridges SL, Jr, Koopman WJ, Schroeder HW, Jr, Gaskin F, Fu SM. A rheumatoid factor from a normal individual encoded by VH2 and V kappa II gene segments. Arthritis Rheum. 1992 Aug;35(8):900–904. [PubMed]
  • Matthyssens G, Rabbitts TH. Structure and multiplicity of genes for the human immunoglobulin heavy chain variable region. Proc Natl Acad Sci U S A. 1980 Nov;77(11):6561–6565. [PubMed]
  • Pascual V, Randen I, Thompson K, Sioud M, Forre O, Natvig J, Capra JD. The complete nucleotide sequences of the heavy chain variable regions of six monospecific rheumatoid factors derived from Epstein-Barr virus-transformed B cells isolated from the synovial tissue of patients with rheumatoid arthritis. Further evidence that some autoantibodies are unmutated copies of germ line genes. J Clin Invest. 1990 Oct;86(4):1320–1328. [PMC free article] [PubMed]
  • Sanz I, Casali P, Thomas JW, Notkins AL, Capra JD. Nucleotide sequences of eight human natural autoantibody VH regions reveals apparent restricted use of VH families. J Immunol. 1989 Jun 1;142(11):4054–4061. [PubMed]
  • Sanz I, Kelly P, Williams C, Scholl S, Tucker P, Capra JD. The smaller human VH gene families display remarkably little polymorphism. EMBO J. 1989 Dec 1;8(12):3741–3748. [PubMed]
  • Shin EK, Akamizu T, Matsuda F, Sugawa H, Fujikura J, Mori T, Honjo T. Variable regions of Ig heavy chain genes encoding antithyrotropin receptor antibodies of patients with Graves' disease. J Immunol. 1994 Feb 1;152(3):1485–1492. [PubMed]
  • Schettino EW, Chai SK, Kasaian MT, Schroeder HW, Jr, Casali P. VHDJH gene sequences and antigen reactivity of monoclonal antibodies produced by human B-1 cells: evidence for somatic selection. J Immunol. 1997 Mar 1;158(5):2477–2489. [PubMed]
  • Yamada M, Wasserman R, Reichard BA, Shane S, Caton AJ, Rovera G. Preferential utilization of specific immunoglobulin heavy chain diversity and joining segments in adult human peripheral blood B lymphocytes. J Exp Med. 1991 Feb 1;173(2):395–407. [PMC free article] [PubMed]
  • Huang C, Stollar BD. A majority of Ig H chain cDNA of normal human adult blood lymphocytes resembles cDNA for fetal Ig and natural autoantibodies. J Immunol. 1993 Nov 15;151(10):5290–5300. [PubMed]
  • Shlomchik M, Mascelli M, Shan H, Radic MZ, Pisetsky D, Marshak-Rothstein A, Weigert M. Anti-DNA antibodies from autoimmune mice arise by clonal expansion and somatic mutation. J Exp Med. 1990 Jan 1;171(1):265–292. [PMC free article] [PubMed]
  • Logtenberg T, Schutte ME, Inghirami G, Berman JE, Gmelig-Meyling FH, Insel RA, Knowles DM, Alt FW. Immunoglobulin VH gene expression in human B cell lines and tumors: biased VH gene expression in chronic lymphocytic leukemia. Int Immunol. 1989;1(4):362–366. [PubMed]
  • Zouali M, Theze J. Probing VH gene-family utilization in human peripheral B cells by in situ hybridization. J Immunol. 1991 Apr 15;146(8):2855–2864. [PubMed]
  • Gauss GH, Lieber MR. Mechanistic constraints on diversity in human V(D)J recombination. Mol Cell Biol. 1996 Jan;16(1):258–269. [PMC free article] [PubMed]
  • Basu M, Hegde MV, Modak MJ. Synthesis of compositionally unique DNA by terminal deoxynucleotidyl transferase. Biochem Biophys Res Commun. 1983 Mar 29;111(3):1105–1112. [PubMed]
  • Stewart AK, Huang C, Long AA, Stollar BD, Schwartz RS. VH-gene representation in autoantibodies reflects the normal human B-cell repertoire. Immunol Rev. 1992 Aug;128:101–122. [PubMed]
  • Cook GP, Tomlinson IM. The human immunoglobulin VH repertoire. Immunol Today. 1995 May;16(5):237–242. [PubMed]
  • Mantovani L, Wilder RL, Casali P. Human rheumatoid B-1a (CD5+ B) cells make somatically hypermutated high affinity IgM rheumatoid factors. J Immunol. 1993 Jul 1;151(1):473–488. [PubMed]
  • Ezaki I, Kanda H, Sakai K, Fukui N, Shingu M, Nobunaga M, Watanabe T. Restricted diversity of the variable region nucleotide sequences of the heavy and light chains of a human rheumatoid factor. Arthritis Rheum. 1991 Mar;34(3):343–350. [PubMed]
  • Van Es JH, Aanstoot H, Gmelig-Meyling FH, Derksen RH, Logtenberg T. A human systemic lupus erythematosus-related anti-cardiolipin/single-stranded DNA autoantibody is encoded by a somatically mutated variant of the developmentally restricted 51P1 VH gene. J Immunol. 1992 Sep 15;149(6):2234–2240. [PubMed]
  • Kipps TJ, Tomhave E, Pratt LF, Duffy S, Chen PP, Carson DA. Developmentally restricted immunoglobulin heavy chain variable region gene expressed at high frequency in chronic lymphocytic leukemia. Proc Natl Acad Sci U S A. 1989 Aug;86(15):5913–5917. [PubMed]
  • Stewart AK, Huang C, Stollar BD, Schwartz RS. High-frequency representation of a single VH gene in the expressed human B cell repertoire. J Exp Med. 1993 Feb 1;177(2):409–418. [PMC free article] [PubMed]
  • Suzuki I, Pfister L, Glas A, Nottenburg C, Milner EC. Representation of rearranged VH gene segments in the human adult antibody repertoire. J Immunol. 1995 Apr 15;154(8):3902–3911. [PubMed]
  • Milner EC, Hufnagle WO, Glas AM, Suzuki I, Alexander C. Polymorphism and utilization of human VH Genes. Ann N Y Acad Sci. 1995 Sep 29;764:50–61. [PubMed]
  • Hufnagle WO, Huang SC, Suzuki I, Milner EC. A complete preimmune human VH3 repertoire. Ann N Y Acad Sci. 1995 Sep 29;764:293–295. [PubMed]
  • Sasso EH, Johnson T, Kipps TJ. Expression of the immunoglobulin VH gene 51p1 is proportional to its germline gene copy number. J Clin Invest. 1996 May 1;97(9):2074–2080. [PMC free article] [PubMed]
  • Pascual V, Liu YJ, Magalski A, de Bouteiller O, Banchereau J, Capra JD. Analysis of somatic mutation in five B cell subsets of human tonsil. J Exp Med. 1994 Jul 1;180(1):329–339. [PMC free article] [PubMed]
  • Lee SK, Bridges SL, Jr, Kirkham PM, Koopman WJ, Schroeder HW., Jr Evidence of antigen receptor-influenced oligoclonal B lymphocyte expansion in the synovium of a patient with longstanding rheumatoid arthritis. J Clin Invest. 1994 Jan;93(1):361–370. [PMC free article] [PubMed]
  • Ziff M. Relation of cellular infiltration of rheumatoid synovial membrane to its immune response. Arthritis Rheum. 1974 May-Jun;17(3):313–319. [PubMed]
  • Iguchi T, Ziff M. Electron microscopic study of rheumatoid synovial vasculature. Intimate relationship between tall endothelium and lymphoid aggregation. J Clin Invest. 1986 Feb;77(2):355–361. [PMC free article] [PubMed]
  • van Dinther-Janssen AC, Pals ST, Scheper R, Breedveld F, Meijer CJ. Dendritic cells and high endothelial venules in the rheumatoid synovial membrane. J Rheumatol. 1990 Jan;17(1):11–17. [PubMed]
  • Randen I, Mellbye OJ, Førre O, Natvig JB. The identification of germinal centres and follicular dendritic cell networks in rheumatoid synovial tissue. Scand J Immunol. 1995 May;41(5):481–486. [PubMed]
  • Girard JP, Springer TA. High endothelial venules (HEVs): specialized endothelium for lymphocyte migration. Immunol Today. 1995 Sep;16(9):449–457. [PubMed]
  • Dechanet J, Merville P, Durand I, Banchereau J, Miossec P. The ability of synoviocytes to support terminal differentiation of activated B cells may explain plasma cell accumulation in rheumatoid synovium. J Clin Invest. 1995 Feb;95(2):456–463. [PMC free article] [PubMed]
  • Schröder AE, Greiner A, Seyfert C, Berek C. Differentiation of B cells in the nonlymphoid tissue of the synovial membrane of patients with rheumatoid arthritis. Proc Natl Acad Sci U S A. 1996 Jan 9;93(1):221–225. [PubMed]
  • Kumon I. In situ characterization of mononuclear cell phenotype in intrahepatic lymphoid follicles in patients with chronic viral hepatitis. Gastroenterol Jpn. 1992 Oct;27(5):638–645. [PubMed]
  • Freni MA, Artuso D, Gerken G, Spanti C, Marafioti T, Alessi N, Spadaro A, Ajello A, Ferraù O. Focal lymphocytic aggregates in chronic hepatitis C: occurrence, immunohistochemical characterization, and relation to markers of autoimmunity. Hepatology. 1995 Aug;22(2):389–394. [PubMed]
  • Mosnier JF, Degott C, Marcellin P, Hénin D, Erlinger S, Benhamou JP. The intraportal lymphoid nodule and its environment in chronic active hepatitis C: an immunohistochemical study. Hepatology. 1993 Mar;17(3):366–371. [PubMed]
  • Steere AC, Duray PH, Butcher EC. Spirochetal antigens and lymphoid cell surface markers in Lyme synovitis. Comparison with rheumatoid synovium and tonsillar lymphoid tissue. Arthritis Rheum. 1988 Apr;31(4):487–495. [PubMed]
  • Berek C, Kim HJ. B-cell activation and development within chronically inflamed synovium in rheumatoid and reactive arthritis. Semin Immunol. 1997 Aug;9(4):261–268. [PubMed]
  • Schröder AE, Sieper J, Berek C. Antigen-dependent B cell differentiation in the synovial tissue of a patient with reactive arthritis. Mol Med. 1997 Apr;3(4):260–272. [PMC free article] [PubMed]

Articles from Molecular Medicine are provided here courtesy of The Feinstein Institute for Medical Research at North Shore LIJ