T Cell Receptor Clonotypes in Skin Lesions from Patients with Systemic Lupus Erythematosus
1998; Elsevier BV; Volume: 110; Issue: 1 Linguagem: Inglês
10.1046/j.1523-1747.1998.00072.x
ISSN1523-1747
AutoresYasuhiko Kita, Kei Kuroda, Tsuneyo Mimori, Takashi Hashimoto, Kazuhiko Yamamoto, Yasushi Saitō, Itsuo Iwamoto, Takayuki Sumida,
Tópico(s)Monoclonal and Polyclonal Antibodies Research
ResumoSystemic lupus erythematosus is an autoimmune disease characterized by the presence of autoantibodies and by lymphocytic infiltration into lesions at several sites such as skin, kidney, and other organs. Immunohistologic studies have clarified that the majority of lymphocytes in the skin are CD4+ αβ T cells. In the present work, to clarify the pathologic role of T cells in the skin of systemic lupus erythematosus patients, we analyzed T cell receptor (TCR) clonotypes of T cells infiltrating into skin lesions. TCR Vβ gene transcripts from T cells from discoid lesions of the skin and peripheral blood lymphocytes of four systemic lupus erythematosus patients were amplified by reverse transcriptase polymerase chain reaction. Southern blot analysis of polymerase chain reaction product demonstrated the heterogeneous TCR Vβ repertoire of T cells in the skin of systemic lupus erythematosus. Single-strand conformation polymorphism analysis showed several distinct bands for smears of most TCR Vβ genes from T cells infiltrating the skin, whereas smears with few bands were found for all TCR Vβ genes from peripheral blood lymphocytes of the same patients. The number of bands encoding each TCR Vβ gene from the skin was significantly higher compared with peripheral blood lymphocytes. Sequencing analysis showed a Leucine-X-Glycine amino acid motif at position 96–98 in the CDR3 region at the frequency of 23–24% in skin-accumulated T cells from two patients, whereas the frequency of this motif in peripheral T cells was only 0–3%, indicating limited T cell epitopes. In conclusion, T cells infiltrating into the skin of systemic lupus erythematosus patients might recognize restricted T cell epitopes on autoantigens and trigger the autoimmune reaction in skin lesions. Systemic lupus erythematosus is an autoimmune disease characterized by the presence of autoantibodies and by lymphocytic infiltration into lesions at several sites such as skin, kidney, and other organs. Immunohistologic studies have clarified that the majority of lymphocytes in the skin are CD4+ αβ T cells. In the present work, to clarify the pathologic role of T cells in the skin of systemic lupus erythematosus patients, we analyzed T cell receptor (TCR) clonotypes of T cells infiltrating into skin lesions. TCR Vβ gene transcripts from T cells from discoid lesions of the skin and peripheral blood lymphocytes of four systemic lupus erythematosus patients were amplified by reverse transcriptase polymerase chain reaction. Southern blot analysis of polymerase chain reaction product demonstrated the heterogeneous TCR Vβ repertoire of T cells in the skin of systemic lupus erythematosus. Single-strand conformation polymorphism analysis showed several distinct bands for smears of most TCR Vβ genes from T cells infiltrating the skin, whereas smears with few bands were found for all TCR Vβ genes from peripheral blood lymphocytes of the same patients. The number of bands encoding each TCR Vβ gene from the skin was significantly higher compared with peripheral blood lymphocytes. Sequencing analysis showed a Leucine-X-Glycine amino acid motif at position 96–98 in the CDR3 region at the frequency of 23–24% in skin-accumulated T cells from two patients, whereas the frequency of this motif in peripheral T cells was only 0–3%, indicating limited T cell epitopes. In conclusion, T cells infiltrating into the skin of systemic lupus erythematosus patients might recognize restricted T cell epitopes on autoantigens and trigger the autoimmune reaction in skin lesions. complementarity determining region 3 Leucine-X-Glycine myelin basic protein peripheral blood lymphocytes single-strand conformation polymorphism Systemic lupus erythematosus (SLE) is a potentially serious autoimmune disease characterized by a broad diversity of clinical features and autoantibodies. B cells that produce antoantibodies seem to be selected by autoantigens and subsequent somatic mutation of IgV regions (Shlomchik et al., 1987Shlomchik M.J. Marshak-Rothstein A. Wolfowicz C.B. Rothstein T.L. Weigert M.G. The role of clonal selection and somatic mutation in autoimmunity.Nature. 1987; 328: 805-811Crossref PubMed Scopus (579) Google Scholar, Shlomchik et al., 1990Shlomchik M. Mascelli M. Shan H. Radic M.Z. Pisetsky D. Marshak-Rothstein A. Weigert M. 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In this study, to address the questions of whether the TCR repertoire of infiltrating T cells in the skin lesions of patients with SLE is restricted and whether these cells expand by antigen stimulation, we analyzed the TCR Vβ gene and TCR clonotypes of infiltrating T cells using polymerase chain reaction (PCR)-Southern blot analysis and subsequent single-strand conformation polymorphism (SSCP) analysis. We obtained evidence that the TCR Vβ repertoire is not limited, but that T cells accumulate clonally in skin lesions, indicating antigen-driven stimulation. Sequencing analysis on skin-specific TCR Vβ genes showed that a Leucine-X-Glycine (LXG) amino acid sequence motif occured preferentially at position 96–98 in the complementarity determining region 3 (CDR3) at the frequency of 23–24% in two of four patients examined, whereas the frequency was only 3% in TCR clones encoding the same Vβ family genes from peripheral blood lymphocytes (PBL). These findings suggest relatively limited T cell epitopes. Possible mechanisms for the progression of SLE are discussed. Four patients with SLE were referred to Chiba University Hospital or Keio University Hospital; all met the criteria for a diagnosis of SLE (Tan et al., 1982Tan E.M. Cohen A.S. Fries J.F. et al.Special article: The 1982 revised criteria for the classification of systemic lupus erythematosus.Arthritis Rheum. 1982; 25: 1271-1277Crossref PubMed Scopus (12087) Google Scholar). All patients were of Japanese ancestry and had skin lesions. None of the patients were related. All four patients had not received treatment, including corticosteroids, in the past. All patients were thought to be “active” because serum complement levels were decreased and anti-DNA antibodies were elevated. Patient LE-2 and LE-4 showed proteinuria. The types of skin lesions that were biopsied from all patients were discoid lesions. The biopsy samples that were obtained from lesional skin included lymphocytic infiltration in all patients. Four disease-free subjects were also examined as controls. The skin biopsy samples did not include lymphocytic infiltration except for a few lymphocytes in control subjects. Typing of the human histocompatibility leukocyte antigens (HLA)-DR and -DQ alleles was performed by PCR combined with dot-blot hybridization with sequence-specific oligonucleotide probes following the protocol of the Eleventh Histocompatibility Workshop (Sumida et al., 1994Sumida T. Sakamaki T. Yonaha F. et al.HLA-DR alleles in patients with Sjögren’s syndrome over-representing Vβ2 and Vβ13 genes in the labial salivary glands.Br J Rheum. 1994; 33: 420-424Crossref PubMed Scopus (19) Google Scholar). Skin biopsy samples were stained with hematoxylin and eosin for histologic examination. For immunohistologic examination, a portion of a sample was snap frozen, and cryostat sections were cut and stained with anti-CD4 and anti-CD8 monoclonal antibodies (Becton Dickinson, Mountain View, CA). Cryostat sections were incubated with biotinylated rabbit anti-mouse immunoglobulins (Dako, Copenhagen, Denmark), then with streptABComplex/HRP (Dako), and finally with a peroxidase substrate. Total RNA (5–10 mg) from skin specimens and PBL of four SLE patients and four healthy subjects was prepared using Isogen solution (Nippon Gene, Tokyo, Japan). The first strand complementary DNA (cDNA) was synthesized in a 20 ml reaction mixture from 5 mg of total RNA using a cDNA synthesis module (Amersham International, Buckinghamshire, U.K.). Amplification was performed with Taq polymerase in 50 ml of standard buffer using 0.2 ml of cDNA (corresponding to 50 ng of total RNA) with 20 different Vβ and Cβ primers to detect each Vβ gene. The sequences of the Vβ and Cβ primers were obtained from previously published data (Sumida et al., 1992Sumida T. Yonaha F. Maeda T. Tanabe E. Koike T. Tomioka H. Yoshida S. T cell receptor of infiltrating T cells in lips of Sjögren’s syndrome patients.J Clin Invest. 1992; 89: 681-685Crossref PubMed Scopus (135) Google Scholar). Oligonucleotides were synthesized with a DNA synthesizer (Applied Biosystems, Foster City, CA). The denaturing step was carried out at 95°C for 1.5 min, the annealing step at 60°C for 1 min, and the extension step at 72°C for 1 min for 30 cycles on a DNA thermal cycler (Perkin-Elmer, Norwalk, CT). One-fifth of the PCR product was subjected to 2% agarose gel electrophoresis and hybridized with a 32P-labeled Pst I fragment of the JUR-β gene (Cβ) probe (Sumida et al., 1992Sumida T. Yonaha F. Maeda T. Tanabe E. Koike T. Tomioka H. Yoshida S. T cell receptor of infiltrating T cells in lips of Sjögren’s syndrome patients.J Clin Invest. 1992; 89: 681-685Crossref PubMed Scopus (135) Google Scholar). Amplified DNA was diluted (1:2) in a denaturing solution (95% formamide, 10 mM ethylenediamine tetraacetic acid, 0.1% bromophenol blue, 0.1% xylene cyanol) and incubated at 90°C for 2 min. The diluted sample (6 ml) was electrophoresed in nondenaturing 5% polyacrylamide gels containing 10% glycerol to discriminate the three-dimensional conformations of the amplified TCR Vβ genes (Yamamoto et al., 1992Yamamoto K. Sakoda H. Nakajima T. et al.Accumulation of multiple T cell clonotypes in the synovial lesions of patients with rheumatoid arthritis revealed by a novel clonality analysis.Int Immunol. 1992; 4: 1219-1223Crossref PubMed Scopus (149) Google Scholar). The gel was run at 35 W constant power for 2.5 h. After electrophoresis, the DNA was transferred to Immobilon-S (Millipore Intertech, Bedford, MA) and hybridized with biotinylated Cb probe (5'-A(AC)AA(GC)GTGTTCCCACCCGAGGTCGC-TGTGTT-3'). The DNA was then visualized with a Phototope detection kit (New England Biolabs, Beverly, MA). Complementary DNA encoding the TCR genes from clonally expanded T cells in the skin were eluted from the SSCP gels and amplified by secondary PCR using inside primers with an EcoRI enzyme site sensitive for each Vβ gene. The sequences of the Vβ and Cβ inside primers were as follows:Vβ1 (5'-TCTAGAATTCGATTCTCCGCACAACAGT-3')Vβ2 (5'-TCTAGAATTCAGAAGGACAAGTTTCTCA-3')Vβ4 (5'-TCTAGAATTCTTGACAAGTTTCCCATCA-3')Vβ5 (5'-TCTAGAATTCTTCTCAGGGCGCCAGTTC-3')Vβ6 (5'-TCTAGAATTCCGGTTCTTTGCAGTCAGG-3')Vβ7 (5'-TCTAGAATTCCCTGAATGCCCCAACAGC-3')Vβ8 (5'-TCTAGAATTCGATTCTCAGCTAAGATGC-3')Vβ12 (5'-TCTAGAATTCCTCTAGATCAAAGACAGAGG-3')Vβ13.1 (5'-TCTAGAATTCACAGAGGATTTCCCGCTC-3')Vβ13.2 (5'-TCTAGAATTCCAGAATTTCCTGCTGGGG-3')Vβ14 (5'-TCTAGAATTCCTCTCGAAAAGAGAAGAGGA-3')Vβ15 (5'-TCTAGAATTCTACAGTGTCTCTCGACAG-3')Vβ17 (5'-TCTAGAATTCAGCGTCTCTCGGGAGAAG-3')Vβ19 (5'-TCTAGAATTCCTCAATGCCCCAAGAACGC-3')Cβ (5'-TCTAGAATTCAGCGACCTCGGGTGGGAA-3') Oligonucleotides were synthesized with a DNA synthesizer (Applied Biosystems). PCR products were purified by phenol extraction, precipitated with ethanol, and digested with excess amounts of EcoRI. The DNA fragments were ligated to M13 mp19 phage vectors obtained by EcoRI digestion. Phages were grown on TG-1 Escherichia coli cells. After hybridization with a Cβ probe, a single phage was allowed to grow, and recombinant phage DNA was purified for DNA sequence determination. Three different plaques encoding TCR genes from one band on SSCP gel were sequenced independently and these DNA showed the identical TCR sequence. Sequencing reactions were performed by automated sequencing (Applied Biosystems). Statistical analysis of the results was carried out by the unpaired t test. Histologic examination (hematoxylin and eosin staining) of skin lesions of patients with SLE was performed. The epidermal basal layer showed focal liquefaction degeneration. Lymphocytic infiltration was observed in the upper dermis, periappendageal, and perivascular areas. Immunohistochemical studies using monoclonal antibodies against CD4 or CD8 confirmed that the majority of infiltrating lymphocytes in skin were CD4+ T cells. The HLA-DR and DQ typing of the individual patients was performed (data not shown). HLA-DR2 (DRB1*1502, DRB1*1602, DRB5*0102, and DRB5*02) was found in all patients. Patient LE-3 is an HLA-DR2/DR4 heterozygote. In two patients (LE-1 and LE-4), DRB3*0202 was found. To analyze the mechanism of skin manifestations in patients with SLE, we examined the TCR repertoire of infiltrating T cells in the skin using the family PCR method (Fig 1). Four skin biopsy specimens from four SLE patients with skin manifestations were analyzed. As control, four PBL samples from the same individuals were used. Most TCR Vβ genes on T cells were detected in the skin, whereas all TCR Vβ genes were detected in PBL. These results suggest that the TCR Vβ repertoire of T cells in the skin is heterogenous as well as PBL and there were no common predominant Vβ genes. To analyze T cell clonotypes in the skin lesions from patients with SLE, TCR Vβ clonotypes in the skin and on PBL of four SLE patients were examined by the PCR-SSCP method. Figure 2 shows several distinct bands in most Vβ genes from T cells infiltrating the skin, whereas smear was found for all Vβ genes from PBL. As shown in Table I, the numbers of bands encoding TCR Vβ genes from skin lesions (5.3 ± 0.7, 5.7 ± 1.0, 7.4 ± 1.2, and 6.4 ± 1.0; means ± SEM) were significantly higher (p < 0.001) in all of four SLE patients compared with PBL (2.5 ± 0.4, 1.8 ± 0.3, 3.2 ± 0.4, and 2.8 ± 0.4). The numbers of T-cell clones expressing the TCR Vβ1, Vβ2, Vβ4, Vβ5, Vβ7, Vβ12, and Vβ13 genes (6.5 ± 0.5, 10.3 ± 1.7, 8.3 ± 0.7, 7.0 ± 0.6, 6.3 ± 0.3, 7.5 ± 0.5, and 11.5 ± 2.2) were specifically increased in the skin compared with PBL (1.8 ± 0.6, 2.3 ± 0.3, 1.5 ± 1.0, 3.8 ± 0.5, 3.5 ± 0.3, 3.3 ± 0.7, and 4.3 ± 0.6) (p < 0.05 to p < 0.005). These results suggest that some T cells in skin lesions of SLE patients expand clonally, indicating antigen stimulation. In contrast, the numbers of bands from skin in all four healthy subjects (Table II; 4.2 ± 1.0, 3.3 ± 0.7, 4.5 ± 0.9, and 5.3 ± 0.8) were not significantly higher compared with PBL (2.3 ± 0.6, 2.8 ± 0.6, 4.7 ± 0.8, and 3.3 ± 0.7). The number of bands from skin in SLE patients (6.1 ± 0.5) was significantly higher than that in healthy subjects (4.4 ± 0.4) (p < 0.01).Table INumbers of accumulated clonotypes of TCR Vβ genes in T cells from patients with SLEVβ1234567891011121314151617181920means ± SEMLE-1PBL323036402224443034002.5 ± 0.4skin6–eNeither smear nor distinct bands are detected.–9736523471343–53555.3±0.7dp < 0.001.LE-2PBL0–104132342–320–13101.8 ± 0.3skin–123––66342––853–11–––5.7 ± 1.0dp < 0.001.LE-3PBL223256466412453033213.2 ± 0.4skin–12–96778–1–8178––4–2–7.4± 1.2dp < 0.001.LE-4PBL232432312414662044112.8 ± 0.4skin77–7810–244––812–––––16.4± 1.0dp < 0.001.MeanPBL1.82.32.31.53.83.83.52.33.33.51.53.34.34.32.00.02.83.51.00.5skin6.5bp < 0.01.10.3bp < 0.01.3.08.3cp < 0.005.7.0bp < 0.01.6.56.3cp < 0.005.4.53.32.54.07.5ap < 0.05.11.5ap < 0.05.7.33.0–6.73.03.53.0a p < 0.05.b p < 0.01.c p < 0.005.d p < 0.001.e Neither smear nor distinct bands are detected. Open table in a new tab Table IINumbers of accumulated clonotypes of TCR Vβ genes in T cells from healthy subjectsVβ1234567891011121314151617181920means ± SEMCont-1PBL410511484314010–aNeither smear nor distinct bands are detected.0–––2.3 ± 0.6skin4302–141141–719––8–––4.2 ± 1.0Cont-2PBL4–0022294214260–52–12.8 ± 0.6skin5113––292127–4––3–––3.3 ± 0.7Cont-3PBL70751–887––4083–71––4.7 ± 0.8skin8–391–885215080–81––4.5 ± 0.9Cont-4PBL5–031–18221562––73––3.3 ± 0.7skin71472111864–708––8–––5.3 ± 0.8MeanPBL5.00.51.83.31.31.03.88.34.32.31.04.32.04.31.0–4.82.0–1.0skin6.01.72.05.31.51.36.39.04.32.01.56.5bp < 0.01.0.37.30.0–6.81.0–3.0a Neither smear nor distinct bands are detected.b p < 0.01. Open table in a new tab As shown in Fig 2 and Table I, several T cell clones accumulate in the skin but not in peripheral blood. We selected skin-specific T cell clones on SSCP gel in Fig 2 and DNA encoding the TCR genes from the skin-specific bands were eluted from the gels and sequenced. To examine the amino acid sequences of the TCR Vβ region of these skin-specific T cell clones, DNA-encoding TCR genes were eluted from the skin-specific bands on SSCP and sequenced. Three different plaques encoding TCR genes from one band on SSCP gel were sequenced independently and these DNA showed the identical TCR sequence. In one patient (LE-1), the numbers of T cell clones expressing TCR Vβ4, Vβ5, Vβ13, and Vβ19 genes (9, 7, 13, and 5) in the skin were specifically increased in comparison with PBL (0, 3, 4, and 0) and there were no identical TCR in skin and PBL. As shown in Table III, an LXG amino acid sequence motif was found at position 96–98 in CDR3 for five of 21 clones (24%). Leucine (L) was conserved at position 96 or 97 in seven of 21 (33%). In another patient (LE-4), the numbers of T cell clones expressing TCR Vβ2, Vβ5, Vβ6, and Vβ13 genes (7, 8, 10, and 8) in the skin were increased in comparison with the numbers in PBL (3, 3, 2, and 6), and there were no identical TCR between skin and PBL. An LXG sequence motif was also found in position 96–98 at a frequency of 23%. Leucine was conserved at position 96 or 97 in seven of 13 clones (54%). In the other two patients (LE-2 and LE-3) there was no conserved amino acid sequence motif in CDR3 on skin-specific T cells. Contrary to SLE patients, the LXG sequence motif (position 96–98) was not found in the skin from all of four healthy subjects (data not shown). In healthy subjects, leucine at position 96 or 97 on skin-specific T cells was found in two of 13 clones (15%) (Control 1), two of eight clones (25%) (Control 2), two of nine clones (22%) (Control 3), and one of 11 clones (9%) (Control 4). The LXG sequence motif (position 96–98) and leucine at position 96 or 97 were found at higher frequency in the SLE skin (LE-1 and LE-4) than in normal skin. To analyze whether the LXG amino acid sequence motif in CDR3 is specific to the skin, cDNA encoding TCR Vβ genes from PBL of two patients (LE-1 and LE-4) were cloned and sequenced (data not shown). An LXG sequence motif in CDR3 was found in only one of 36 clones (2.8%) (LE-1) or none of 19 clones (0%) (LE-4). Leucine at position 96 or 97 was found in only four of 36 clones (11%) (LE-1) or two of 19 clones (11%) (LE-4). The LXG sequence motif (position 96–98) and leucine at position 96 or 97 were found at high frequency in the skin from these patients (LE-1 and LE-4). These results suggest that T cells that infiltrate the skin might recognize relatively limited epitopes on the autoantigen in skin.Table IIIJunctional sequences of TCR β genes of skin-specific T cells from patients with SLEaThe single letter amino acid sequences at the 3' position of TCR Vβ, CDR3, and the 5' position of theJ region are given.PatientCloneVN-D-NJJβ95106LE-1Vβ4-1L C S VD QGV G VbLeucines (L) at position 96 or 97 are in bold type.T E A F F GJβ1.1Vβ4-2L C S VP S G I GY N E Q F F GJβ2.1Vβ4-3L C S VP R D TY N E Q F F GJβ2.1Vβ4-4L C S V EG P SG IY N E Q F F GJβ2.1Vβ4-5L C S A GL A G A EcAmino acids encoded by Dβ genes are in italic letters., dLXG amino acid sequence motifs at position 96–98 in the CDR3 are underlined.E T Q Y F GJβ2.5Vβ4-6L C S ATGA P V KE Q Y F GJβ2.7Vβ5-1L C A S SL R G GSTDTQYFGJβ2.3Vβ5-2L C A S SS QGA TT D T Q Y F GJβ2.3Vβ5-3L C A S SL A GG P SD T Q Y F GJβ2.3Vβ5-4L C A S SRTSGRL GD T Q Y F GJβ2.3Vβ5-5L C A S SL Q GA TT D T Q Y F GJβ2.3Vβ5-6L C A S SF L AV G P G GD T Q Y F GJβ2.3Vβ5-7L C A S SH R DR G TY E Q Y F GJβ2.7Vβ13-1F C A S SDEWTGN Q P Q H F GJβ1.5Vβ13-2F C A S SH RG QN E R F F GJβ2.1Vβ13-3F C A SR L SGTNHN E Q F F GJβ2.1Vβ13-4F C A SR L ALSTDTQYFGJβ2.3Vβ13-5F C A S SD S S A YE Q Y F GJβ2.7Vβ19-1L C ATYAVDW GQGCDE Q F F GJβ2.1Vβ19-2L C A S SS R A TG RNTGELFFGJβ2.2Vβ19-3L C AR P T Y PSTDTQYFGJβ2.3LE-2Vβ2-1I C S AT S N RG A AG E L F F GJβ2.2Vβ2-2I C S AN E G GE Q Y F GJβ2.7Vβ7-1L C A S S QD Q H PE A F F GJβ1.1Vβ7-2L C A S S QG IE Q F F GJβ2.1Vβ7-3L C AT H C E GMSTDTQYFGJβ2.3Vβ7-4L C ATHEEGTSGFST Q Y F GJβ2.3Vβ13F C A S SF R S Q Y TG AN T E A F F GJβ1.1Vβ15F C A TK D S R AG E L F F GJβ2.2Vβ17-1L C A S SI W P SGHE Q Y F GJβ2.1Vβ17-2L C A S SR R E VY E Q Y F GJβ2.7LE-3Vβ2-1I C S AIQG P PS Y E Q Y F GJβ2.7Vβ2-2I C S AR G Q P GE Q Y F GJβ2.7Vβ4-1L C S V EG GTSTDTQYFGJβ2.3Vβ4-2L C S AK EE T Q Y F GJβ2.5Vβ6L CASSL A S V Q V TY N E Q F F GJβ2.1Vβ8-1F C A S SQ G P DN S P L H F GJβ1.6Vβ8-2F C A SRGTDTQYFGJβ2.3Vβ13F C A S SR LA D EE T Q Y F GJβ2.5LE-4Vβ2-1I C S AR D E SSYNEQFFGJβ2.1Vβ2-2I C S ARG L A G AN E Q F F GJβ2.1Vβ2-3I C S AR DR E SD T Q Y F GJβ2.3Vβ2-4I C S AI R G P PS Y E Q Y F GJβ2.7Vβ5-1L C A S SL R G R ST D T Q Y F GJβ2.3Vβ5-2L C ATA L G GG R AD T Q Y F GJβ2.3Vβ6-1L C A S SL T S G DT E A F F GJβ1.1Vβ6-2L C A S SL A S A H V TY N E Q F F GJβ2.1Vβ6-3L C A S SL T AG G L GD T Q Y F GJβ2.3Vβ6-4L C A S SL T T GA GT D T Q Y F GJβ2.3Vβ6-5L C A S SS G AG G GD T Q Y F GJβ2.3Vβ13-1F C A S SY S A V L NG E L F F GJβ2.2Vβ13-2F C A SR F A LSTDTQYFGJβ2.3a The single letter amino acid sequences at the 3' position of TCR Vβ, CDR3, and the 5' position of the J region are given.b Leucines (L) at position 96 or 97 are in bold type.c Amino acids encoded by Dβ genes are in italic letters.d LXG amino acid sequence motifs at position 96–98 in the CDR3 are underlined. Open table in a new tab Activated T cells have been detected in the peripheral blood of SLE patients (Alocer-Varela et al., 1991Alocer-Varela J. Alarcon-Riquelme M. Laffon A. Sanchez-Madrid F. Alarcon-Sgovia D. Activation markers on peripheral blood T cells from patients with active or inactive systemic lupus erythematosus. Correlation with proliferative responses and production of IL-2.J Autoimmun. 1991; 4: 935-945Crossref PubMed Scopus (23) Google Scholar) and may mediate B cell autoantibody production (Shivakumar et al., 1989Shivakumar S. Tsokos G.C. Datta S.K. T cell receptor α/β expressing double-negative (CD4−/CD8−) and CD4+ T helper cells in humans augment the production of pathogenic anti-DNA autoantibodie associated with lupus nephritis.J Immunol. 1989; 143: 103-112PubMed Google Scholar; Rajagopalan et al,1990; Murakami et al., 1992Murakami M. K
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