Human Immunology 75 (2014) 466–469
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www.ashi-hla.org
journal homepage: www.elsevier.com/locate/humimm
CXCR3 polymorphism is associated with male gender and pleuritis in patients with systemic lupus erythematosus Churl Hyun Im a, Ji Ah Park a, Jeong Yeon Kim a, Eun Young Lee a, Eun Bong Lee a, Yeonjung Kim b,⇑, Yeong Wook Song a,c,⇑ a
Division of Rheumatology, Department of Internal Medicine, Seoul National University Medical College, Seoul, Republic of Korea Center for Genomic Science, The National Institute of Health, Seoul, Republic of Korea WCU Department of Molecular Medicine and Biopharmaceutical Sciences, Medical Research Center, Seoul National University College of Medicine, 103 Daehak-ro, Jongno-gu, 110-799 Seoul, Republic of Korea b c
a r t i c l e
i n f o
Article history: Received 29 May 2013 Accepted 4 February 2014 Available online 12 February 2014
a b s t r a c t Background: Systemic lupus erythematosus (SLE) is a chronic inflammatory autoimmune disease involving multiple organs. Chemokines and their receptors play an important role in the pathogenesis of SLE. Lymphocytes expressing CXCR3, chemokine receptors of CXCL4, 9, 10, and 11, increase in patients with SLE and animal models, particularly in those with skin manifestations and nephritis. We investigated CXCR3 genetic polymorphisms in patients with SLE and their association with clinical manifestations. Methods: A total of 346 patients with SLE and 540 healthy controls were investigated for CXCR3 intron 1 polymorphisms rs2280964 and rs34334103 by Taqman analysis. Results: rs2280964 and rs34334103 were not associated with all patients with SLE, but rs34334103 showed a significant association with male patients with SLE. Among the clinical manifestations, pleuritis was associated with the rs34334103 polymorphism. Conclusion: The CXCR3 polymorphism rs34334103 was associated with male gender and pleuritis in patients with SLE. Ó 2014 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.
1. Introduction Systemic lupus erythematosus (SLE) is a chronic inflammatory autoimmune disease of multiple organ systems. The clinical manifestations and organ involvement of SLE are polymorphic; however, a common pathological finding is inflammatory cell infiltrations in affected tissue, which are introduced via chemotaxis by chemokines and their receptors. CXCR3 is a G protein-coupled cell surface receptor that has been identified as a receptor for
Abbreviations: SLE, systemic lupus erythematosus; SLEDAI, systemic lupus erythematosus disease activity index; SNP, single nucleotide polymorphism; DNA, deoxyribonucleic acid; PCR, polymerase chain reaction; OR, odds ratio; CI, confidence interval; TLR, toll-like receptor. ⇑ Corresponding authors. Address: Center for Genomic Science, Korea National Institute of Health, Osong Health Technology Administration Complex, 187 Osongsaengmyeong2(i)-ro, Osong-eup, Cheongwon-gun, Chungcheongbuk-do 363-951, Republic of Korea (Y. Kim). Address: Division of Rheumatology, Department of Internal Medicine, Department of Immunology, Medical Research Center, Seoul National University College of Medicine, 103 Daehak-ro, Jongno-gu, Seoul 110-799, Republic of Korea. Fax: +82 2 762 9662 (Y.W. Song). E-mail addresses: [email protected] (Y. Kim), [email protected] (Y.W. Song).
T-lymphocyte-specific CXC chemokines including CXCL4, 9, 10, and 11. CXCR3 is preferentially expressed on activated effector T lymphocytes, although it has also been observed on subsets of B lymphocytes and natural killer cells [1]. CXCL10 and its receptor CXCR3 are associated with the Th1 and interferon-c response and have received attention as a pathogenic chemokine and receptor in SLE [2]. CXCL10 levels increase in serum of patients with SLE compared to those in the general population and are associated with disease activity, such as anti-double stranded DNA antibody titer, serum complement level, and SLE disease activity index (SLEDAI) score [3,4]. CXCR3-expressing CD4 T lymphocytes in peripheral blood of patients with active SLE decrease compared to that in patients with inactive SLE and a control group and show a trend toward a negative correlation with SLEDAI score [5]. CXCR3-expressing T lymphocytes increase in skin lesions of patients with SLE but decrease in peripheral blood lymphocytes, which reflects recruitment of CXCR3-expressing lymphocytes into the skin [6]. CXCL9, 10, and 11 mRNA expression increase, and their specific receptor CXCR3 is expressed by infiltrating lymphocytes in discoid LE tissues [7]. T cells expressing CXCR3 increase in peripheral blood of patients with lupus and renal involvement [8], and infiltrate in
http://dx.doi.org/10.1016/j.humimm.2014.02.005 0198-8859/Ó 2014 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.
C.H. Im et al. / Human Immunology 75 (2014) 466–469
the tubule-interstitium in renal tissue from patients with lupus nephritis, which correlates with kidney dysfunction [9]. In an animal model of lupus nephritis, immune-mediated kidney disease is promoted by CXCR3 and one of its ligands, CXCL9, but not CXCL10, via activated T cell and macrophage recruitment into the kidney [10]. Th1 and Th17 cell infiltration into the inflamed kidney and morphological and functional damage of nephritic kidney are suppressed in a CXCR3 knock-out lupus nephritis mice model (MRL/ lpr) [10,11]. CXCR3 expression in urinary CD4 T cells correlates with SLEDAI scores in patients with lupus nephritis [12]. CXCR3bearing memory B cell subset is related to autoantibody production and adverse clinical outcomes in SLE [13]. The CXCR3 gene (NC_000023.10) is located on the X chromosome (Xq13). Four single nucleotide polymorphism (SNP) sites were located within the CXCR3 gene in a previous Korean population study [14]. According to the NCBI database, the frequencies of the other CXCR3 gene polymorphism are extremely low (frequency < 0.05), except rs2280964, rs34334103, and rs111516068. The first two SNPs were evaluated in Korean patients with allergic asthma, but the last SNP (rs111516068) was not detected in the Korean population [14,15]. In this study, we investigated CXCR3 polymorphisms in patients with SLE, and their association with clinical manifestations.
2. Methods A total of 346 patients with SLE were enrolled at the Rheumatology Clinic, Seoul National University Hospital, Korea. They were diagnosed by SLE classification criteria, which were revised in 1997 by the American College of Rheumatology [16]. After informed consent, the patient’s peripheral blood was collected, and medical records were reviewed as follows: age, sex, mucocutaneous (malar rash, discoid rash, oral ulcer, and other skin manifestations), serosa (pleuritis and pericarditis), renal (proteinuria, 24 h collected urine protein amount, creatinine clearance, and renal histology), hematologic (leukopenia, lymphopenia, hemolytic anemia, and thrombocytopenia), arthritis, neuropsychiatric (psychosis and seizure), anti-nuclear antibody, and other autoantibodies (anti-double stranded DNA antibody, anti-Smith antibody, lupus anticoagulant, anti-cardiolipin antibody IgG/IgM, anti-Ro (SSA) antibody, and anti-La (SSB) antibody). This study was approved by the institutional review board of the hospital. A total of 540 healthy control DNA samples were collected from hospital workers during their annual medical checkup. Two CXCR3 polymorphism sites in intron 1, rs2280964 and rs3433413, were investigated by Taq-man analysis, as shown in previous studies [14,15]. Primer Express (Applied Biosystems, Foster City, CA, USA) was used to design both the polymerase chain reaction (PCR) primers and the MGB Taqman probe. One allelic probe was labeled with FAM dye and the other with fluorescent VIC dye. The primer and probe sequences are available on the website (http:// www.snp-genetics.com/reference/CXCR3_additional_information. doc). The PCR was run with TaqMan Universal Master mix without UNG (Applied Biosystems) at a primer concentration of 900 nM and a TaqMan MGB-probe concentration of 200 nM. The reaction was performed in a 384-well format in a total reaction volume of 5 ll using 20 ng genomic DNA. The plate was then placed in a thermal cycler (PE 9700, Applied Biosystems) and heated for 2 min at 50 °C and for 10 min at 95 °C followed by 40 cycles of 95 °C for 15 s and 60 °C for 1 min. The TaqMan assay plate was then transferred to a Prism 7900HT instrument (Applied Biosystems) where the fluorescence intensity of each well was read. Fluorescence data files from each plate were analyzed using automated software (SDS 2.1). Baseline statistics was analyzed with logistic regression models controlling for age and sex. False discovery rate adjusted P-values
467
(Pc) were calculated for the multiple comparison correction. The association with each SLE manifestation and the laboratory results were analyzed with SPSS program version 12.0 K (SPSS Inc., Chicago, IL, USA). The Haploview program version 4.2, (Broad Institute of MIT and Harvard, Cambridge, MA, USA) was used for the haplotype analysis with the two loci of the CXCR3 polymorphisms. 3. Results A total of 346 patients with SLE and 540 healthy controls were analyzed. The mean ages (± standard deviation) of patients with SLE and healthy controls were 35.6 ± 13.1 and 43.3 ± 15.6 years, respectively. The male gender ratios of the groups were 9.2% vs. 12.4% respectively. Neither polymorphism was significantly different between the SLE and normal control groups (Table 1). However, after stratification by gender, the A allele of the rs34334103 polymorphism was associated with male patients with SLE compared to that in male controls (Pc value < 0.001, odds ratio [OR] 14.279; 95% confidence interval [CI] 4.153–49.095) (Table 2). In the haplotype analysis for male gender (rs2280964-rs34334103), both polymorphisms were in linkage disequilibrium (D’ = 1.000, r2 = 0.115), and the CA haplotype (C allele from rs2280964 and A allele from rs34334103) was significantly associated with an increased risk of male lupus compared to that in the male control group (Pc value = 0.002; OR 10.833; 95% CI 2.147–54.669) (Table 2). In females, neither polymorphism was different between the SLE and control groups, and the haplotype analysis was not significant either (Table 2). Mucocutaneous, musculoskeletal, hematological, renal, and neurologic manifestations were not associated with the CXCR3 polymorphisms in patients with SLE. However, the A allele of CXCR3 SNP rs34334103 increased significantly in patients with pleuritis (Pc value = 0.013; OR 2.250; 95% CI 1.228–4.122) (Table 3). Both polymorphisms were in linkage disequilibrium (D’ = 0.913, r2 = 0.102), and haplotype CA was associated with pleuritis (Pc value = 0.009; OR 2.415; 95% CI 1.309–4.453) (Table 3). Significance disappeared in male patients with pleuritis after stratifying by gender, but not in female patients (Supplementary Table 1). Among laboratory abnormalities, anti-phospholipid antibody IgG tended to be associated with both rs2280964 and rs34334103 polymorphisms (P value = 0.038 and 0.046, respectively, but was not significant after correction of multiple comparison). Anti-SSB (La) antibodies were associated with the A allele of rs34334103 (Pc value = 0.046). The other laboratory results such as antinuclear antibody, anti-double stranded DNA antibody, anti-Smith antibody, lupus anticoagulant, anti-cardiolipin antibody IgG/M, anti-Ro (SSA) antibody, 24 h collected urine protein amount, and creatinine clearance were not associated with the polymorphisms or haplotypes. 4. Discussion CXCR3 is a receptor for chemokines CXCL4, 9, 10, and 11, which are involved in lymphocyte chemotaxis to target organs in patients with SLE. CXCL9, CXCL10, and their receptor CXCR3 are involved with target tissues such as the skin and kidney and are associated with disease activity in SLE [3–9]. Taken together, CXCR3-bearing autoreactive T lymphocytes are recruited from the circulation to the site of autoimmune attack by chemokine CXCR9 or CXCL10, released from local cells at target tissues during the initiating event of SLE, and the CXCR3-brearing memory B cell subset is engaged in the extended stage of the disease [10]. Despite the importance of CXCR3 in autoimmune diseases, CXCR3 polymorphism studies have been rarely reported. Kato et al. studied variations in the coding region of human CXCR1, 2,
468
C.H. Im et al. / Human Immunology 75 (2014) 466–469
Table 1 CXCR3 polymorphisms in patients with systemic lupus erythematosus (SLE) and healthy controls. Polymorphisms
SLE (n = 346) (%)
Controls (n = 540) (%)
Pc valuea
OR
(95% CI)
NS
1.066
(0.875–1.298)
rs2280964
C allele T allele
50.0 50.0
50.5 49.5
rs34334103
G allele A allele
88.4 11.6
90.6 9.4
NS
1.283
(0.933–1.764)
Haplotypeb
TG CG CA TA
49.6 39.5 10.6 0.4
49.4 40.8 9.8 0.1
1.000 0.933 0.935 0.400
1.008 0.950 1.095 4.614
(0.828–1.226) (0.906–1.161) (0.793–1.513) (0.479–44.451)
NS, Not significant. a Pc-values for the allele frequencies were adjusted for age and sex (correction by false delivery rate for multiple comparisons), and those of each haplotype were corrected by the permutation test. b Haplotypes consisted of the rs2280964 and rs34334103 CXCR3 polymorphisms in linkage disequilibrium (D’ = 0.962, r2 = 0.104).
Table 2 CXCR3 polymorphism analysis between patients with systemic lupus erythematosus (SLE) and healthy controls stratified by gender. Gender
Polymorphisms
Male
rs2280964 rs34334103 Haplotypeb
Female
rs2280964
rs34334103
Haplotypeb
SLE (male 32, female 314) (%)
Controls (male 67, female 473) (%)
C allele T allele G allele A allele TG CG CA TA
50.0 50.0 75.0 25.0 50.0 25.0 25.0 0.0
49.3 50.7 97.0 3.0 50.7 46.3 3.0 0.0
CC CT TT C allele T allele GG GA AA G allele A allele TG CG CA TA
70 (22.4) 172 (55.1) 70 (22.4) 50.0 50.0 252 (80.5) 58 (18.5) 3 (1.0) 89.8 10.2 49.5 40.2 9.8 0.4
129 (27.6) 216 (46.2) 123 (26.3) 49.3 50.7 380 (80.3) 88 (18.6) 5 (1.1) 89.6 10.4 49.3 40.4 10.3 0.1
Pc valuea
OR
NS
0.752
Contents lists available at ScienceDirect
www.ashi-hla.org
journal homepage: www.elsevier.com/locate/humimm
CXCR3 polymorphism is associated with male gender and pleuritis in patients with systemic lupus erythematosus Churl Hyun Im a, Ji Ah Park a, Jeong Yeon Kim a, Eun Young Lee a, Eun Bong Lee a, Yeonjung Kim b,⇑, Yeong Wook Song a,c,⇑ a
Division of Rheumatology, Department of Internal Medicine, Seoul National University Medical College, Seoul, Republic of Korea Center for Genomic Science, The National Institute of Health, Seoul, Republic of Korea WCU Department of Molecular Medicine and Biopharmaceutical Sciences, Medical Research Center, Seoul National University College of Medicine, 103 Daehak-ro, Jongno-gu, 110-799 Seoul, Republic of Korea b c
a r t i c l e
i n f o
Article history: Received 29 May 2013 Accepted 4 February 2014 Available online 12 February 2014
a b s t r a c t Background: Systemic lupus erythematosus (SLE) is a chronic inflammatory autoimmune disease involving multiple organs. Chemokines and their receptors play an important role in the pathogenesis of SLE. Lymphocytes expressing CXCR3, chemokine receptors of CXCL4, 9, 10, and 11, increase in patients with SLE and animal models, particularly in those with skin manifestations and nephritis. We investigated CXCR3 genetic polymorphisms in patients with SLE and their association with clinical manifestations. Methods: A total of 346 patients with SLE and 540 healthy controls were investigated for CXCR3 intron 1 polymorphisms rs2280964 and rs34334103 by Taqman analysis. Results: rs2280964 and rs34334103 were not associated with all patients with SLE, but rs34334103 showed a significant association with male patients with SLE. Among the clinical manifestations, pleuritis was associated with the rs34334103 polymorphism. Conclusion: The CXCR3 polymorphism rs34334103 was associated with male gender and pleuritis in patients with SLE. Ó 2014 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.
1. Introduction Systemic lupus erythematosus (SLE) is a chronic inflammatory autoimmune disease of multiple organ systems. The clinical manifestations and organ involvement of SLE are polymorphic; however, a common pathological finding is inflammatory cell infiltrations in affected tissue, which are introduced via chemotaxis by chemokines and their receptors. CXCR3 is a G protein-coupled cell surface receptor that has been identified as a receptor for
Abbreviations: SLE, systemic lupus erythematosus; SLEDAI, systemic lupus erythematosus disease activity index; SNP, single nucleotide polymorphism; DNA, deoxyribonucleic acid; PCR, polymerase chain reaction; OR, odds ratio; CI, confidence interval; TLR, toll-like receptor. ⇑ Corresponding authors. Address: Center for Genomic Science, Korea National Institute of Health, Osong Health Technology Administration Complex, 187 Osongsaengmyeong2(i)-ro, Osong-eup, Cheongwon-gun, Chungcheongbuk-do 363-951, Republic of Korea (Y. Kim). Address: Division of Rheumatology, Department of Internal Medicine, Department of Immunology, Medical Research Center, Seoul National University College of Medicine, 103 Daehak-ro, Jongno-gu, Seoul 110-799, Republic of Korea. Fax: +82 2 762 9662 (Y.W. Song). E-mail addresses: [email protected] (Y. Kim), [email protected] (Y.W. Song).
T-lymphocyte-specific CXC chemokines including CXCL4, 9, 10, and 11. CXCR3 is preferentially expressed on activated effector T lymphocytes, although it has also been observed on subsets of B lymphocytes and natural killer cells [1]. CXCL10 and its receptor CXCR3 are associated with the Th1 and interferon-c response and have received attention as a pathogenic chemokine and receptor in SLE [2]. CXCL10 levels increase in serum of patients with SLE compared to those in the general population and are associated with disease activity, such as anti-double stranded DNA antibody titer, serum complement level, and SLE disease activity index (SLEDAI) score [3,4]. CXCR3-expressing CD4 T lymphocytes in peripheral blood of patients with active SLE decrease compared to that in patients with inactive SLE and a control group and show a trend toward a negative correlation with SLEDAI score [5]. CXCR3-expressing T lymphocytes increase in skin lesions of patients with SLE but decrease in peripheral blood lymphocytes, which reflects recruitment of CXCR3-expressing lymphocytes into the skin [6]. CXCL9, 10, and 11 mRNA expression increase, and their specific receptor CXCR3 is expressed by infiltrating lymphocytes in discoid LE tissues [7]. T cells expressing CXCR3 increase in peripheral blood of patients with lupus and renal involvement [8], and infiltrate in
http://dx.doi.org/10.1016/j.humimm.2014.02.005 0198-8859/Ó 2014 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.
C.H. Im et al. / Human Immunology 75 (2014) 466–469
the tubule-interstitium in renal tissue from patients with lupus nephritis, which correlates with kidney dysfunction [9]. In an animal model of lupus nephritis, immune-mediated kidney disease is promoted by CXCR3 and one of its ligands, CXCL9, but not CXCL10, via activated T cell and macrophage recruitment into the kidney [10]. Th1 and Th17 cell infiltration into the inflamed kidney and morphological and functional damage of nephritic kidney are suppressed in a CXCR3 knock-out lupus nephritis mice model (MRL/ lpr) [10,11]. CXCR3 expression in urinary CD4 T cells correlates with SLEDAI scores in patients with lupus nephritis [12]. CXCR3bearing memory B cell subset is related to autoantibody production and adverse clinical outcomes in SLE [13]. The CXCR3 gene (NC_000023.10) is located on the X chromosome (Xq13). Four single nucleotide polymorphism (SNP) sites were located within the CXCR3 gene in a previous Korean population study [14]. According to the NCBI database, the frequencies of the other CXCR3 gene polymorphism are extremely low (frequency < 0.05), except rs2280964, rs34334103, and rs111516068. The first two SNPs were evaluated in Korean patients with allergic asthma, but the last SNP (rs111516068) was not detected in the Korean population [14,15]. In this study, we investigated CXCR3 polymorphisms in patients with SLE, and their association with clinical manifestations.
2. Methods A total of 346 patients with SLE were enrolled at the Rheumatology Clinic, Seoul National University Hospital, Korea. They were diagnosed by SLE classification criteria, which were revised in 1997 by the American College of Rheumatology [16]. After informed consent, the patient’s peripheral blood was collected, and medical records were reviewed as follows: age, sex, mucocutaneous (malar rash, discoid rash, oral ulcer, and other skin manifestations), serosa (pleuritis and pericarditis), renal (proteinuria, 24 h collected urine protein amount, creatinine clearance, and renal histology), hematologic (leukopenia, lymphopenia, hemolytic anemia, and thrombocytopenia), arthritis, neuropsychiatric (psychosis and seizure), anti-nuclear antibody, and other autoantibodies (anti-double stranded DNA antibody, anti-Smith antibody, lupus anticoagulant, anti-cardiolipin antibody IgG/IgM, anti-Ro (SSA) antibody, and anti-La (SSB) antibody). This study was approved by the institutional review board of the hospital. A total of 540 healthy control DNA samples were collected from hospital workers during their annual medical checkup. Two CXCR3 polymorphism sites in intron 1, rs2280964 and rs3433413, were investigated by Taq-man analysis, as shown in previous studies [14,15]. Primer Express (Applied Biosystems, Foster City, CA, USA) was used to design both the polymerase chain reaction (PCR) primers and the MGB Taqman probe. One allelic probe was labeled with FAM dye and the other with fluorescent VIC dye. The primer and probe sequences are available on the website (http:// www.snp-genetics.com/reference/CXCR3_additional_information. doc). The PCR was run with TaqMan Universal Master mix without UNG (Applied Biosystems) at a primer concentration of 900 nM and a TaqMan MGB-probe concentration of 200 nM. The reaction was performed in a 384-well format in a total reaction volume of 5 ll using 20 ng genomic DNA. The plate was then placed in a thermal cycler (PE 9700, Applied Biosystems) and heated for 2 min at 50 °C and for 10 min at 95 °C followed by 40 cycles of 95 °C for 15 s and 60 °C for 1 min. The TaqMan assay plate was then transferred to a Prism 7900HT instrument (Applied Biosystems) where the fluorescence intensity of each well was read. Fluorescence data files from each plate were analyzed using automated software (SDS 2.1). Baseline statistics was analyzed with logistic regression models controlling for age and sex. False discovery rate adjusted P-values
467
(Pc) were calculated for the multiple comparison correction. The association with each SLE manifestation and the laboratory results were analyzed with SPSS program version 12.0 K (SPSS Inc., Chicago, IL, USA). The Haploview program version 4.2, (Broad Institute of MIT and Harvard, Cambridge, MA, USA) was used for the haplotype analysis with the two loci of the CXCR3 polymorphisms. 3. Results A total of 346 patients with SLE and 540 healthy controls were analyzed. The mean ages (± standard deviation) of patients with SLE and healthy controls were 35.6 ± 13.1 and 43.3 ± 15.6 years, respectively. The male gender ratios of the groups were 9.2% vs. 12.4% respectively. Neither polymorphism was significantly different between the SLE and normal control groups (Table 1). However, after stratification by gender, the A allele of the rs34334103 polymorphism was associated with male patients with SLE compared to that in male controls (Pc value < 0.001, odds ratio [OR] 14.279; 95% confidence interval [CI] 4.153–49.095) (Table 2). In the haplotype analysis for male gender (rs2280964-rs34334103), both polymorphisms were in linkage disequilibrium (D’ = 1.000, r2 = 0.115), and the CA haplotype (C allele from rs2280964 and A allele from rs34334103) was significantly associated with an increased risk of male lupus compared to that in the male control group (Pc value = 0.002; OR 10.833; 95% CI 2.147–54.669) (Table 2). In females, neither polymorphism was different between the SLE and control groups, and the haplotype analysis was not significant either (Table 2). Mucocutaneous, musculoskeletal, hematological, renal, and neurologic manifestations were not associated with the CXCR3 polymorphisms in patients with SLE. However, the A allele of CXCR3 SNP rs34334103 increased significantly in patients with pleuritis (Pc value = 0.013; OR 2.250; 95% CI 1.228–4.122) (Table 3). Both polymorphisms were in linkage disequilibrium (D’ = 0.913, r2 = 0.102), and haplotype CA was associated with pleuritis (Pc value = 0.009; OR 2.415; 95% CI 1.309–4.453) (Table 3). Significance disappeared in male patients with pleuritis after stratifying by gender, but not in female patients (Supplementary Table 1). Among laboratory abnormalities, anti-phospholipid antibody IgG tended to be associated with both rs2280964 and rs34334103 polymorphisms (P value = 0.038 and 0.046, respectively, but was not significant after correction of multiple comparison). Anti-SSB (La) antibodies were associated with the A allele of rs34334103 (Pc value = 0.046). The other laboratory results such as antinuclear antibody, anti-double stranded DNA antibody, anti-Smith antibody, lupus anticoagulant, anti-cardiolipin antibody IgG/M, anti-Ro (SSA) antibody, 24 h collected urine protein amount, and creatinine clearance were not associated with the polymorphisms or haplotypes. 4. Discussion CXCR3 is a receptor for chemokines CXCL4, 9, 10, and 11, which are involved in lymphocyte chemotaxis to target organs in patients with SLE. CXCL9, CXCL10, and their receptor CXCR3 are involved with target tissues such as the skin and kidney and are associated with disease activity in SLE [3–9]. Taken together, CXCR3-bearing autoreactive T lymphocytes are recruited from the circulation to the site of autoimmune attack by chemokine CXCR9 or CXCL10, released from local cells at target tissues during the initiating event of SLE, and the CXCR3-brearing memory B cell subset is engaged in the extended stage of the disease [10]. Despite the importance of CXCR3 in autoimmune diseases, CXCR3 polymorphism studies have been rarely reported. Kato et al. studied variations in the coding region of human CXCR1, 2,
468
C.H. Im et al. / Human Immunology 75 (2014) 466–469
Table 1 CXCR3 polymorphisms in patients with systemic lupus erythematosus (SLE) and healthy controls. Polymorphisms
SLE (n = 346) (%)
Controls (n = 540) (%)
Pc valuea
OR
(95% CI)
NS
1.066
(0.875–1.298)
rs2280964
C allele T allele
50.0 50.0
50.5 49.5
rs34334103
G allele A allele
88.4 11.6
90.6 9.4
NS
1.283
(0.933–1.764)
Haplotypeb
TG CG CA TA
49.6 39.5 10.6 0.4
49.4 40.8 9.8 0.1
1.000 0.933 0.935 0.400
1.008 0.950 1.095 4.614
(0.828–1.226) (0.906–1.161) (0.793–1.513) (0.479–44.451)
NS, Not significant. a Pc-values for the allele frequencies were adjusted for age and sex (correction by false delivery rate for multiple comparisons), and those of each haplotype were corrected by the permutation test. b Haplotypes consisted of the rs2280964 and rs34334103 CXCR3 polymorphisms in linkage disequilibrium (D’ = 0.962, r2 = 0.104).
Table 2 CXCR3 polymorphism analysis between patients with systemic lupus erythematosus (SLE) and healthy controls stratified by gender. Gender
Polymorphisms
Male
rs2280964 rs34334103 Haplotypeb
Female
rs2280964
rs34334103
Haplotypeb
SLE (male 32, female 314) (%)
Controls (male 67, female 473) (%)
C allele T allele G allele A allele TG CG CA TA
50.0 50.0 75.0 25.0 50.0 25.0 25.0 0.0
49.3 50.7 97.0 3.0 50.7 46.3 3.0 0.0
CC CT TT C allele T allele GG GA AA G allele A allele TG CG CA TA
70 (22.4) 172 (55.1) 70 (22.4) 50.0 50.0 252 (80.5) 58 (18.5) 3 (1.0) 89.8 10.2 49.5 40.2 9.8 0.4
129 (27.6) 216 (46.2) 123 (26.3) 49.3 50.7 380 (80.3) 88 (18.6) 5 (1.1) 89.6 10.4 49.3 40.4 10.3 0.1
Pc valuea
OR
NS
0.752
