Genetic variants of RANTES are associated with serum RANTES level and protection for type 1 diabetes

Abstract

RANTES (regulated on activation, normal T-cell expressed and secreted) is a T-helper type 1 (Th1) chemokine that promotes T-cell activation and proliferation. RANTES is genetically associated with asthma, sarcoidosis and multiple sclerosis. The concentration of RANTES is increased at inflammation sites in different autoimmune diseases. Type 1 diabetes (T1D) is a Th1-mediated disease with complex genetic predisposition. We tested RANTES as a candidate gene for association with T1D using three single-nucleotide polymorphism (SNP) variants (rs4251719, rs2306630 and rs2107538) to capture haplotype information. The minor alleles of all SNPs were transmitted less frequently to T1D offspring (transmission rates 37.3% (P=0.002), 38.7% (P=0.007) and 41.0% (P=0.01)) and were less frequently present in patients compared to controls (P=0.009, 0.03 and 0.04, respectively). A similar protective effect was observed for the haplotype carrying three minor alleles (transmission disequilibrium test (TDT): P=0.003; odds ratio (OR)=0.55; confidence interval (CI): 0.37–0.83; case/control: P=0.03; OR=0.74; CI: 0.55–0.98). Both patients and controls carrying the protective haplotype express significantly lower serum levels of RANTES compared to non-carriers. Subsequently, we tested a cohort of 310 celiac disease patients, but failed to detect association. RANTES SNPs are significantly associated with RANTES serum concentration and development of T1D. The rs4251719*A–rs2306630*A–rs2107538*A haplotype associated with low RANTES production confers protection from T1D. Our data imply that RANTES is associated with T1D both genetically and functionally, and contributes to diabetes-prone Th1 cytokine profile.

Introduction

Type 1 diabetes (T1D) is an autoimmune disease characterized by a T-cell-mediated progressive destruction of pancreatic beta cells.1 The genetic predisposition to T1D is complex, implying that multiple susceptibility genes with different effects play a role in the development of the disease. Approximately, 50% of the genetic susceptibility to T1D is explained by four established genetic risk factors, including human leukocyte antigen class II genes, the insulin gene, cytotoxic T-lymphocyte-associated antigen 4 (CTLA4) and protein tyrosine phosphatase PTPN22.2, 3, 4 These findings indicate that analyzing candidate genes is a fruitful approach in the search for T1D susceptibility genes, but that other genetic risk factors still need to be identified.

An interesting candidate gene is RANTES (‘regulated on activation, normal T-cell expressed and secreted’, CCL5). This gene has not yet been investigated in the context of T1D, although it has been shown to be associated with other autoimmune diseases, such as atopic dermatitis, asthma, sarcoidosis, rheumatoid arthritis and multiple sclerosis.5, 6, 7, 8, 9, 10, 11, 12 RANTES belongs to the family of CC chemokines, which are involved in immunoregulatory and inflammatory processes owing to their ability to recruit, activate and co-stimulate T cells and monocytes.13, 14 In addition to the trafficking effect, RANTES, like other CC chemokines, plays an important role in co-stimulation of T-cell proliferation15, 16 and activation of the T cells localized in the inflammatory lesion.17 Activation of T cells by RANTES leads to interleukin-2 (IL-2) receptor expression, cytokine release and T-cell proliferation.16 Micromolar concentrations of RANTES can induce T-cell proliferation in an antigen-independent manner that is especially important in autoimmunity, whereas the increased level of RANTES in the inflammatory sites may lead to the enhanced activation and proliferation of local T cells.17 Recent studies suggest that RANTES is also involved in the process of T-cell differentiation. Administration of RANTES and its interaction with the CCR5 receptor bias the T-helper response toward a T-helper type 1 (Th1)-associated cytokine profile.18, 19 A RANTES−/− mouse model showed decreased T-cell proliferation and impaired production of Th1 cytokines IL-2 and interferon-γ.20

RANTES is located in a cluster with several CC chemokine genes on 17q12. Three RANTES single-nucleotide polymorphisms (SNPs) in the promoter (rs2280788: −28*C/G and rs2107538: −403*G/A) and in the first intron (rs2280789: Int1.1*T/C) have been shown in different studies to modify the transcriptional activity of RANTES.7, 21, 22 However, establishing which of these SNPs has functional consequences is complicated by the high linkage disequilibrium (LD) between them.

Given the strong involvement of RANTES in the immune process, we decided to perform a genetic analysis of the RANTES gene region in T1D. To assess the functional significance of the genetic variants, we tested whether the serum level of RANTES was dependent on the underlying genotypes.

Results

Haplotype structure of the RANTES region

To comprehensively test RANTES for association with T1D, we first investigated the LD structure of the RANTES gene (www.hapmap.org).23 We observed that RANTES is located in an extended block of strong LD and that two SNPs from the block is sufficient to tag all the haplotype information (data not shown; available at http://humgen.med.uu.nl/publications/zhernakova2006_1.html).

Next, we assessed whether the haplotype structure in a Dutch population is similar to the CEHP families included in HAPMAP. In 90 healthy controls that were picked at random from the whole control group, we determined that SNPs rs2306630 and rs3817655 were mutually redundant, meaning that only one of this pair needed to be used for testing. Thus, only three SNPs – rs4251719, rs2306630 and rs2107538 – are informative for the RANTES gene region. Although the infrequent rs2280788 SNP did not differ between haplotypes, we included this SNP in our study owing to its possible functionality. Consequently, four SNPs (rs4251719, rs2306630, rs2280788 and rs2107538) were tested in further studies.

Association studies

All four SNPs were tested for association with transmission disequilibrium test (TDT) in our cohort of 218 families. We observed a significant decrease in transmission of the minor alleles of rs4251719*A (OR=0.55; 95% CI: 0.37–0.81; P=0.002); rs2306630*A (OR=0.59; 95% CI: 0.40–0.87; P=0.007) and rs2107538*A (OR=0.64; 95% CI: 0.45–0.91; P=0.01) to affected children (Table 1). The transmission of the rare rs2280788*G allele was also decreased, but failed to reach significance (Table 1). To confirm our findings, we tested the same SNPs in a case–control design using 350 T1D cases and 540 controls. We observed a significant decrease in frequency of rs4251719*A (OR=0.70; 95% CI: 0.53–0.92; P=0.009), rs2306630*A (OR=0.74; 95% CI: 0.56–0.98; P=0.03) and rs2107538*A (OR=0.79; 95% CI: 0.63–0.99; P=0.04) in T1D cases compared to controls (Table 2).

Table 1 TDT results in T1D families (n=218)
Table 2 Allele and genotype frequencies of the SNPs in the RANTES region in T1D (n=350) and control (n=540) groups

We then tested whether any particular RANTES haplotype was associated with T1D. Three frequent haplotypes were observed in both cases and controls that together comprise 97% of all haplotypes. As all three haplotypes contained the rs2280788*C allele, the rs2280788 SNP was omitted from the haplotype analysis. Transmission of the rs4251719*A–rs2306630*A–rs2107538*A haplotype (further referred to as haplotype A–A–A) to T1D offspring was decreased (OR=0.55; 95% CI: 0.37–0.83; P=0.003). The frequency of this A–A–A haplotype was also decreased in T1D cases compared to controls (OR=0.74; 95% CI: 0.55–0.98; P=0.03; Table 4). The frequency of the rs4251719*G–rs2306630*G–rs2107538*A haplotype (G–G–A) was similar in cases and controls and transmission of this haplotype was not distorted (Tables 3 and 4). The overall haplotype association was also significant (P=0.01 and P=0.02 in TDT and the case–control study, respectively; Tables 3 and 4).

Table 4 Frequency of RANTES haplotype in T1D cases (n=350) and controls (n=540)
Table 3 Transmission of RANTES haplotypes to affected offspring (n=218)

Association with celiac disease

The positive association found in T1D prompted us to investigate these RANTES polymorphism in another autoimmune disease, celiac disease (CD). We therefore genotyped RANTES SNPs rs2306630, rs2280788 and rs2107538 in a group of 310 CD patients and compared them with controls. The allele distribution of all three SNPs in celiac patients was not significantly different from that in controls (Supplementary Table 1, also available online at http://humgen.med.uu.nl/publications/zhernakova2006_1.html).

Relation to RANTES serum level

To investigate the functional relevance of the RANTES genotype, we tested for correlation between serum level and genotypes in an independent cohort of 15 recent onset T1D patients and 14 age-matched controls. The patients and controls were genotyped for the SNPs rs2280788, rs2107538 and rs2306630. Only three patients and one control were heterozygous for the rs2280788*G allele. This SNP was excluded from the analysis because of its low frequency.

Carriers of the T1D-associated alleles, rs2107538*A and rs2306630*A (A–A haplotype), had lower levels of RANTES compared to non-carriers (Figure 1). The mean level of RANTES was significantly lower in control carriers for the A–A haplotype (mean 25456.3±2841.1 pg/ml) than in non-carriers (44274.6±2617.4 pg/ml) (P=0.01). In patient carriers and non-carriers of the A–A haplotype, the RANTES serum concentrations were 27 129.8±3044.1 and 35 591.9±3369.2 pg/ml, respectively (P=0.09). When patients and controls were combined, the difference in RANTES serum level between carriers and non-carriers of the protective haplotype reached significance (P=0.003), suggesting that carriers of the protective A–A haplotype have a lower RANTES serum level than non-carriers.

Figure 1
figure1

Correlation of RANTES serum level with the presence or absence of haplotype 2. In the controls (n=14), two individuals carried haplotype 2 (white), 12 were non-carriers (black); in the patients (n=15), there were five carriers (white) and 10 non-carriers (black). P-value was calculated with Student's t-test by comparing the RANTES serum level in carriers and non-carriers of the haplotype 2.

Discussion

We demonstrate that both in families and in a case–control cohort, the minor alleles of SNPs rs4251719*A, rs2306630*A and rs2107538*A are associated with protection from T1D. We further show that these alleles are related to decreased serum protein levels. This implies that RANTES is involved in the pathogenesis of T1D, presumably owing to its key role in the process of polarization, activation and differentiation of T cells.

A number of molecular mechanisms can explain the association between the RANTES gene and T1D: (i) binding of RANTES to its receptor CCR5 leads to the activation of Janus kinases (JAK).24, 25 Remarkably, the administration of JAK3 inhibitor JANEX1 delays the onset of autoimmune diabetes in non-obese diabetic (NOD) mice.26 (ii) RANTES is one of the activation factors of Erk and p38 members of the mitogen-activated protein kinase (MAPK) family that are involved in regulating cell proliferation and differentiation.17, 25, 27, 28 Interestingly, p38 is considered to be a master switch between benign and destructive insulitis and therefore provokes the development of T1D.29 Inhibition of p38 MAPK leads to the inhibition of Th1 immunity and prevents NOD mice from developing diabetes.29 It is therefore conceivable that a decreased level of RANTES could offer protection from development of T1D via reduced activation of one of these pathways.

Genetic associations of RANTES SNPs with asthma, atopic dermatitis, sarcoidosis, multiple sclerosis and other inflammatory and autoimmune diseases have been reported.5, 6, 7, 8, 9, 10, 11, 12 Therefore, RANTES could be considered a gene generally predisposing to autoimmune disease. To test this hypothesis, we investigated RANTES SNPs rs2306630, rs2280788 and rs2107538 in a group of 310 CD patients. Although RANTES genotypes did not differ significantly between patients and controls (Supplementary Table 1), our microarray studies on intestinal biopsies indicated increased levels of RANTES in the intestinal biopsies of CD patients compared to controls (B Diosdado, personal communication). The increase in the level of RANTES correlated with the severity of the morphological changes in the intestine. Despite a lack of genetic association for RANTES in CD, the fact that the levels of RANTES are increased in patient biopsies, and in combination with the genetic association with RANTES in other autoimmune diseases, renders this gene a potentially relevant candidate biological marker for autoimmune disease.

Increased protein levels of RANTES in the primary sites of inflammation have been reported in different autoimmune diseases. RANTES levels are increased in the cerebrospinal fluid of patients suffering active attacks of multiple sclerosis.14 RANTES levels are also increased in allograft rejection,14 in patients with system lupus erythematosis,30 in the synovium of patients with rheumatoid arthritis31 and in the granulomas in Crohn's disease.32 Similarly, we here report that the RANTES alleles associated with lower serum level of the protein are associated with protection from T1D.

Investigating the functionality of RANTES SNPs is complicated by the presence of high LD in this region. The transcriptional activity of two promoter SNPs (rs2280788: −28*C/G and rs2107538: −403*G/A) and an SNP in intron 1 (rs2280789: Int1.1*T/C) has been investigated.7, 21, 22 An et al.22 showed that transcription activity was mainly regulated by the Int1.1 genotype. Int1.1*C allele strongly downregulates the transcriptional activity of RANTES, whereas the −403*G/A SNP has no effect on transcription and the −28*G allele shows a modest upregulation.22 We have investigated the SNP rs2306630, which, according to the HAPMAP data, is a perfect proxy of the Int1.1 SNP (where the Int1.1*C allele could be tagged by the rs2306630*G allele and the Int1.1*T allele corresponds to the rs2306630*A allele). We observed that the haplotype 2, which is tagged by the rs2306630*A allele, is associated with a lower protein level of RANTES. Our results therefore corroborate An et al.'s22 findings.

RANTES functions via the CC receptors, mainly CCR5, and the association of T1D with the functional variant in the CCR5 gene (CCR5Δ32) has been reported previously.33, 34 We therefore considered testing for epistasis between RANTES and CCL5 genetic variants. However, the observed frequency of the CCR5Δ32 variant in our patient cohort was too low to provide conclusive evidence for interactions between these genes.

We propose RANTES as a functional candidate gene for T1D. However, as RANTES is located in an extended block of high LD, the observed genetic association could be owing to variants in other proximal genes. The block of strong LD includes six known and predicted genes (MMP28, TAF15, FLJ32830, CCL5, LOC440427 and RAD52B). Among these, the MMP28 (the epilysin member of matrix metalloproteinase (MMP)) could serve as an alternative functional candidate for predisposition to autoimmune disease. The MMP28 gene is important for tissue remodelling and wound repair. Interestingly, the level of several MMP genes is reported to be influenced by RANTES stimulation.35, 36, 37 The CC chemokine genes located telomeric of RANTES are also in relatively strong LD with RANTES, and therefore cannot be excluded. A detailed study of this region is therefore necessary to elucidate the contribution of RANTES to the observed association.

Subjects and methods

Subjects

A total of 350 patients with juvenile onset T1D were included in the case–control study. T1D in all the children was diagnosed according to the International Society of Pediatric and Adolescent Diabetes (ISPAD) criteria. All these patients were diagnosed at age 17 years or younger (median 8.7 years, range 1–17 years); 159 were girls. Both parents were available for 218 of these patients, so that we could perform a family-based association study on them by means of a TDT.

The control group consisted of 540 independent Dutch individuals, comprising 180 healthy spouses of non-diabetic patients included in different (not immune-mediated) studies, and 360 healthy blood donor controls, as described previously.38 Written informed consent was obtained from all participants.

Serum levels of RANTES were measured in 15 T1D patients and 14 matched controls from the REMTrial project by double-sandwich enzyme-linked immunosorbent assay, as described previously.39 The detection level for RANTES was set at 14 pg/ml.

A total of 310 independent CD patients were studied. The diagnosis of CD was made according to the revised European Society of Paediatric Gastroenterology and Nutrition (ESPGAN) criteria.40 In addition, the intestinal biopsies on which the initial diagnoses were based were re-evaluated for all patients by one experienced pathologist. Only patients with a Marsh III lesion were included in our study. Of the 310 CD patients, 214 were female and 96 were male patients. The average age at diagnosis of CD patients was 34.3 years (range from 1 to 83 years).

SNP selection

The RANTES gene is located in a region of strong LD of 155.5 kb extent and it is limited by SNPs rs4796105 and rs9899870. We selected two SNPs (rs2306630 and rs3817655) that could tag all the frequent haplotypes in the block according to the HAPMAP data. We also included three additional SNPs (rs4251719, rs2280788 and rs2107538), two of which were located in the promoter region of RANTES (rs2280788 and rs2107538) and have previously been reported to influence the protein level of RANTES. One additional previously reported functional SNP – Int1.1 (rs2280789) – was typed in the HAPMAP project and, according to the HAPMAP data, could be replaced by rs2306630 owing to the strong LD and the identical allele frequency.

Genotyping

SNPs rs2306630, rs4251719, rs3817655 and rs2107538 (−403*G/A) were genotyped using Taqman assays-on-demand (Applied Biosystems, 2910 AH Nieuwerkerk a/d IJssel, The Netherlands) (assay numbers C_26924101_10, C_3143173_10, C_2957192_10 and C_15874407_10, respectively). The rs2280788 (−28*C/G) SNPs was genotyped using Taqman assay-by-design (Applied Biosystems). Assays were performed according to the manufacturer's specifications. Genotypes were analyzed using a TaqMan 7900HT (Applied Biosystems). The DNA samples were processed in 384-well plates. Each plate contained eight negative controls and 16 genotyping controls (four duplicates of four different CEPH samples).

In the controls, the frequency of all the SNPs was in Hardy–Weinberg (HW) proportions, except the rs2107538 (P=0.02), which was most likely owing to a low number of individuals homozygous for the minor allele. To exclude possible genotyping mistakes, we typed this SNP in the additional cohort of 360 healthy individuals. The allelic and genotypes distribution were in HW proportions in the total control group (P=0.35).

Data analysis

The haplotype structure and LD in the region was investigated using the HAPMAP data (www.hapmap.org)41 with the Haploview application.42 Allele and genotype distribution in cases and controls were compared using the COCAPHASED module of the UNPHASED statistical package.43 Association in the T1D families was tested using the TDT phase module of the same package.43 Haplotypes association were estimated using the UNPHASED package.43 HW equilibrium was tested by comparing the expected and observed genotypes in 2 × 3 χ2 table. ORs were calculated and the CIs were approximated using Woolf's method with Haldane's correction.44 The Student's t-test and analysis of variance (ANOVA) were used to compare the mean levels of serum RANTES level among the RANTES genotypes.

References

  1. 1

    Atkinson MA, Eisenbarth GS . Type 1 diabetes: new perspectives on disease pathogenesis and treatment. Lancet 2001; 358: 221–229.

  2. 2

    Ueda H, Howson JM, Esposito L, Heward J, Snook H, Chamberlain G et al. Association of the T-cell regulatory gene CTLA4 with susceptibility to autoimmune disease. Nature 2003; 423: 506–511.

  3. 3

    Bottini N, Musumeci L, Alonso A, Rahmouni S, Nika K, Rostamkhani M et al. A functional variant of lymphoid tyrosine phosphatase is associated with type I diabetes. Nat Genet 2004; 36: 337–338.

  4. 4

    Kelly MA, Rayner ML, Mijovic CH, Barnett AH . Molecular aspects of type 1 diabetes. Mol Pathol 2003; 56: 1–10.

  5. 5

    Hizawa N, Yamaguchi E, Konno S, Tanino Y, Jinushi E, Nishimura M . A functional polymorphism in the RANTES gene promoter is associated with the development of late-onset asthma. Am J Respir Crit Care Med 2002; 166: 686–690.

  6. 6

    Al-Abdulhadi SA, Helms PJ, Main M, Smith O, Christie G . Preferential transmission and association of the −403 G → A promoter RANTES polymorphism with atopic asthma. Genes Immun 2005; 6: 24–30.

  7. 7

    Nickel RG, Casolaro V, Wahn U, Beyer K, Barnes KC, Plunkett BS et al. Atopic dermatitis is associated with a functional mutation in the promoter of the C–C chemokine RANTES. J Immunol 2000; 164: 1612–1616.

  8. 8

    Takada T, Suzuki E, Ishida T, Moriyama H, Ooi H, Hasegawa T et al. Polymorphism in RANTES chemokine promoter affects extent of sarcoidosis in a Japanese population. Tissue Antigens 2001; 58: 293–298.

  9. 9

    Leung TF, Tang NL, Lam CW, Li AM, Fung SL, Chan IH et al. RANTES G−401A polymorphism is associated with allergen sensitization and FEV1 in Chinese children. Respir Med 2005; 99: 216–219.

  10. 10

    Kozma GT, Falus A, Bojszko A, Krikovszky D, Szabo T, Nagy A et al. Lack of association between atopic eczema/dermatitis syndrome and polymorphisms in the promoter region of RANTES and regulatory region of MCP-1. Allergy 2002; 57: 160–163.

  11. 11

    Makki RF, al Sharif F, Gonzalez-Gay MA, Garcia-Porrua C, Ollier WE, Hajeer AH . RANTES gene polymorphism in polymyalgia rheumatica, giant cell arteritis and rheumatoid arthritis. Clin Exp Rheumatol 2000; 18: 391–393.

  12. 12

    Gade-Andavolu R, Comings DE, MacMurray J, Vuthoori RK, Tourtellotte WW, Nagra RM et al. RANTES: a genetic risk marker for multiple sclerosis. Mult Scler 2004; 10: 536–539.

  13. 13

    Luster AD . Chemokines – chemotactic cytokines that mediate inflammation. N Engl J Med 1998; 338: 436–445.

  14. 14

    Gerard C, Rollins BJ . Chemokines and disease. Nat Immunol 2001; 2: 108–115.

  15. 15

    Taub DD, Turcovski-Corrales SM, Key ML, Longo DL, Murphy WJ . Chemokines and T lymphocyte activation: I. Beta chemokines costimulate human T lymphocyte activation in vitro. J Immunol 1996; 156: 2095–2103.

  16. 16

    Bacon KB, Premack BA, Gardner P, Schall TJ . Activation of dual T cell signaling pathways by the chemokine RANTES. Science 1995; 269: 1727–1730.

  17. 17

    Wong MM, Fish EN . Chemokines: attractive mediators of the immune response. Semin Immunol 2003; 15: 5–14.

  18. 18

    Zou W, Borvak J, Marches F, Wei S, Galanaud P, Emilie D et al. Macrophage-derived dendritic cells have strong Th1-polarizing potential mediated by beta-chemokines rather than IL-12. J Immunol 2000; 165: 4388–4396.

  19. 19

    Frauenschuh A, DeVico AL, Lim SP, Gallo RC, Garzino-Demo A . Differential polarization of immune responses by co-administration of antigens with chemokines. Vaccine 2004; 23: 546–554.

  20. 20

    Makino Y, Cook DN, Smithies O, Hwang OY, Neilson EG, Turka LA et al. Impaired T cell function in RANTES-deficient mice. Clin Immunol 2002; 102: 302–309.

  21. 21

    Liu H, Chao D, Nakayama EE, Taguchi H, Goto M, Xin X et al. Polymorphism in RANTES chemokine promoter affects HIV-1 disease progression. Proc Natl Acad Sci USA 1999; 96: 4581–4585.

  22. 22

    An P, Nelson GW, Wang L, Donfield S, Goedert JJ, Phair J et al. Modulating influence on HIV/AIDS by interacting RANTES gene variants. Proc Natl Acad Sci USA 2002; 99: 10002–10007.

  23. 23

    The International HapMap Consortium. The International HapMap Project. Nature 2003; 426: 789–796.

  24. 24

    Rodriguez-Frade JM, Vila-Coro AJ, Martin A, Nieto M, Sanchez-Madrid F, Proudfoot AE et al. Similarities and differences in RANTES- and (AOP)-RANTES-triggered signals: implications for chemotaxis. J Cell Biol 1999; 144: 755–765.

  25. 25

    Wong M, Uddin S, Majchrzak B, Huynh T, Proudfoot AE, Platanias LC et al. Rantes activates Jak2 and Jak3 to regulate engagement of multiple signaling pathways in T cells. J Biol Chem 2001; 276: 11427–11431.

  26. 26

    Cetkovic-Cvrlje M, Dragt AL, Vassilev A, Liu XP, Uckun FM . Targeting JAK3 with JANEX-1 for prevention of autoimmune type 1 diabetes in NOD mice. Clin Immunol 2003; 106: 213–225.

  27. 27

    Kondoh K, Torii S, Nishida E . Control of MAP kinase signaling to the nucleus. Chromosoma 2005; 114 (2): 86–91.

  28. 28

    Brill A, Hershkoviz R, Vaday GG, Chowers Y, Lider O . Augmentation of RANTES-induced extracellular signal-regulated kinase mediated signaling and T cell adhesion by elastase-treated fibronectin. J Immunol 2001; 166: 7121–7127.

  29. 29

    Ando H, Kurita S, Takamura T . The specific p38 mitogen-activated protein kinase pathway inhibitor FR167653 keeps insulitis benign in nonobese diabetic mice. Life Sci 2004; 74: 1817–1827.

  30. 30

    Eriksson C, Eneslatt K, Ivanoff J, Rantapaa-Dahlqvist S, Sundqvist KG . Abnormal expression of chemokine receptors on T-cells from patients with systemic lupus erythematosus. Lupus 2003; 12: 766–774.

  31. 31

    Pierer M, Rethage J, Seibl R, Lauener R, Brentano F, Wagner U et al. Chemokine secretion of rheumatoid arthritis synovial fibroblasts stimulated by Toll-like receptor 2 ligands. J Immunol 2004; 172: 1256–1265.

  32. 32

    Oki M, Ohtani H, Kinouchi Y, Sato E, Nakamura S, Matsumoto T et al. Accumulation of CCR5+ T cells around RANTES+ granulomas in Crohn's disease: a pivotal site of Th1-shifted immune response? Lab Invest 2005; 85: 137–145.

  33. 33

    Yang B, Houlberg K, Millward A, Demaine A . Polymorphisms of chemokine and chemokine receptor genes in Type 1 diabetes mellitus and its complications. Cytokine 2004; 26: 114–121.

  34. 34

    Buhler MM, Craig M, Donaghue KC, Badhwar P, Willis J, Manolios N et al. CCR5 genotyping in an Australian and New Zealand type 1 diabetes cohort. Autoimmunity 2002; 35: 457–461.

  35. 35

    Masuko-Hongo K, Sato T, Nishioka K . Chemokines differentially induce matrix metalloproteinase-3 and prostaglandin E2 in human articular chondrocytes. Clin Exp Rheumatol 2005; 23: 57–62.

  36. 36

    Locati M, Deuschle U, Massardi ML, Martinez FO, Sironi M, Sozzani S et al. Analysis of the gene expression profile activated by the CC chemokine ligand 5/RANTES and by lipopolysaccharide in human monocytes. J Immunol 2002; 168: 3557–3562.

  37. 37

    Garcia-Vicuna R, Gomez-Gaviro MV, Dominguez-Luis MJ, Pec MK, Gonzalez-Alvaro I, Alvaro-Gracia JM et al. CC and CXC chemokine receptors mediate migration, proliferation, and matrix metalloproteinase production by fibroblast-like synoviocytes from rheumatoid arthritis patients. Arthritis Rheum 2004; 50: 3866–3877.

  38. 38

    Schipper RF, Koeleman BP, Bruining GJ, Schreuder GM, Verduijn W, De Vries RR et al. HLA class II associations with type 1 diabetes mellitus: a multivariate approach. Tissue Antigens 2001; 57: 144–150.

  39. 39

    Hanifi-Moghaddam P, Schloot NC, Kappler S, Seissler J, Kolb H . An association of autoantibody status and serum cytokine levels in type 1 diabetes. Diabetes 2003; 52: 1137–1142.

  40. 40

    Revised Criteria for Diagnosis of Coeliac Disease. Report of working group of European Society of Paediatric Gastroenterology and Nutrition. Arch Dis Child 1990; 65: 909–911.

  41. 41

    Altshuler D, Brooks LD, Chakravarti A, Collins FS, Daly MJ, Donnelly P . A haplotype map of the human genome. Nature 2005; 437: 1299–1320.

  42. 42

    Barrett JC, Fry B, Maller J, Daly MJ . Haploview: analysis and visualization of LD and haplotype maps. Bioinformatics 2005; 21: 263–265.

  43. 43

    Dudbridge F . Pedigree disequilibrium tests for multilocus haplotypes. Genet Epidemiol 2003; 25: 115–121.

  44. 44

    Haldane JB . The estimation and significance of the logarithm of a ratio of frequencies. Ann Hum Genet 1956; 20: 309–311.

Download references

Acknowledgements

We are grateful to all the patients and their families and physicians for participating in the study. We thank Jackie Senior for improving the manuscript. The study was supported by grants from the Dutch Diabetes Research Foundation (97.137), The Dutch Digestive Disease Foundation (WS 03-06), The Netherlands Organization for Health Research and Development (ZonMW 912-02-028), The Juvenile Diabetes Research Foundation International (JDRF) (2001.10.004) and the Celiac Disease Consortium, an Innovative Cluster approved by the Netherlands Genomics Initiative and partially funded by the Dutch Government (BSIK03009).

Author information

Correspondence to B P C Koeleman.

Additional information

Supplementary Information accompanies the paper on Genes and Immunity website (http://www.nature.com/gene)

Supplementary information

Supplementary Table 1 (DOC 23 kb)

Rights and permissions

Reprints and Permissions

About this article

Keywords

  • RANTES
  • CCL5
  • autoimmune diseases
  • type 1 diabetes
  • celiac disease
  • Th1 cytokine profile

Further reading