FokI Polymorphism, Vitamin D Receptor, and Interleukin-1 Receptor Haplotypes Are Associated with Type 1 Diabetes in the Dalmatian Population
2005; Elsevier BV; Volume: 7; Issue: 5 Linguagem: Inglês
10.1016/s1525-1578(10)60593-4
ISSN1943-7811
AutoresTatijana Zemunik, Veselin Škrabić, Vesna Boraska Perica, D Diklić, Ivana Marinovic‐Terzić, Vesna Čapkun, Marijana Peruzović, Janoš Terzić,
Tópico(s)Digestive system and related health
ResumoVitamin D and interleukin (IL)-1 have been suggested to function in the pathogenesis of type 1 diabetes mellitus (T1DM). Therefore, we examined the influence of gene polymorphisms in vitamin D receptor (VDR) and interleukin-1 receptor type I (IL-1-R1) on susceptibility to T1DM in the Dalmatian population of South Croatia. We genotyped 134 children with T1DM and 132 controls; for FokI polymorphism studies, we extended the control group to an additional 102 patients. The VDR gene polymorphism FokI displayed unequal distribution (P = 0.0049) between T1DM and control groups, with the ff genotype occurring more frequently in T1DM individuals whereas the VDR gene polymorphism Tru9I did not differ in frequency between studied groups. All tested polymorphisms of the IL-1-R1 gene [PstI, HinfI, and AluI (promoter region) and PstI-e (exon 1B region)] displayed no differences between cases and controls. Haplotype analysis of the VDR gene (FokI, BsmI, ApaI, TaqI, Tru9I) and of the IL-1-R1 gene (PstI, HinfI, AluI, PstI-e) found haplotypes VDR FbATu (P = 0.0388) and IL-1-R1 phap' (P = 0.0419) to be more frequent in T1DM patients whereas the BatU haplotype occurred more often in controls (P = 0.0064). Our findings indicate that the VDR FokI polymorphism and several VDR and IL-1-R1 haplotypes are associated with susceptibility to T1DM in the Dalmatian population. Vitamin D and interleukin (IL)-1 have been suggested to function in the pathogenesis of type 1 diabetes mellitus (T1DM). Therefore, we examined the influence of gene polymorphisms in vitamin D receptor (VDR) and interleukin-1 receptor type I (IL-1-R1) on susceptibility to T1DM in the Dalmatian population of South Croatia. We genotyped 134 children with T1DM and 132 controls; for FokI polymorphism studies, we extended the control group to an additional 102 patients. The VDR gene polymorphism FokI displayed unequal distribution (P = 0.0049) between T1DM and control groups, with the ff genotype occurring more frequently in T1DM individuals whereas the VDR gene polymorphism Tru9I did not differ in frequency between studied groups. All tested polymorphisms of the IL-1-R1 gene [PstI, HinfI, and AluI (promoter region) and PstI-e (exon 1B region)] displayed no differences between cases and controls. Haplotype analysis of the VDR gene (FokI, BsmI, ApaI, TaqI, Tru9I) and of the IL-1-R1 gene (PstI, HinfI, AluI, PstI-e) found haplotypes VDR FbATu (P = 0.0388) and IL-1-R1 phap' (P = 0.0419) to be more frequent in T1DM patients whereas the BatU haplotype occurred more often in controls (P = 0.0064). Our findings indicate that the VDR FokI polymorphism and several VDR and IL-1-R1 haplotypes are associated with susceptibility to T1DM in the Dalmatian population. Type 1, or insulin-dependent, diabetes mellitus (T1DM) presents with ketosis-prone hyperglycemia associated with an almost complete loss of insulin-producing pancreatic β cells. T1DM is a multifactorial autoimmune disease for which susceptibility is determined by both environmental and genetic factors.1Atkinson MA Maclaren NK Mechanisms of disease: the pathogenesis of insulin-dependent diabetes mellitus.N Engl J Med. 1994; 331: 1428-1436Crossref PubMed Scopus (967) Google Scholar, 2Bach JF Insulin-dependent diabetes mellitus as an autoimmune disease.Endocrinol Rev. 1994; 15: 516-542Crossref PubMed Scopus (770) Google Scholar, 3Cordell HJ Todd JA Multifactorial inheritance in type 1 diabetes.Trends Genet. 1995; 11: 449-503Abstract Full Text PDF PubMed Scopus (86) Google Scholar, 4Todd JA Genetic control of autoimmunity in type 1 diabetes.Immunol Today. 1990; 11: 122-129Abstract Full Text PDF PubMed Scopus (163) Google Scholar There are differences in the prevalence of T1DM in different populations, with disease being more common in Europeans and less prevalent in Asians and Africans.5Karvonen M Tuomilehto J Libman I LaPorte R For the World Health Organization DIAMOND Project group. A review of the recent epidemiological data on the worldwide incidence of type 1 (insulin-dependent) diabetes mellitus.Diabetologia. 1993; 36: 883-892Crossref PubMed Scopus (406) Google Scholar The main genetic contribution to T1DM susceptibility resides in the major histocompatibility complex.6Cox NJ Wapelhorst B Morrison VA Johnson L Punchuk L Spielman RS Todd JA Concannon P Seven regions of the genome show evidence of linkage to type 1 diabetes in a consensus analysis of 767 multiplex families.Am J Hum Genet. 2001; 69: 820-830Abstract Full Text Full Text PDF PubMed Scopus (243) Google Scholar, 7Undlien DE Lie BA Thorsby E HLA complex genes in type 1 diabetes and other autoimmune disease. 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43: 1093-1098Crossref PubMed Scopus (310) Google Scholar The VDR gene is located on chromosome 12q (12-12q14) and is highly polymorphic.15Pani MA Knapp M Donner H Braun J Baur MP Usadel KH Badenhoop K Vitamin D receptor allele combinations influence genetic susceptibility to type I diabetes in Germans.Diabetes. 2000; 49: 504-507Crossref PubMed Scopus (224) Google Scholar There are six VDR polymorphisms that have been mainly studied: the FokI restriction fragment length polymorphism (RFLP) in exon 2; BsmI, Tru9I, and ApaI RFLPs located between exons 8 and 9; the TaqI restriction site in exon 9; and the poly-A polymorphism downstream of the 3′ untranslated region.16Ban Y Taniyama M Yanagawa T Yamada S Maruyama T Kasuga A Ban Y Vitamin D receptor initiation codon polymorphism influences genetic susceptibility to type 1 diabetes mellitus in the Japanese population.BMC Med Genet. 2001; 2: 7Crossref PubMed Scopus (84) Google Scholar However, the four polymorphisms located at the 3′ end of the VDR gene [BsmI (A to G), Tru9I (G to A), ApaI (G to T), and TaqI (T to C)] have unknown functional effects.17Uitterlinden AG Fang Y Van Meurs JB Pols HA Van Leeuwen JP Genetics and biology of vitamin D receptor polymorphisms.Gene. 2004; 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Promoter IL-1-R1 gene polymorphisms HinfI (G to A), PstI (G to A), and AluI (T to C) are not known to affect transcriptional activity; thus, there is no evidence to support any functional significance of the identified polymorphisms.35Bergholdt R Larsen ZM Andersen NA Johannesen J Kristiansen OP Mandrup-Poulsen T Nerup J Pociot F The Danish Study Group of Diabetes in Childhood Danish IDDM Epidemiology and Genetics Group Characterization of new polymorphisms in the 5′UTR of the human interleukin-1 type 1 (IL1R1) gene: linkage to type 1 diabetes and correlation to IL-1RI plasma level.Genes Immun. 2000; 1: 495-500Crossref PubMed Scopus (37) Google Scholar The PstI-e polymorphism in exon 1B has been associated with T1DM in a British data set but not in the high-risk Finnish or low-risk South Indian population.10Metcalfe KA Hitman GA Pociot F Bergholdt R Toumilehto-Wolf E Tuomileht Viswanathan M Ramachandran A Nerup J An association between type 1 diabetes and the interleukin-receptor type 1 gene.Hum Immunol. 1996; 51: 41-48Crossref PubMed Scopus (32) Google Scholar In the high-risk population of Finland, this polymorphism only associated with T1DM in patients lacking high-risk HLA-DR3 and DR4 haplotypes.10Metcalfe KA Hitman GA Pociot F Bergholdt R Toumilehto-Wolf E Tuomileht Viswanathan M Ramachandran A Nerup J An association between type 1 diabetes and the interleukin-receptor type 1 gene.Hum Immunol. 1996; 51: 41-48Crossref PubMed Scopus (32) Google Scholar The IL-1-R1 PstI-e polymorphism, which is situated in exon 1B of the gene, is caused by a C to T transition that does not affect any of the recognition sites for known transcriptional regulatory factor. Also, the different allelic variants do not appear to affect the predicted mRNA secondary structure.10Metcalfe KA Hitman GA Pociot F Bergholdt R Toumilehto-Wolf E Tuomileht Viswanathan M Ramachandran A Nerup J An association between type 1 diabetes and the interleukin-receptor type 1 gene.Hum Immunol. 1996; 51: 41-48Crossref PubMed Scopus (32) Google Scholar As we have previously studied BsmI, ApaI, and TaqI VDR polymorphisms and their influence on T1DM susceptibility in the Dalmatian population, we extended our studies to the genotyping of the FokI and Tru9I polymorphisms in the same study population. We also analyzed PstI, HinfI, and AluI polymorphisms in the promoter region of IL-1-R1 gene and the PstI-e polymorphism in the exon 1B region of the same gene. The aim of the present study was also to evaluate the strength of the linkage disequilibrium (LD), ie, allelic association between investigated polymorphisms, and to correlate LD with the disease. We also performed haplotype analysis of the VDR gene (by inclusion of the results for BsmI, ApaI, TaqI) and of the IL-1-R1 gene and examined their influence on susceptibility to T1DM. From the Dalmatian region of South Croatia, 134 unrelated patients (72 boys and 62 girls) were recruited for the study. T1DM was diagnosed according to the World Health Organization criteria.12Sandler S Buschard K Bendtzen K Effects of 1,25-dihydroxyvitamin D3, and the analogues MC903 and KH1060 on inteleukin-1 beta-cell function in vitro.Immunol Lett. 1994; 41: 73-77Crossref PubMed Scopus (36) Google Scholar The mean age at onset of T1DM was 8.6 ± 4.3 years (mean ± SD). Informed consent from patients and their parents was obtained before blood sampling. The control group, 132 unrelated, consecutive normal patients (62 boys and 70 girls), were recruited from individuals who visited the hospital for general health check-ups (mean age, 8.2 ± 4.9 years). For the analysis of the FokI polymorphism, we extended the control group with 102 healthy blood donors from the Dalmatian region. Genomic DNA was extracted from peripheral blood leukocytes using the Perfect gDNA kit (Eppendorf, Hamburg, Germany). Genotypes for four gene polymorphisms in IL-1-R1 gene (PstI, HinfI, AluI, and PstI-e) and two gene polymorphisms in VDR gene (FokI and Tru9I) were identified by polymerase chain reaction followed by RFLP analysis, according to previous reports.16Ban Y Taniyama M Yanagawa T Yamada S Maruyama T Kasuga A Ban Y Vitamin D receptor initiation codon polymorphism influences genetic susceptibility to type 1 diabetes mellitus in the Japanese population.BMC Med Genet. 2001; 2: 7Crossref PubMed Scopus (84) Google Scholar, 35Bergholdt R Larsen ZM Andersen NA Johannesen J Kristiansen OP Mandrup-Poulsen T Nerup J Pociot F The Danish Study Group of Diabetes in Childhood Danish IDDM Epidemiology and Genetics Group Characterization of new polymorphisms in the 5′UTR of the human interleukin-1 type 1 (IL1R1) gene: linkage to type 1 diabetes and correlation to IL-1RI plasma level.Genes Immun. 2000; 1: 495-500Crossref PubMed Scopus (37) Google Scholar Genotypes were designated with a capital letter for the absence of a restriction site and with a lowercase letter for its presence. Therefore, the genotypes are as follow: FokI = FF, ff (homozygous for the absence or presence of the cut site, respectively), and Ff (heterozygous); Tru9I = UU, uu, Uu; PstI = PP, pp, Pp; HinfI = HH, hh, Hh; AluI = AA, aa, Aa; PstI-e = P'P', p'p', P'p'. The digested fragments were separated in 3% agarose gels and 8% acrylamide gels and were visualized by ethidium bromide staining. The EH+ program was used to estimate haplotype frequencies and the disequilibrium measure for VDR and IL-1-R1 gene polymorphisms.36Zhao JH Curtis D Sham PC Model-free analysis and permutation tests for allelic associations.Hum Hered. 2000; 50: 133-139Crossref PubMed Scopus (382) Google Scholar Conclusions about association of certain haplotypes with susceptibility to T1DM have been derived from the χ2 test. A comparison between groups was made on the basis of χ2 test and power calculation. We used the Statistica 6.0 (StatSoft, Inc., Tulsa, OK) software package for statistical analysis. A P value of less than 0.05 was considered statistically significant. The frequency of the VDR and IL-1-R1 gene polymorphisms in the T1DM patients and controls is shown in Table 1. The distribution of genotype frequencies of the FokI VDR polymorphism among T1DM patients and controls differed significantly (χ2 = 10.646, P = 0.0049) with the ff genotype occurring more frequently in the T1DM patients. Another VDR gene polymorphism, Tru9I, did not differ in frequency between both studied groups. There were no differences in the genotype frequencies of the PstI, HinfI, and AluI polymorphisms of promoter region or the PstI-e polymorphism of exon 1B region of IL-1-R1 gene in T1DM patients and controls (Table 1). As genotype frequencies for the AluI polymorphism were at the limit of statistical significance, P = 0.07, we performed power calculation and found no association between this polymorphism and T1DM (power = 0.606).Table 1Vitamin D Receptor and Interleukin 1 Receptor Type 1 Genotype Frequency in Dalmatian Population among T1DM Patients and ControlsRestriction site and genotypeT1DM, no. (%) n = 134Controls, no. (%) n = 232χ2P value (df = 2)VDR*VDR–vitamin D receptor FokIff29 (22)23 (10)FF42 (31)73 (31)10.640.0049Ff63 (47)136 (59) Tru9Iuu3 (2)4 (3)UU105 (78)90 (68)3.530.17Uu26 (20)38 (29)IL-1-R1†IL-1-R1–interleukin-1 type I receptor Promoter region PstIpp73 (54)72 (55)PP9 (7)7 (5)0.250.88Pp52 (39)53 (40) HinfIhh75 (56)76 (57)HH11 (8)9 (7)0.20.9Hh48 (36)47 (36) AluIaa28 (21)34 (26)AA20 (15)31 (23)5.290.07Aa86 (64)67 (51) Exon 1B region PstI-ep'p'51 (38)54 (41)P'P'13 (10)16 (12)0.870.65P'p'70 (52)62 (47)* VDR–vitamin D receptor† IL-1-R1–interleukin-1 type I receptor Open table in a new tab Recently, we analyzed several other VDR gene polymorphisms (ApaI, BsmI, and TaqI) among the same study participants.24Škrabić V Zemunik T Šitum M Terzić J Vitamin D receptor polymorphism and susceptibility to type 1 diabetes in the Dalmatian population.Diabetes Res Clin Pract. 2003; 59: 31-35Abstract Full Text Full Text PDF PubMed Scopus (66) Google Scholar Thus, we were able to combine results from that study with our new results, enabling estimation of haplotype frequencies present among tested individuals. BsmI, ApaI, TaqI, and Tru9I polymorphisms were found to possess a strong LD (P < 0.0001) whereas no significant LD between FokI and any other polymorphism was detectable. The most frequent haplotypes in our population were BAtU (35%) and baTU (29%). The BatU haplotype was observed in 10 controls but in no T1DM individuals (P = 0.0064). When integrated with the FokI polymorphism, several other allele combinations were also found to be more common in the control group, such as FbATu (P = 0.00163) and fBATU (P = 0.0374). On the other hand, the FbATU (P = 0.0388) haplotype was found to be a risk factor for susceptibility to T1DM. The IL-1-R1 gene polymorphisms PstI, HinfI, AluI, and PstI-e were also found to possess a strong LD (P < 0.0001). The most frequent haplotype in our population was PHap' (39%) followed by PHAP' (21%) and phAp' (20%). The allelic combination phap' was found to be linked with T1DM (P = 0.0419). Genotype frequencies for each RFLP fit the Hardy-Weinberg equilibrium except for the FokI polymorphism in which we found a slight deviation among controls. For this reason, we extended the control group to an additional 102 new patients, but the deviation remained. We have found an association between the VDR FokI polymorphism and T1DM in our study population (χ2 = 10.646, P = 0.0049). The genotype ff was twice as common in T1DM patients as in controls. Because the f allele corresponds to a less active VDR protein,17Uitterlinden AG Fang Y Van Meurs JB Pols HA Van Leeuwen JP Genetics and biology of vitamin D receptor polymorphisms.Gene. 2004; 338: 143-156Crossref PubMed Scopus (1228) Google Scholar, 37Gross C Eccleshall TR Malloy PJ Villa ML Marcus R Feldman D The presence of a polymorphism at the translation initiation site of the vitamin D receptor gene is associated with low bone mineral density in postmenopausal Mexican-American women.J Bone Miner Res. 1996; 11: 1850-1855Crossref PubMed Scopus (423) Google Scholar the ff genotype might contribute to the development of the T1DM either by causing weaker insulin production or by affecting vitamin D immunosuppressive properties.38Haussler MR Whitfield GK Haussler CA Hsieh JC Thompson PD Selznick SH Dominguez CE Jurutka PW The nuclear vitamin D receptor: biological and molecular regulatory properties revealed.J Bone Miner Res. 1998; 13: 325-349Crossref PubMed Scopus (1259) Google Scholar The extended control group for the VDR FokI polymorphism did not fit into Hardy-Weinberg equilibrium, and we explain this by hidden subpopulation stratification. Several other studies found the FokI polymorphism to contribute to genetic heterogeneity of T1DM. In agreement with our results, the less active VDR protein occurs more frequently in T1DM patients and also associates with GAD65-Ab-positive T1DM in the Japanese population,16Ban Y Taniyama M Yanagawa T Yamada S Maruyama T Kasuga A Ban Y Vitamin D receptor initiation codon polymorphism influences genetic susceptibility to type 1 diabetes mellitus in the Japanese population.BMC Med Genet. 2001; 2: 7Crossref PubMed Scopus (84) Google Scholar although it was not informative in the German population.15Pani MA Knapp M Donner H Braun J Baur MP Usadel KH Badenhoop K Vitamin D receptor allele combinations influence genetic susceptibility to type I diabetes in Germans.Diabetes. 2000; 49: 504-507Crossref PubMed Scopus (224) Google Scholar However, a similar study was performed in two Spanish populations with different genetic backgrounds: the Mediterranean population (Barcelona) exhibited no difference regarding FokI polymorphism whereas the Navarra population exhibited decreased ff genotype (the less active form) in T1DM patients.39Marti G Audi L Esteban C Oyarzabal M Chueca M Gussinye M Yeste D Fernandez-Cancio M Andaluz P Carrascosa A Association of vitamin D receptor gene polymorphism with type 1 diabetes mellitus in two Spanish populations.Med Clin (Barc). 2004; 123: 286-290Crossref PubMed Google Scholar We were also able to compute the strength of LD between studied polymorphisms and to estimate the haplotype frequencies. The observed LD between BsmI, ApaI, TaqI, and FokI are in accordance with previously reported findings for Caucasian populations.27Colotta F Re F Muzio M Bertini R Polentarutti N Sironi M Giri JG Dower SK Sims JE Mantovani A Interleukin 1 type II receptor: a decoy target for IL-1 that is regulated by IL-4.Science. 1993; 261: 472-475Crossref PubMed Scopus (896) Google Scholar, 40Uitterlinden AG Pols HA Burger H Huang Q Van Daele PL Van Duijn CM Hofman A Birkenhager JC Van Leeuwen JP A large-scale population-based study of the association of vitamin D receptor gene polymorphisms with bone mineral density.J Bone Miner Res. 1996; 11: 1241-1248Crossref PubMed Scopus (229) Google Scholar, 41Peltekova VD Cole DE Pavlova A Rubin LA Improved method for direct haplotyping at the vitamin D receptor gene locus.Clin Biochem. 1998; 31: 191-194Crossref PubMed Scopus (7) Google Scholar, 42Ingles SA Haile RW Henderson BE Kolonel LN Nakaichi G Shi CY Yu MC Ross RK Coetzee GA Strength of linkage disequilibrium between two vitamin D receptor markers in five ethnic groups: implications for association studies.Cancer Epidemiol Biomarkers Prev. 1997; 6: 93-98PubMed Google Scholar, 43Nejentsev S Godfrey L Snook H Rance H Nutland S Walker NM Lam AC Guja C Ionescu-Tirgoviste C Undlien DE Ronningen KS Tuomilehto-Wolf E Tuomilehto J Newport MJ Clayton DG Todd JA Comparative high-resolution analysis of linkage disequilibrium and tag single nucleotide polymorphisms between populations in the vitamin D receptor gene.Hum Mol Genet. 2004; 13: 1633-1639Crossref PubMed Scopus (167) Google Scholar The most frequent haplotypes were BAtU (35%) and baTU (29%), which is also a confirmation of the previous findings for Caucasians.24Škrabić V Zemunik T Šitum M Terzić J Vitamin D receptor polymorphism and susceptibility to type 1 diabetes in the Dalmatian population.Diabetes Res Clin Pract. 2003; 59: 31-35Abstract Full Text Full Text PDF PubMed Scopus (66) Google Scholar, 40Uitterlinden AG Pols HA Burger H Huang Q Van Daele PL Van Duijn CM Hofman A Birkenhager JC Van Leeuwen JP A large-scale population-based study of the association of vitamin D receptor gene polymorphisms with bone mineral density.J Bone Miner Res. 1996; 11: 1241-1248Crossref PubMed Scopus (229) Google Scholar BsmI, ApaI, TaqI, and Tru9I RFLPs are located near the 3′ end of the VDR gene, which contains many polymorphisms, and the LD extends into the 3′ regulatory region containing the UTR. The 3′ UTR of genes is known to be involved in regulation of expression, especially through regulation of mRNA stability and degradation.17Uitterlinden AG Fang Y Van Meurs JB Pols HA Van Leeuwen JP Genetics and biology of vitamin D receptor polymorphisms.Gene. 2004; 338: 143-156Crossref PubMed Scopus (1228) Google Scholar Although the RFLPs we studied were anonymous, ie, have no known functional effect, the associations we found might be explained by their strong LD with some truly functional polymorphism in the 3′ UTR region. The VDR protein can exist in less active (f allele) and more active (F allele) variants. Individuals with the identical heterozygous genotypes for FokI, BsmI, ApaI, and TaqI might produce different levels of VDR protein. This occurs as a result of the particular haplotype combinations, which produce more or less active VDR protein depending on their linkage with the 3′ UTR.17Uitterlinden AG Fang Y Van Meurs JB Pols HA Van Leeuwen JP Genetics and biology of vitamin D receptor polymorphisms.Gene. 2004; 338: 143-156Crossref PubMed Scopus (1228) Google Scholar, 44Whitfield GK Remus LS Jurutka PW Zitzer H Oza AK Dang HT Haussler CA Galligan MA Thatcher ML Encinas Dominguez C Haussler MR Functionally relevant polymorphisms in the human nuclear vitamin D receptor gene.Mol Cell Endocrinol. 2001; 177: 145-159Crossref PubMed Scopus (370) Google Scholar This could not have been predicted by analyzing single SNPs and/or by looking at genotypes of individual SNPs but is only evident on analysis of gene-wide haplotypes.17Uitterlinden AG Fang Y Van Meurs JB Pols HA Van Leeuwen JP Genetics and biology of vitamin D receptor polymorphisms.Gene. 2004; 338: 143-156Crossref PubMed Scopus (1228) Google Scholar In light of these descriptions, we can offer an explanation for why two of our possibly protective haplotypes, FbATu and fBATU, carry different FokI alleles. The fBATU haplotype carries the f allele, which is a susceptibility allele in our population; however, this haplotype is found more commonly in the control group. One possible explanation is that the BATU haplotype is linked with some truly functional polymorphism in the 3′ UTR region that determines higher transcription and/or stability of mRNA and diminishes the effect of susceptibility of the f allele and of a less active VDR protein. This study also demonstrates that IL-1-R1 gene single polymorphisms are not associated with an increased risk of T1DM in the Dalmatian population. Thus far the HinfI polymorphism of the promoter region has been linked with T1DM whereas PstI and AluI polymorphism have not.30Busfield SJ Comrack CA Yu G Chickering TW Smutko JS Zhou H Leiby KR Holmgren LM Gearing DP Pan Y Identification and gene organization of three novel members of the IL-1 family on human chromosome 2.Genomics. 2000; 66: 213-216Crossref PubMed Scopus (147) Google Scholar In the high-risk population of Finland, the PstI-e polymorphism in exon 1B of the gene only showed association to T1DM in patients without high-risk HLA-DR3 and DR4 haplotypes.10Metcalfe KA Hitman GA Pociot F Bergholdt R Toumilehto-Wolf E Tuomileht Viswanathan M Ramachandran A Nerup J An association between type 1 diabetes and the interleukin-receptor type 1 gene.Hum Immunol. 1996; 51: 41-48Crossref PubMed Scopus (32) Google Scholar However, the results change when looking at the haplotype level. We found strong LD between all four studied polymorphisms within the IL-1R1 gene. Cox and colleagues6Cox NJ Wapelhorst B Morrison VA Johnson L Punchuk L Spielman RS Todd JA Concannon P Seven regions of the genome show evidence of linkage to type 1 diabetes in a consensus analysis of 767 multiplex families.Am J Hum Genet. 2001; 69: 820-830Abstract Full Text Full Text PDF PubMed Scopus (243) Google Scholar reported a strong LD between PstI and HinfI polymorphisms but not between PstI and AluI. One specific allele combination, phap', is distinguished as being more frequent in T1DM patients of the Dalmatian population. This could be explained with the extended LD throughout the region; thus, the studied polymorphisms could be in LD either with a mutation in a regulatory region of the IL-1-R1 gene or with a different disease locus in this chromosomal region.25Dinarello CA Interleukin-1, interleukin-1 receptors and interleukin-1 receptor antagonists.Int Rev Immunol. 1998; 16: 457-499Crossref PubMed Scopus (682) Google Scholar It has been suggested that the haplotypes, rather than individual genes, incorporating different allelic variants of genes in the IL-1 gene cluster may be important in determining susceptibility to T1DM.25Dinarello CA Interleukin-1, interleukin-1 receptors and interleukin-1 receptor antagonists.Int Rev Immunol. 1998; 16: 457-499Crossref PubMed Scopus (682) Google Scholar To our knowledge this is the first report of linkage of haplotypes of these four IL-1-R1 gene polymorphisms with T1DM. The results of our study provide more evidence of the polygenic nature of T1DM and of the genetic heterogeneity between different populations. We confirm that in the Dalmatian population of South Croatia the VDR FokI polymorphism and several VDR and IL-1-R1 haplotypes correlate with the susceptibility to and development of T1DM. We thank Prof. Mark McCarthy and Eleftheria Zeggini for advice and help in haplotype estimation.
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