- Email: [email protected]
Association of interleukin-1α and interleukin-1β gene alleles with ulcerative colitis: Evidence for genetic susceptibility
susceptibility and phenotype may be determined by MICNB alleles. Methods: A PCR-SSP system was developedto identify the 45 known MICAand 9 MICBalleles.This typing technique was applied to 319 UC patients divided into 3 phenotypic subgroups (116 total colitis + no surgery; 100 total colitis + proctocolectomy / lieu-anal pouch formation; 103 proctitis + no surgery), 110 CD patientswith stricturing small bowel diseaseand 296 healthyOxfordshire controls recruitedfrom Generalpractice health screeningclinics. Results: Of the 45 previously reported MICA alleles 19 and 18 were identified in the patient and control populations respectively.5 of the previously reported MICB alleles were identified in both the control and patient groups. 10 new alleles (frequency less than 1%) were identified (4 controls, 6 UC or CD). No significant difference was.found in allele frequencies between controls and UC or CD patients when analysedby IBD overall, diseasetype (CD or UC), or phenotypicsubgroup. Conclusion: Polymorphisms in exons 2-4 of the MICNB genesappearnot determinesusceptibility or phenotype in patients with UC or CD. Further work is now being carded out to evaluate the role of polymorphisms in exon 5 that encodes the transmembrane portion of the MIC molecule.
IL-IA and IL-IB GenotypesIn UlcerativeColitis PatientsAnd Controls +3954 ILl B Controls
1,1 609
UC
204
Surgical Nonsurgical
148 56
1,2 43 5 10 2 77 25
-511 IL1B
+4845ILIA
2,2 72
1,1 490
1,2 486
2,2 140
1,1 524
1,2 487
2,2 105
12
129
154
36
177
122
20
9 3
102 27
110 44
23 13
132 45
87 35
16 4
2337 An Estimate Of Critical Region For Ulcerative Colitis in The Human Major Histocompatibility Complex Using Refined Microsatellite Mapping Kazuhito Sugimura, Niigata Univ Sch of Medicine, Niigata Japan; Masao Ota, Shinshu Univ Sch of Medicine, Matsumoto Japan; Jun Matsuzawa,Niigata Univ Sch of Medicine, Niigata Japan; Yoshihiko Katsuyama, Shinshu Univ Sch of Medicine, Matsumoto Japan; Suzuko S. Seki, Terasu Honma, Niigata Univ Sch of Medicine, Niigata Japan; Hidetoshi Inoko, Tokai Univ Sch of Medicine, IseharaJapan; Hitoshi Asakura, Niigata Univ Sch of Medicine, Niigata Japan
2335 Mannan-binding LecUn Gene Polymorphismsin Ulcerative Colitis and Crohn's Disease Annabel Rector, Philippe Lemey, Wire Laffut, EIs Keyaerts,Frank Struyf, Elke Woliants, Rega Institute, Univ of Leuven, Leuven Belgium; SeverineVermeire, Paul Rutgeerts, Dept of Gastroenterology,UZ Gasthuisberg, Leuven Belgium; Marc Van Ranst, Rega Institute, Univ of Leuven, Leuven Belgium
Background & Aims: As ulcerative colitis (UC) is the complex multifactorial traits involving both environmental and genetic factors, the responsible loci for disease susceptibility is difficult to be determined. In Japanese patients with UC, we have reported a conserved haplotype of HLA A24-B52-DR2(15)-DPw9 and a significantly high frequency of the MHC class I chain-relatedgene A (MICA)A6 allele. To precisely map the responsiblegene involved in the developmentof UC, twenty-five markers distributed over a whole human MHC region were subjectedto associationanalysis.Methods: Forty-ninepatientswith UCand 236 unrelated controls were included in this study. All subjects were Japanese.Twenty-two microsatellite markers, sevenof which werefound recentlyby our largegenomesequencing,were determined by direct sequencing procedures. HLA-A, B, and DR serotype were also decided. The significance of associationwas tested by the ;<2method. All Pvalues were corrected by multiplication by the number of decided alleles of each locus (Pc). In addition, linkage disequilibria among associatedmarkers were evaluatedwith t value. Results: One or more alleles showed significantly higher frequencies in patients with UC at many loci, compared with controls. Two patients were homozygousfor susceptible haplotype(B52-DR2(15)), and more detailedalleles about the susceptible haplotype were determined. Among these alleles, the maximum odds ratio (OR) and lowest Pcvalue of phenotypefrequency were observed at the marker of C12-A (allele 238), located approximately 147 kb centromeric to the HLA-B gene (0R=4.37, 95%CI 2.35-8.12, Pc = 0.000042). No significant associationwas observedat the microsatellite markers distributed both centromeric to HLA-DP and telomeric to HLA-A locus. The highest t value was observed between C1-2-A (allele 238) and TNF-a (allele 119) (t value= 14.62), and this region of susceptible haplotype was strongly conserved in Japanese patients with UC. Conclusions: These observations may indicate the existence of important determinants of diseasesusceptibility to UC betweenC1-2-A and TNF-a microsatellite markers on the short arm of chromosome 6. Interestingly, this conserved region does not contain the classical HLA locus. High resolution mapping of responsible gene locus in this region will provide a new view on the pathogenesisof UC.
Introduction: Ulcerativecolitis (UC) and Crohn's disease(CD) are complex multifactorial traits involving both environmental and genetic factors. Mannan binding lectin (MBL) plays an important role in non-specific immunity as well as in complement activation. Point mutations have been identified in codons 52, 54 and 57 of exon 1 in the MBL gene, and are associated with decreasedMBL plasma concentrationsand increasedsusceptibility to various infectious diseases. It is plausiblethat these MBL mutations could lead to susceptibility to putative IBDetiological microbial agents, or could temper the complement-mediatedmucosal damage in IBD. MBL could function as the link between microbial, immunological and genetic factors in IBO. Aim: We investigatedthe presenceof point mutations at codon 52, 54 and 57 of the MBL gene in a large cohort of IBO-patientsand in an unaffected control population. Methods: Blood sampleswere obtainedfrom 340 unrelated IBD-patients(216 CD, 124 UC). DNAsamples of 308 controls were obtained through a non-invasive swish-and-spit technique. The MSL gene polymcrphisms were investigated using polymerase chain reaction (PCR) restriction fragment length polymorphism for detection of the codon 54 and 57 mutations, and an amplification refractory mutation system PCR(ARMS-PCR)for the codon 52 mutation. Results: The genotypes in all study populations were in Hardy-Weinbergequilibrium. The frequency of the investigated MBL mutations was significantly lower in UC patients when compared with CD patients (X2=6.703, p=0.0096) and with the control population (X2=5.445, p =O.0196)(Table 1). Conclusion:These results suggest a protective effect of MBL mutations in UC, but not in CD. This can be explained by the differential T-helper response in both diseases.The predominant TH2-responsein UC increases IL-6 production, and IL-6 increases the expressionof MBL. MBL could therefore be responsiblefor more extensivecomplementmediated mucosal damage in UC than in CD. MBL mutations that decreasethe expression of functional MBL could protect against the clinical developmentof UC. Table1: MBL variantgenefrequenciesin IBD (CD, UC) and controls MBLvadants
IBD (n=340)
CO (n=216)
UC (n=124)
Co (n=308)
Homozygotes and compound heterozygotes Heterozyotes
13 (3.8%)
11 (5.1%)
2 (1.6%)
16 (5.2%)
2338
116(34.1%)
82(38°/o)
34(27.4%)
110(35.7%}
The IKGL Gene Confers High Risk to Extensive Ulcerative Colitis. Gontrand Lopez-Nava,Julio Garcia Parades, Miguel Fernandez-Arquero,Emilio G. Concha, Manuel Diaz-Rubio,San Carlos Clin Hosp, Madrid Spain BACKGROUND&AIMSThe pathogenesisof UC remains largely unclear.There is an increasing interest in the role of cytokines in UC pathogenesisand on the polymorphic genes that may influence cytokine secretion. Among them we have chosen to study a substitution in the IKBL gene.The IKBL protein is most similar to IKBa, which regulatesthe nuclear localization of NFKB, a nuclear factor that stimulates the transcription of TNF and other cytokines. IKBL gene lies telomeric of the TNFcluster in the central major HLA complex and carries a structural polymorphism at position + 738. In the Spanishwhite population we found the IKBL+ 738(C) allele in haplotypes carrying either HLA-ORB1*1501 or DRB1"0103. As these HLA class II alleles may confer susceptibility to ulcerativecolitis, we investigatedan association between IKBL+738(C) and this disease. PATIENTS&METHODSDNA based techniques were used to type individual alleles of HLA-DRB1 and IKBL+738.The frequencies were compared among ethnically matched populations comprising 155 patients and 298 controls. RESULTS: 1)A statistically significant increase of the IKBL+738(C) allele was observed in UC patients (16.8%versus 6.7% in controls; OR= 2.80; p
2336 Association of Interleukin-lot and Interleukin-l,8 Gene Alleles with Ulcerative Colitis: Evidence for Genetic Susceptibility Martyn J. Carter, Gastroenterologyand Liver Unit, P,oyal Haliamshire Hosp, $heflietd United Kingdom; FrancescoS. Di Giovine, Angeia Cox, Julian Sorrell, Gordon W. Duff, Alan J. Lobo, Div of MGM, Univ of Sheffield, Sheffield United Kingdom Background; Case-control studies in ulcerative colitis (UC) assessing single nucleotide polymorphisms (SNP) within the intereukin-1 genes have not demonstrated significant associations. However,these studies had limited power for the detection of genes of modest effect on disease susceptibility. Aims; To further assess the IL-I~ (IL1A) and IL-1/~(IL1B) genes as candidate loci for disease susceptibility and severity in UC in a large well characterised cohort of patients. Patientsand Methods; 319 UC patientswhose diseasediagnosisand extent had beenconfirmed by endoscopic and histological criteria were studied. 84 of these patients had undergonecolectomy. 1116 anonymousblood donors were usedas controls. All individuals were genotyped for the -511 and +3954 SNPs in IL18 and the +4845 SNP in IL1A using the Taqmanallelic discrimination system. Chi-squaredstatistics were used to compare allele carriage ratesand odds ratios were catcalated.Resorts;Homozygouscarriageof + 3954 allele 1 was associatedwith UC (p = 0.002; OR 1.5(95% C1:12-1.9)) but not surgery (p>O.05). The -511 allele 2 was not associated with disease (p>O.05) but was increased in surgical compared to non-surgical patients (p=O.07; OR 1.6 (95% CI: 0.96-2.7)). Carriage of the + 4845 allele1 was associatedwith disease(p = 0.007; OR1.4 (95% CI: 1.1-1.8)). Conclusions; Genes in the IL-1 gene cluster other than the IL-1 receptor antagonist gene may have a role in determining disease susceptibility and behaviour.
A-459
IL-IA and IL-IB GenotypesIn UlcerativeColitis PatientsAnd Controls +3954 ILl B Controls
1,1 609
UC
204
Surgical Nonsurgical
148 56
1,2 43 5 10 2 77 25
-511 IL1B
+4845ILIA
2,2 72
1,1 490
1,2 486
2,2 140
1,1 524
1,2 487
2,2 105
12
129
154
36
177
122
20
9 3
102 27
110 44
23 13
132 45
87 35
16 4
2337 An Estimate Of Critical Region For Ulcerative Colitis in The Human Major Histocompatibility Complex Using Refined Microsatellite Mapping Kazuhito Sugimura, Niigata Univ Sch of Medicine, Niigata Japan; Masao Ota, Shinshu Univ Sch of Medicine, Matsumoto Japan; Jun Matsuzawa,Niigata Univ Sch of Medicine, Niigata Japan; Yoshihiko Katsuyama, Shinshu Univ Sch of Medicine, Matsumoto Japan; Suzuko S. Seki, Terasu Honma, Niigata Univ Sch of Medicine, Niigata Japan; Hidetoshi Inoko, Tokai Univ Sch of Medicine, IseharaJapan; Hitoshi Asakura, Niigata Univ Sch of Medicine, Niigata Japan
2335 Mannan-binding LecUn Gene Polymorphismsin Ulcerative Colitis and Crohn's Disease Annabel Rector, Philippe Lemey, Wire Laffut, EIs Keyaerts,Frank Struyf, Elke Woliants, Rega Institute, Univ of Leuven, Leuven Belgium; SeverineVermeire, Paul Rutgeerts, Dept of Gastroenterology,UZ Gasthuisberg, Leuven Belgium; Marc Van Ranst, Rega Institute, Univ of Leuven, Leuven Belgium
Background & Aims: As ulcerative colitis (UC) is the complex multifactorial traits involving both environmental and genetic factors, the responsible loci for disease susceptibility is difficult to be determined. In Japanese patients with UC, we have reported a conserved haplotype of HLA A24-B52-DR2(15)-DPw9 and a significantly high frequency of the MHC class I chain-relatedgene A (MICA)A6 allele. To precisely map the responsiblegene involved in the developmentof UC, twenty-five markers distributed over a whole human MHC region were subjectedto associationanalysis.Methods: Forty-ninepatientswith UCand 236 unrelated controls were included in this study. All subjects were Japanese.Twenty-two microsatellite markers, sevenof which werefound recentlyby our largegenomesequencing,were determined by direct sequencing procedures. HLA-A, B, and DR serotype were also decided. The significance of associationwas tested by the ;<2method. All Pvalues were corrected by multiplication by the number of decided alleles of each locus (Pc). In addition, linkage disequilibria among associatedmarkers were evaluatedwith t value. Results: One or more alleles showed significantly higher frequencies in patients with UC at many loci, compared with controls. Two patients were homozygousfor susceptible haplotype(B52-DR2(15)), and more detailedalleles about the susceptible haplotype were determined. Among these alleles, the maximum odds ratio (OR) and lowest Pcvalue of phenotypefrequency were observed at the marker of C12-A (allele 238), located approximately 147 kb centromeric to the HLA-B gene (0R=4.37, 95%CI 2.35-8.12, Pc = 0.000042). No significant associationwas observedat the microsatellite markers distributed both centromeric to HLA-DP and telomeric to HLA-A locus. The highest t value was observed between C1-2-A (allele 238) and TNF-a (allele 119) (t value= 14.62), and this region of susceptible haplotype was strongly conserved in Japanese patients with UC. Conclusions: These observations may indicate the existence of important determinants of diseasesusceptibility to UC betweenC1-2-A and TNF-a microsatellite markers on the short arm of chromosome 6. Interestingly, this conserved region does not contain the classical HLA locus. High resolution mapping of responsible gene locus in this region will provide a new view on the pathogenesisof UC.
Introduction: Ulcerativecolitis (UC) and Crohn's disease(CD) are complex multifactorial traits involving both environmental and genetic factors. Mannan binding lectin (MBL) plays an important role in non-specific immunity as well as in complement activation. Point mutations have been identified in codons 52, 54 and 57 of exon 1 in the MBL gene, and are associated with decreasedMBL plasma concentrationsand increasedsusceptibility to various infectious diseases. It is plausiblethat these MBL mutations could lead to susceptibility to putative IBDetiological microbial agents, or could temper the complement-mediatedmucosal damage in IBD. MBL could function as the link between microbial, immunological and genetic factors in IBO. Aim: We investigatedthe presenceof point mutations at codon 52, 54 and 57 of the MBL gene in a large cohort of IBO-patientsand in an unaffected control population. Methods: Blood sampleswere obtainedfrom 340 unrelated IBD-patients(216 CD, 124 UC). DNAsamples of 308 controls were obtained through a non-invasive swish-and-spit technique. The MSL gene polymcrphisms were investigated using polymerase chain reaction (PCR) restriction fragment length polymorphism for detection of the codon 54 and 57 mutations, and an amplification refractory mutation system PCR(ARMS-PCR)for the codon 52 mutation. Results: The genotypes in all study populations were in Hardy-Weinbergequilibrium. The frequency of the investigated MBL mutations was significantly lower in UC patients when compared with CD patients (X2=6.703, p=0.0096) and with the control population (X2=5.445, p =O.0196)(Table 1). Conclusion:These results suggest a protective effect of MBL mutations in UC, but not in CD. This can be explained by the differential T-helper response in both diseases.The predominant TH2-responsein UC increases IL-6 production, and IL-6 increases the expressionof MBL. MBL could therefore be responsiblefor more extensivecomplementmediated mucosal damage in UC than in CD. MBL mutations that decreasethe expression of functional MBL could protect against the clinical developmentof UC. Table1: MBL variantgenefrequenciesin IBD (CD, UC) and controls MBLvadants
IBD (n=340)
CO (n=216)
UC (n=124)
Co (n=308)
Homozygotes and compound heterozygotes Heterozyotes
13 (3.8%)
11 (5.1%)
2 (1.6%)
16 (5.2%)
2338
116(34.1%)
82(38°/o)
34(27.4%)
110(35.7%}
The IKGL Gene Confers High Risk to Extensive Ulcerative Colitis. Gontrand Lopez-Nava,Julio Garcia Parades, Miguel Fernandez-Arquero,Emilio G. Concha, Manuel Diaz-Rubio,San Carlos Clin Hosp, Madrid Spain BACKGROUND&AIMSThe pathogenesisof UC remains largely unclear.There is an increasing interest in the role of cytokines in UC pathogenesisand on the polymorphic genes that may influence cytokine secretion. Among them we have chosen to study a substitution in the IKBL gene.The IKBL protein is most similar to IKBa, which regulatesthe nuclear localization of NFKB, a nuclear factor that stimulates the transcription of TNF and other cytokines. IKBL gene lies telomeric of the TNFcluster in the central major HLA complex and carries a structural polymorphism at position + 738. In the Spanishwhite population we found the IKBL+ 738(C) allele in haplotypes carrying either HLA-ORB1*1501 or DRB1"0103. As these HLA class II alleles may confer susceptibility to ulcerativecolitis, we investigatedan association between IKBL+738(C) and this disease. PATIENTS&METHODSDNA based techniques were used to type individual alleles of HLA-DRB1 and IKBL+738.The frequencies were compared among ethnically matched populations comprising 155 patients and 298 controls. RESULTS: 1)A statistically significant increase of the IKBL+738(C) allele was observed in UC patients (16.8%versus 6.7% in controls; OR= 2.80; p
2336 Association of Interleukin-lot and Interleukin-l,8 Gene Alleles with Ulcerative Colitis: Evidence for Genetic Susceptibility Martyn J. Carter, Gastroenterologyand Liver Unit, P,oyal Haliamshire Hosp, $heflietd United Kingdom; FrancescoS. Di Giovine, Angeia Cox, Julian Sorrell, Gordon W. Duff, Alan J. Lobo, Div of MGM, Univ of Sheffield, Sheffield United Kingdom Background; Case-control studies in ulcerative colitis (UC) assessing single nucleotide polymorphisms (SNP) within the intereukin-1 genes have not demonstrated significant associations. However,these studies had limited power for the detection of genes of modest effect on disease susceptibility. Aims; To further assess the IL-I~ (IL1A) and IL-1/~(IL1B) genes as candidate loci for disease susceptibility and severity in UC in a large well characterised cohort of patients. Patientsand Methods; 319 UC patientswhose diseasediagnosisand extent had beenconfirmed by endoscopic and histological criteria were studied. 84 of these patients had undergonecolectomy. 1116 anonymousblood donors were usedas controls. All individuals were genotyped for the -511 and +3954 SNPs in IL18 and the +4845 SNP in IL1A using the Taqmanallelic discrimination system. Chi-squaredstatistics were used to compare allele carriage ratesand odds ratios were catcalated.Resorts;Homozygouscarriageof + 3954 allele 1 was associatedwith UC (p = 0.002; OR 1.5(95% C1:12-1.9)) but not surgery (p>O.05). The -511 allele 2 was not associated with disease (p>O.05) but was increased in surgical compared to non-surgical patients (p=O.07; OR 1.6 (95% CI: 0.96-2.7)). Carriage of the + 4845 allele1 was associatedwith disease(p = 0.007; OR1.4 (95% CI: 1.1-1.8)). Conclusions; Genes in the IL-1 gene cluster other than the IL-1 receptor antagonist gene may have a role in determining disease susceptibility and behaviour.
A-459