Marco-Antonio Montes-Cano

SLC11A1 promoter gene polymorphisms and fibrosis progression in chronic hepatitis C

Background and aims: The solute carrier family 11 member 1 (SLC11A1) gene (formerly Nramp1) encodes for the protein solute carrier family 11, member 1. It affects susceptibility and clinical outcome of autoimmune and infectious diseases. We investigated the possible role of the functional polymorphism located in the promoter region of SLC11A1 and tumour necrosis factor (TNF) genes in the progression of fibrosis in chronic hepatitis C. Methods: A total of 242 Caucasian Spanish patients with biopsy proven chronic hepatitis C and 194 healthy control subjects were genotyped for SLC11A1 and TNF promoter polymorphisms. Results: No significant differences in the distribution of frequencies among patient and control groups were observed. The SCL11A1 homozygous 2/2 genotype was rarely detected among patients showing advanced fibrosis (2/82; 2.4%) but was highly represented in those with mild fibrosis (29/160; 18.1%; odds ratio (OR) 8.85 (95% confidence interval (CI) 1.9–55.2, pc = 0.002). In patients carrying allele 3 of SLC11A1, the presence of −238 TNF A/G was associated with advanced fibrosis (14/26 (53.8%) v 68/216 (31.4%); OR 2.53 (95% CI 1.03–6.23); p = 0.02). Conclusions: SLC11A1 gene promoter polymorphism could influence fibrosis progression in chronic hepatitis C in that the homozygous genotype 2/2 exerts a protective effect against cirrhosis development. Also, the combination of TNF −238 A/G and the presence of allele 3 is conducive to progression to pre-cirrhotic or cirrhotic stages of the disease.

Clinical Significance of Anti-Multiple Nuclear Dots/Sp100 Autoantibodies

BACKGROUND Autoantibodies against discrete variable-sized dots observed in HEp2 cells by indirect immunofluorescence (IIF) test, called multiple nuclear dots (MND), have been closely associated with primary biliary cirrhosis (PBC). Some authors have argued that this antibody is also present in connective tissue diseases or liver diseases other than PBC as autoimmune chronic active hepatitis, particularly of the cholestatic type. We studied an unselected group of patients routinely tested for autoantibodies and positive for the MND pattern and tried to establish the correlation between the presence of this antibody and their diagnosis. METHODS A commercial ELISA test, using a recombinant 26 kD truncated sequence of the Sp100 protein, corresponding to an immunodominant molecular region, was used to assess the clinical correlation of these autoantibodies in 110 patients showing an anti-MND immunofluorescence pattern. RESULTS One-hundred-and-ten patients were MND positive by IIF. Of these, 100 were Sp100 positive by ELISA. In the Sp100 positive group, 34 had a diagnosis of PBC (30 definite and 4 suspected) while 15 patients had a non-PBC hepatopathy. Unexpectedly, 13 of the MND/Sp100 positive pattern corresponded to systemic lupus erythematosus (SLE) patients and 5 cases to collagen diseases. Another divergence with previous reports was that 34 of the positive patients showed very heterogeneous clinical pictures, different from hepatopathies or collagen diseases. CONCLUSIONS Anti-Sp100 antibodies can be found in many clinical conditions. Testing for MND/Sp100 positivity is useful for the diagnosis of PBC, but only when the right clinical context is present. Other diseases cannot be excluded in first line SLE.Background: Autoantibodies against discrete variable-sized dots observed in HEp2 cells by indirect immunofluorescence (IIF) test, called multiple nuclear dots (MND), have been closely associated with primary biliary cirrhosis (PBC). Some authors have argued that this antibody is also present in connective tissue diseases or liver diseases other than PBC as autoimmune chronic active hepatitis, particularly of the cholestatic type. We studied an unselected group of patients routinely tested for autoantibodies and positive for the MND pattern and tried to establish the correlation between the presence of this antibody and their diagnosis. Methods: A commercial ELISA test, using a recombinant 26 r kD truncated sequence of the Sp100 protein, corresponding to an immunodominant molecular region, was used to assess the clinical correlation of these autoantibodies in 110 patients showing an anti-MND immunofluorescence pattern. Results: One-hundred-and-ten patients were MND positive by IIF. Of these, 100 were Sp100 positive by ELISA. In the Sp100 positive group, 34 had a diagnosis of PBC (30 definite and 4 suspected) while 15 patients had a non-PBC hepatopathy. Unexpectedly, 13 of the MND/Sp100 positive pattern corresponded to systemic lupus erythematosus (SLE) patients and 5 cases to collagen diseases. Another divergence with previous reports was that 34 of the positive patients showed very heterogeneous clinical pictures, different from hepatopathies or collagen diseases. Conclusions: Anti-Sp100 antibodies can be found in many clinical conditions. Testing for MND/Sp100 positivity is useful for the diagnosis of PBC, but only when the right clinical context is present. Other diseases cannot be excluded in first line SLE.

Association of the AIRE gene with susceptibility to rheumatoid arthritis in a European population: a case control study.

IntroductionAIRE is a transcriptional regulator playing a functional role in thymocyte education and negative selection by controlling the expression of peripheral antigens in the thymus. Recently, the AIRE gene was identified as a genetic risk factor for rheumatoid arthritis (RA) in genome wide association (GWA) studies performed in the Japanese population. According to the available data this association is restricted to the Asian population. However, different facts could influence the lack of association in Caucasian populations. The aim of this study was to further investigate the possible role of the AIRE gene in susceptibility to RA in a Caucasian population.MethodsA total of 472 Spanish Caucasian RA patients and 475 ethnically matched controls were included in the study. Three single-nucleotide polymorphisms (SNPs) (rs2776377, rs878081 and rs1055311) with a minor allele frequency >0.05 in the Caucasian population which were not included in the high-throughput platforms used in the GWA studies performed in susceptibility to RA, and two SNPs (rs2075876 and rs1800520) associated with RA in the Japanese population, were selected and genotyped using TaqMan assays.ResultsNo significant differences in the distribution of the alleles of rs2776377, rs2075876, rs1055311 and rs1800520 SNPs between RA patients and controls were observed. Nevertheless, the frequency of the C allele of rs878081 was significantly higher among RA patients (80.5% vs. 74.6% in the control group, pc = 0.012, OR = 1.41, 95%CI 1.13-1.75). Regarding the distribution of the rs878081 genotypes, a higher frequency of CC homozygous individuals was found in the RA patient group (65.56% vs. 56.47% in the control group, pc = 0.013, OR = 1.47, 95%CI 1.12-1.93). The in silico analysis predicted lower affinity to the binding-site of a motif of the transcription NF-κB family and lower transcription levels of AIRE gene for the rs878081C risk variantConclusionsOur findings suggest that the AIRE gene is associated with susceptibility to RA in the Spanish population. Probably, this association has not been detected in the European population in the GWA studies because the earliest high-throughput platforms did not include SNP suitable markers (e.g. rs878081).

Genetic Analysis with the Immunochip Platform in Behçet Disease. Identification of Residues Associated in the HLA Class I Region and New Susceptibility Loci

Behcets disease (BD) is an immuno-mediated vasculitis in which knowledge of its etiology and genetic basis is limited. To improve the current knowledge, a genetic analysis performed with the Immunochip platform was carried out in a population from Spain. A discovery cohort comprising 278 BD cases and 1,517 unaffected controls were genotyped using the Immunochip platform. The validation step was performed on an independent replication cohort composed of 130 BD cases and 600 additional controls. The strongest association signals were observed in the HLA class I region, being HLA-B*51 the highest peak (overall P = 6.82E-32, OR = 3.82). A step-wise conditional logistic regression with classical alleles identified HLA-B*57 and HLA-A*03 as additional independent markers. The amino acid model that best explained the association, includes the position 97 of the HLA-B molecule and the position 66 of the HLA-A. Among the non-HLA loci, the most significant in the discovery analysis were: IL23R (rs10889664: P = 3.81E-12, OR = 2.00), the JRKL/CNTN5 region (rs2848479: P = 5.00E-08, OR = 1.68) and IL12A (rs1874886: P = 6.67E-08, OR = 1.72), which were confirmed in the validation phase (JRKL/CNTN5 rs2848479: P = 3.29E-10, OR = 1.66; IL12A rs1874886: P = 1.62E-08, OR = 1.61). Our results confirm HLA-B*51 as a primary-association marker in predisposition to BD and suggest additional independent signals within the class I region, specifically in the genes HLA-A and HLA-B. Regarding the non-HLA genes, in addition to IL-23R, previously reported in our population; IL12A, described in other populations, was found to be a BD susceptibility factor also in Spaniards; finally, a new associated locus was found in the JRKL/CNTN5 region.

HAVCR1 Gene Haplotypes and Infection by Different Viral Hepatitis C Virus Genotypes

ABSTRACT The hepatitis A virus cellular receptor 1 (HAVCR1) gene is highly polymorphic, and several variants have been associated with susceptibility to allergic and autoimmune diseases. The HAVCR1 gene region was identified as a candidate for hepatitis C virus (HCV) natural clearance in a genotyping study of selected immune response genes in both European-American and African-American populations. The aim of the present study was to explore the influence of HAVCR1 in the outcome of HCV infection in the Spanish population. Three cohorts, consisting of 354 subjects with persistent HCV infection (285 with persistent HCV monoinfection and 69 with natural clearance), 182 coinfected HIV/HCV patients, and 320 controls, were included. Samples were genotyped in several polymorphic positions, insertion/deletion variants in exon 4 and tag single nucleotide polymorphisms (SNPs), in order to define previously described HAVCR1 haplotypes (haplotypes A to D). No statistically significant differences were observed with spontaneous resolution of infection or with viral clearance after treatment. Nevertheless, different rates of infection by viral genotypes (Gs) were observed among the HAVCR1 haplotypes. Individuals bearing haplotype C had the highest viral G1 infection rate when compared to individuals bearing other haplotypes (75.82% versus 57.72%, respectively; corrected P value [Pc], 3.2 × 10−4; odds ratio [OR], 2.30; 95% confidence interval [CI], 1.51 to 3.47). Thus, HAVCR1 could be involved in susceptibility or resistance to infection by a particular HCV genotype.

Variants of the IFI16 Gene Affecting the Levels of Expression of mRNA Are Associated with Susceptibility to Behçet Disease

Objective. Behçet disease (BD) is a multifactorial disease in which infectious agents have been proposed as triggers in genetically predisposed individuals. The aim of our study was to investigate the role of innate immunity receptors, specifically the nucleic acid sensors, in susceptibility to BD. Methods. Seventy-four tag single nucleotide polymorphisms (tSNP) selected in 9 candidate genes (DDX58, IFIH1, TLR3, TLR7, TLR8, AIM2, IFI16, ZBP1, and TLR9) were genotyped in 371 patients and 854 controls. Assays of mRNA expression and allele-specific transcript quantification (ASTQ) were performed in 110 and 50 controls, respectively. Results. Patients and controls were genotyped and 2 tSNP (rs6940 in IFI16 and rs855873 in AIM2) were associated with BD. To confirm this association, these tSNP were genotyped in 850 additional controls, and the total cohort was randomly divided into 2 cohorts. The association of these 2 tSNP with the disease remained in both cohorts. One haplotype (rs6940T-rs855873G) was identified as a risk factor (OR 1.41, 95% CI 1.06–1.86, p = 0.015), and another (rs6940A-rs855873A) as a protective factor (OR 0.65, 95% CI 0.47–0.90, p = 0.009). Samples with the risk haplotype had lower IFI16 expression levels than samples with the protective (0.99 ± 0.29 vs 1.23 ± 0.50, p = 0.022). Consistently, in the ASTQ assays performed with the nonsynonymous rs6940 SNP, the risk allele had lower IFI16 expression levels than the protective (p = 0.027). Conclusion. Our findings suggest association of IFI16, a cytosolic sensor of dsDNA and mediator of the AIM2 inflammasome-dependent pathway, in susceptibility to BD. Differences genetically determined in the levels of this molecule could be the cause of this association.

PTPN22 C1858T Polymorphism and the Outcome of Hepatitis C Virus Infection

The outcome of chronic hepatitis C virus infection varies, depending on viral and host factors. Those mechanisms involved in the control of the innate and adaptive response could have an influence on the outcome of infection. The PTPN22 gene encodes an intracellular lymphoid-specific phosphatase (Lyp) with a lymphocyte activating downregulatory effect. A single-nucleotide polymorphism (SNP) C1858T located on this gene has been associated with autoimmune diseases and bacterial infections. The aim of this study was to assess whether the PTPN22 C1858T polymorphism is related to the outcome of hepatitis C viral infection. A total of 69 patients with spontaneous viral clearance (SVC), 281 patients with chronic hepatitis C (CHC), and 1036 individuals not infected with hepatitis C (NIC) were included in this study. Patients with CHC were stratified according to Scheuer score of hepatic fibrosis from F0-F2 (n = 200) and F3-F4 (n = 81), and according to their response to therapy in patients with sustained responses (SR; n = 103) and non-sustained response (NSR; n = 104). Genotyping of the C1858T polymorphism was performed using TaqMan probes. No statistically significant differences in the distribution of PTPN22 C1858T polymorphism were observed upon comparison of patient group with the NIC group. Also, when the different patient groups were compared to one another, no statistically significant differences were detected: the SVC with the CHC group (10.2% versus 12.5%; p = 0.6), the F0-F2 with the F3-F4 group (11.5% versus 14.8%; p = 0.5), and the NSR with the SR group (11.5% versus 14.6%; p = 0.4). Our results do not support a major role of this polymorphism of the PTPN22 gene in the outcome of chronic hepatitis C virus infection in the Spanish population.

Hereditary spherocytosis associated with mutations in HFE gene.

We report on a Spanish family in which three members of different generations were diagnosed with hereditary spherocytosis (HS). Additionally, one of them II-I (44-years-old), presented iron overload with hepatic deposit and needed treatment with periodic phlebotomies. The rest of the family members presented normal analytical values in iron metabolism. To investigate the presence of H63D and C282Y mutations in the HFE gene, patient II-I was found to be compound heterozygous and was the only family member presenting HS and this genetic condition in HFE. We propose a synergistic effect of HS and mutations in HFE as the cause of the iron deposits.

Mutational profile of rare variants in inflammasome-related genes in Behçet disease: A Next Generation Sequencing approach

Behçet’s disease (BD) is an immune-mediated systemic disorder with a well-established association with HLA class I and other genes. BD has clinical overlap with many autoinflammatory diseases (AIDs). The aim of this study was to investigate the role of rare variants in seven genes involved in AIDs: CECR1, MEFV, MVK, NLRP3, NOD2, PSTPIP1 and TNFRSF1A using a next generation sequencing (NGS) approach in 355 BD patients. To check global association of each gene, 4 tests: SKAT, CollapseBt, C(α) and weighted KBAC were used. Databases: 1000 Genomes Project Phase 3, Infevers, HGMD and ClinVar and algorithms: PolyPhen2 and SIFT were consulted to collect information of the 62 variants found. All the genes resulted associated using SKAT but only 3 (MVK, NOD2 and PSTPIP1) with C(α) and weighted KBAC. When all the genes are considered, 40 variants were associated to AIDs in clinical databases and 25 were predicted as pathogenic at least by one of the algorithms. Including only MVK, NOD2 and PSTPIP1, the associated to AIDs variants found in BD were 20 and the predicted as pathogenic, 12. The maxima contribution corresponds to NOD2. This study supports influence of rare variants in genes involved in AIDs in the pathogenesis of BD.

Reply to Bogdanos et al.

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