Endocrinology and Diabetes Research Group, BioCruces Health Research Institute, UPV-EHU, CIBERDEM, Barakaldo.
Department of Genetics, Physical Anthropology and Animal Physiology, University of the Basque Country (UPV-EHU), BioCruces Health Research Institute, Leioa.
J Pediatr Gastroenterol Nutr. 2018 Aug;67(2):225-231. doi: 10.1097/MPG.0000000000001970.
The aim of the study is to identify additional celiac disease associated loci in the major histocompatibility complex (MHC) independent from classical HLA risk alleles (HLA-DR3-DQ2) and to characterize their potential functional impact in celiac disease pathogenesis at the intestinal level.
We performed a high-resolution single-nucleotide polymorphism (SNP) genotyping of the MHC region, comparing HLA-DR3 homozygous celiac patients and non-celiac controls carrying a single copy of the B8-DR3-DQ2 conserved extended haplotype. Expression level of potential novel risk genes was determined by RT-PCR in intestinal biopsies and in intestinal and immune cells isolated from control and celiac individuals. Small interfering RNA-driven silencing of selected genes was performed in the intestinal cell line T84.
MHC genotyping revealed 2 associated SNPs, one located in TRIM27 gene and another in the non-coding gene HCG14. After stratification analysis, only HCG14 showed significant association independent from HLA-DR-DQ loci. Expression of HCG14 was slightly downregulated in epithelial cells isolated from duodenal biopsies of celiac patients, and eQTL analysis revealed that polymorphisms in HCG14 region were associated with decreased NOD1 expression in duodenal intestinal cells.
We have successfully employed a conserved extended haplotype-matching strategy and identified a novel additional celiac disease risk variant in the lncRNA HCG14. This lncRNA seems to regulate the expression of NOD1 in an allele-specific manner. Further functional studies are needed to clarify the role of HCG14 in the regulation of gene expression and to determine the molecular mechanisms by which the risk variant in HCG14 contributes to celiac disease pathogenesis.
本研究旨在鉴定主要组织相容性复合体（MHC）中与经典 HLA 风险等位基因（HLA-DR3-DQ2）无关的其他乳糜泻相关基因座，并在肠道水平上阐明其在乳糜泻发病机制中的潜在功能影响。
我们对 MHC 区域进行了高分辨率单核苷酸多态性（SNP）基因分型，比较了 HLA-DR3 纯合乳糜泻患者和携带 B8-DR3-DQ2 保守扩展单体型单拷贝的非乳糜泻对照者。通过 RT-PCR 测定潜在新风险基因在肠道活检组织以及从对照和乳糜泻个体分离的肠道和免疫细胞中的表达水平。在肠道细胞系 T84 中用小干扰 RNA 驱动的基因沉默。
MHC 基因分型显示 2 个相关 SNP，一个位于 TRIM27 基因，另一个位于非编码基因 HCG14。分层分析后，只有 HCG14 显示与 HLA-DR-DQ 基因座无关的显著相关性。乳糜泻患者十二指肠活检上皮细胞中 HCG14 的表达略有下调，eQTL 分析显示 HCG14 区域的多态性与十二指肠肠道细胞中 NOD1 表达降低相关。
我们成功地采用了保守扩展单体型匹配策略，鉴定了长链非编码 RNA HCG14 中的乳糜泻新风险变异。该长链非编码 RNA 似乎以等位基因特异性方式调节 NOD1 的表达。需要进一步的功能研究来阐明 HCG14 在基因表达调控中的作用，并确定 HCG14 风险变异在乳糜泻发病机制中的分子机制。
