Table 3_Targeted next-generation sequencing of candidate regions identified by GWAS revealed SNPs associated with IBD in GSDs.docx

Canine Inflammatory bowel disease (IBD) is a chronic multifactorial disease, resulting from complex interactions between the intestinal immune system, microbiota and environmental factors in genetically predisposed dogs. Previously, we identified several single nucleotide polymorphisms (SNP) and regions on chromosomes (Chr) 7, 9, 11 and 13 associated with IBD in German shepherd dogs (GSD) using GWAS and FST association analyses. Here, building on our previous results, we performed a targeted next-generation sequencing (NGS) of a two Mb region on Chr 9 and 11 that included 14 of the newly identified candidate genes, to identify potential functional SNPs that could explain these association signals. Furthermore, correlations between genotype and treatment response were estimated. Results revealed several SNPs in the genes for canine EEF1A1, MDH2, IL3, IL4, IL13 and PDLIM. Based on the known function of their human orthologues, these results further our insight into their potential contribution to the pathogenesis of IBD in dogs. In addition, several pathways involved in innate and adaptive immunity and inflammatory responses (i.e., T helper cell differentiation, Th1 and Th2 activation pathway, communication between innate and adaptive immune cells and differential regulation of cytokine production in intestinal epithelial cells by IL-17A and IL-17F), were constructed involving the gene products in the candidate regions for IBD susceptibility. Interestingly, some of the identified SNPs were present in only one outcome group, suggesting that different genetic factors are involved in the pathogenesis of IBD in different treatment response groups. This also highlights potential genetic markers to predict the response in dogs treated for IBD.

犬类炎症性肠病（Canine Inflammatory Bowel Disease, IBD）是一种慢性多因素疾病，由遗传易感犬的肠道免疫系统、微生物群与环境因素之间的复杂相互作用所引发。此前，我们通过全基因组关联分析（Genome-Wide Association Study, GWAS）和FST关联分析，在德国牧羊犬（German Shepherd Dog, GSD）中鉴定出多个与IBD相关的单核苷酸多态性（single nucleotide polymorphism, SNP）以及7、9、11、13号染色体上的相关区域。本研究基于此前的研究结果，对9号和11号染色体上的2 Mb区域进行了靶向下一代测序（next-generation sequencing, NGS），该区域包含14个新鉴定的候选基因，旨在挖掘能够解释上述关联信号的潜在功能性SNP。此外，本研究还估算了基因型与治疗应答之间的相关性。研究结果在犬EEF1A1、MDH2、IL3、IL4、IL13及PDLIM基因中发现了多个SNP。基于这些基因在人类中的同源基因已知功能，本研究结果进一步阐明了它们在犬IBD发病机制中的潜在作用。此外，本研究构建了多条参与固有免疫、适应性免疫及炎症应答的通路，包括辅助T细胞分化、Th1与Th2细胞激活通路、固有免疫与适应性免疫细胞间的通讯，以及IL-17A和IL-17F对肠上皮细胞细胞因子产生的差异性调控；这些通路均涉及犬IBD易感候选区域内的基因产物。值得注意的是，部分鉴定出的SNP仅存在于单一结局组别中，这表明不同治疗应答组的犬IBD发病机制涉及不同的遗传因素。该发现同时也为预测犬IBD治疗应答提供了潜在的遗传标志物。