Single-cell transcriptome sequencing reveals the cellular heterogeneity of small intestine tissue in celiac disease

Background: Celiac disease (CeD) is an autoimmune small intestinal disease caused by gluten protein ingestion by genetically susceptible individuals. Genome-wide association studies and transcriptomic data have limited capacity to capture intercellular genetic variations. We aim to construct a single-cell transcriptome spectrum, analyze the immune microenvironment and cellular heterogeneity, discover disease-related specific genes and markers, and explore the pathogenesis of CeD. Methods: We performed single-cell RNA sequencing (scRNA-seq) on three small intestine biopsies from patients with CeD and three healthy Chinese controls. Immunohistochemistry (IHC) and quantitative PCR (qPCR) were used to validate potential diagnostic biomarkers of disease-differential genes. Results: A total of 10 cell subpopulations were annotated, including three types of epithelial and stromal cells and seven types of immune cells. IHC revealed a pronounced overexpression of T cell disease-differential genes, TRAT1, BCL11B, and ETS1 in intraepithelial lymphocytes in the CeD group. Further clinical validation using qPCR confirmed that ETS1 (P = 0.010), TRAT1 (P < 0.001), and BCL11B (P = 0.036) w Overall design: Three healthy control individuals (Ctrl group) were recruited from the Health Examination Center of the People's Hospital of Xinjiang Uyghur Autonomous Region. These individuals were carefully selected to match the three patients with CeD in terms of age, sex, and ethnicity, thereby ensuring a controlled comparison. Controls were screened using serum-specific antibodises (anti-tissue transglutaminase antibody and anti-endomysial antibody) to exclude any CeD cases. The exclusion criteria included a history of malignant tumors, other autoimmune diseases, acute cardiovascular or cerebrovascular disease (within the last six months), history of gastrointestinal surgery, contraindications to endoscopic biopsy, and refusal to participate in the study. We performed single-cell RNA sequencing (scRNA-seq) on three small intestine biopsies from patients with CeD and three healthy Chinese controls. Immunohistochemistry (IHC) and quantitative PCR (qPCR) were used to validate potential diagnostic biomarkers of disease-differential genes.