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  • Mechanism underlying the significant role of the miR-4262/SIRT1 axis in children with inflammatory bowel disease.

Mechanism underlying the significant role of the miR-4262/SIRT1 axis in children with inflammatory bowel disease.

Experimental and therapeutic medicine (2020-08-09)
Xiaozhi Deng, Lihong Shang, Min Du, Lan Yuan, Lijing Xiong, Xiaoli Xie
ABSTRACT

Inflammatory bowel disease (IBD) is a term used to describe chronic and recurrent gastrointestinal disease. In total, >2 million individuals worldwide have been diagnosed with IBD, including ulcerative colitis (UC), Crohn's disease (CD) and indeterminate colitis. There is accumulating evidence to indicate that microRNAs (miRNAs or miRs) are involved in the development and progression of IBD. miR-4262, an underlying promoter in tumor diseases, has been reported to regulate inflammatory responses. However, the potential mechanisms underlying the role of miR-4262 in IBD remain unknown. The present study attempted to explore the role and mechanisms of miR-4262 in IBD. Firstly, reverse transcription-quantitative PCR (RT-qPCR) was used to detect the expression of miR-4262 in 30 IBD colonic mucosa tissues, 30 normal tissues, 2% dextran sulfate sodium (DSS)-treated Caco-2 cells and normal cells. It was demonstrated that the expression levels of miR-4262 in IBD colonic mucosa tissues and 2% DSS-stimulated Caco-2 cells were markedly higher compared with those in the control groups. Target gene prediction databases and dual-luciferase reporter assays were then used, and sirtuin 1 (SIRT1) was identified as a target gene of miR-4262. Furthermore, the levels of SIRT1 in 2% DSS-stimulated Caco-2 cells and IBD colonic mucosa tissues were suppressed compared with the corresponding control groups. In addition, it was observed that miR-4262 negatively regulated SIRT1 expression in Caco-2 cells. Thereafter, Caco-2 cells were treated with inhibitor control, miR-4262 inhibitor, control-siRNA or SIRT1-siRNA for 48 h, followed by 2% DSS treatment for 4 days. The secretion of inflammatory factors was analyzed via ELISA and RT-qPCR. MTT assay, flow cytometry and western blot analysis were performed to assess cell viability, apoptosis and NF-κB signaling pathway-related protein levels, respectively. The results indicated that DSS enhanced the inflammatory response, suppressed cell viability and promoted cell apoptosis, and this was decreased following transfection with an miR-4262 inhibitor. In addition, 2% DSS upregulated p-p65 expression and enhanced the ratio of p-p65/p65, while the miR-4246 inhibitor exerted an opposite effect. All the effects of miR-4262 inhibitor on Caco-2 cells were eliminated following transfection with SIRT1-siRNA. It was thus concluded that miR-4262 may serve a role in the progression of IBD via targeting SIRT1, and miR-4262/SIRT1 may represent a potential target for the diagnosis and treatment of IBD.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Anti-SIRT1 antibody produced in rabbit, ~1.0 mg/mL, affinity isolated antibody, buffered aqueous solution
Sigma-Aldrich
Anti-GAPDH antibody produced in rabbit, ~1 mg/mL, affinity isolated antibody, buffered aqueous solution
Sigma-Aldrich
Anti-phospho-NF-kappaB p65 (pSer276) antibody produced in rabbit, affinity isolated antibody
Sigma-Aldrich
Anti-NF-κB p65 antibody produced in rabbit, affinity isolated antibody