Effects of circular RNA circ-SOD2 on intestinal epithelial barrier and ulcerative colitis.
- Author:
Ting Ting WANG
1
;
Ying HAN
1
;
Fang Fang GAO
1
;
Lei YE
1
;
Yu Jun ZHANG
1
Author Information
1. The Central Laboratory, Peking University People's Hospital, Beijing 100044, China.
- Publication Type:Journal Article
- MeSH:
Caco-2 Cells;
Colitis, Ulcerative;
Humans;
In Situ Hybridization, Fluorescence;
RNA;
Superoxide Dismutase/metabolism*
- From:
Journal of Peking University(Health Sciences)
2019;51(5):805-812
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE:To explore the expression profiling of circRNAs in ulcerative colitis(UC) and then determine the significantly changed circRNA and its influences on intestinal epithelial barrier.
METHODS:In this study, we selected 5 pairs of inflamed and normal colorectal mucosa tissues from UC patients to perform circRNAs microarray and identified the differentially expressed circRNAs in the UC inflamed colorectal mucosa tissues, and quantitative real-time PCR was used to identify the expression change of circ-SOD2 in 30 UC patients' inflamed and normal colorectal mucosa tissues. We detected the expression of circ-SOD2 in Caco2 and NCM460 cells after being treated with inflammatory factors (LPS, TNF-α, IL1-β). Fluorescence in situ hybridization (FISH) was used to determine the cellular location of circ-SOD2 in the UC colorectal mucosal tissues. The circ-SOD2 overexpression vector was constructed and produced and then transfected into Caco2 cells to examine the cells' trans-epithelial electrical resistance (TEER), permeability of FITC-dextran and the alterations of epithelial barrier related molecules.
RESULTS:We found 264 circRNAs (111 increased and 153 decreased) differentially expressed in the inflamed colon mucosa compared with normal colon mucosa using a P-value <0.05 and a >1.5-fold change cutoff. To validate the circRNA microarray results, we selected some circRNAs to perform qRT-PCR based on the following criteria: (1) circRNAs raw data >100 in each sample, (2) fold-change >2, (3) P<0.05. We identified 10 dysregulated circRNA, among them, circ-SOD2 was upregulated with maximum fold-change in the UC inflamed colorectal mucosa tissues. Then we identified circ-SOD2 was upregulated significantly through quantitative real-time PCR (qRT-PCR) in expanded 30 paired colorectal mucosa tissues(P<0.001). After treatments with LPS, TNF-α and IL1-β, circ-SOD2 was upregulated in Caco2 and NCM460 cells at different points from 1 to 7 h. Fluorescence in situ hybridization (FISH) indicated that circ-SOD2 located in intestinal epithelium mostly and few in mesenchyme and inflammatory cells. The overexpression of circ-SOD2 in Caco2 cells resulted in a decrease of transepithelial electrical resistance (TEER), an increase of the FITC-dextran permeability and the downregulation of epithelial barrier related molecule CLDN-8 (P<0.05).
CONCLUSION:The dysregulation of circRNAs existed in UC inflamed colorectal mucosa, among which, the upregulated circ-SOD2 weakened the intestinal epithelial barrier and thus might promote the occurrence of ulcerative colitis.
