S Wang1, L Cui1, X Liu1, Z Luo1, H Li1, J Pu1. 1. Department of Gastroenterology, First Medical Center of Chinese PLA General Hospital, Beijing 100853, China.
Abstract
OBJECTIVE: To explore the effect of WDSUB1 on dextran sulfate sodium (DSS)-induced inflammatory colon injury in mice and the underlying mechanism. METHODS: Different WDSUB1 siRNA sequences were transfected into mouse fibroblast L929 cells and the optimal sequence was selected by Western blotting. Twelve male C57BL/6 mice were randomized into two groups for injection of siWDSUB1 or siControl via the caudal vein, followed by treatment with 2.5% DSS in drinking water to establish mouse models of DSS- induced colitis (n=6). The expression level of WDSUB1 in the colon tissue of the mice was detected with Western blotting and RT-PCR, the changes in body weight and fecal condition were recorded, and the clinical symptoms of the mice were evaluated. The mRNA expression levels of IL-6, COX-2 and TNF-α and the protein expression of IκBα and P65 in the colon tissues were detected with RT- PCR and Western blotting, respectively. RESULTS: The mRNA and protein expressions of WDSUB1 in the colon tissues were significantly lower in colitis mice with WDSUB1 knock-down than in the control mice. Compared with the control mice, the mice receiving siWDSUB1 injection showed obviously milder weight loss, diarrhea and hematochezia with significantly lower mRNA expressions of COX2, IL-6 and TNFα (P < 0.05) and protein expression of IκBα but without obvious changes in P65 expression in the colon tissue. CONCLUSION: WDSUB1 knockdown can alleviate DSS- induced colitis in mice possibly by inhibiting the NF-κB signaling pathway and decreasing the expression of inflammatory factors in the colon tissues.
OBJECTIVE: To explore the effect of WDSUB1 on dextran sulfate sodium (DSS)-induced inflammatory colon injury in mice and the underlying mechanism. METHODS: Different WDSUB1 siRNA sequences were transfected into mouse fibroblast L929 cells and the optimal sequence was selected by Western blotting. Twelve male C57BL/6 mice were randomized into two groups for injection of siWDSUB1 or siControl via the caudal vein, followed by treatment with 2.5% DSS in drinking water to establish mouse models of DSS- induced colitis (n=6). The expression level of WDSUB1 in the colon tissue of the mice was detected with Western blotting and RT-PCR, the changes in body weight and fecal condition were recorded, and the clinical symptoms of the mice were evaluated. The mRNA expression levels of IL-6, COX-2 and TNF-α and the protein expression of IκBα and P65 in the colon tissues were detected with RT- PCR and Western blotting, respectively. RESULTS: The mRNA and protein expressions of WDSUB1 in the colon tissues were significantly lower in colitis mice with WDSUB1 knock-down than in the control mice. Compared with the control mice, the mice receiving siWDSUB1 injection showed obviously milder weight loss, diarrhea and hematochezia with significantly lower mRNA expressions of COX2, IL-6 and TNFα (P < 0.05) and protein expression of IκBα but without obvious changes in P65 expression in the colon tissue. CONCLUSION: WDSUB1 knockdown can alleviate DSS- induced colitis in mice possibly by inhibiting the NF-κB signaling pathway and decreasing the expression of inflammatory factors in the colon tissues.
Authors: Stefan Wirtz; Vanessa Popp; Markus Kindermann; Katharina Gerlach; Benno Weigmann; Stefan Fichtner-Feigl; Markus F Neurath Journal: Nat Protoc Date: 2017-06-01 Impact factor: 13.491