The Mechanisms of the Anti-Inflammatory and Anti-Apoptotic Effects of Omega-3 Polyunsaturated Fatty Acids during Methotrexate-Induced Intestinal Damage in Cell Line and in a Rat Model

Background: The aim of this study was to examine the anti-inflammatory and anti-apoptotic patterns of omega-3 polyunsaturated fatty acids (n-3 PUFAs) during methotrexate (MTX) induced intestinal damage in cell culture and in a rat model. Methods: Non-treated and treated with MTX HT 29 and HCT116cells were exposed to increasing doses of n-3 PUFAs and cell viability was evaluated using PrestoBlue(R) assay. Male Sprague-Dawley rats were divided into 4 experimental groups: Control rats, CONTR+n-3 PUFA rats that were treated with oral n-3 PUFA, MTX rats were treated with MTX given IP, and MTX+n-3 PUFA rats were treated with oral n-3 PUFA before and following injection of MTX. Intestinal mucosal parameters and mucosal inflammation, enterocyte proliferation and apoptosis, TNF-alpha in mucosal tissue and plasma (ELISA), NF-kappa B, COX-2, TNF-alpha, Fas, FasL, Fadd, Bid, Bax and Bcl-2gene and protein levels were determined 72 h following MTX injection. Results: Exposure of HT 29 and HCT116cells to n-3 PUFA attenuated inhibiting effects of MTX on cell viability. MTX-n-3 PUFA rats demonstrated a lower intestinal injury score and enhanced intestinal repair. A significant decrease in enterocyte apoptosis in MTX+n-3 PUFA rats was accompanied by decreased TNF-alpha, FAS, FasL, FADD and BID mRNA levels. Decreased NF-kappa B, COX-2 and TNF-alpha levels in mucosa was accompanied by a decreased number of IELs and macrophages. Conclusions: n-3 PUFAs inhibit NF-kappa B/COX-2 induced production of pro-inflammatory cytokines and inhibit cell apoptosis mainly by extrinsic pathway in rats with MTX-induced intestinal damage.

Automated summary

This study showed that n-3 PUFAs attenuate the inhibitory effects of MTX on cell viability, reduce intestinal injury scores, enhance intestinal repair, and protect the intestine from damage by inhibiting cell apoptosis, decreasing inflammatory cytokines, and reducing inflammation in rats with MTX-induced intestinal damage.