Thymoquinone, a bioactive phytochemical alleviates colon inflammation through Nrf2/Keap1 system: An experimental study using both in vivo and in vitro model of colon inflammation

Introduction Inflammatory bowel diseases (IBD) comprises of Crohn's disease (CD) and ulcerative colitis (UC), which causes inflammation in both small and large intestine. The aetiology of IBD is unclear however, seems to be multifactorial. Therefore, impetus to search for the new therapeutic compounds is warranted. Phytochemicals (a group of secondary plant metabolites) are identified to inhibit a variety of biochemical pathways. One such dietary phytochemical, thymoquinone (TQ) is an active ingredient present in seed oil extract of Nigella sativa exhibits various pharmacological properties including anti‐oxidant, anti‐inflammatory properties however, the primary molecular targets remain to be clarified in colon inflammation. Aim Therefore, the main aim of this study is to investigate the molecular targets of TQ responsible for its anti‐inflammatory property in the experimental model of colon inflammation using both in vivo and in vitro studies. Methodology C57BL/6J black mice were administered with 2% dextran sodium sulfate (DSS) in drinking water for 7days to induce colitis. The treatment group received TQ using oral gavage at concentrations of 20 & 40mg/kg body wt and compared with control and DSS along with a positive control through clinically used drug (sulfasalazine) group. The disease activity index (DAI) was established throughout the treatment period and colon length, changes in myeloperoxidase (MPO) and histology was performed at the end of the experiment. Changes in myeloperoxidase (MPO) activity and pro‐inflammatory cytokines (IL‐1β, TL‐6 and TNF‐α) protein level were measured using ELISA and mRNA using real time PCR. Cox‐2, iNOS, Nrf2 & KEAP1 protein and mRNA expression were carried out. Human colon carcinoma cell line, HT‐29 was used for in vitro studies. HT‐29 cells were cultured in 10% FBS containing DMEM media and were stimulated using LPS (1μg/ml) to mimic inflammation and were treated with various concentrations of TQ. Proinflammatory cytokine mRNA level were measured using real time PCR. Cox‐2, iNOS, Nrf2 & KEAP1 protein and mRNA expression were carried out. Results TQ treatment significantly ( p<0.01 ) decreased DAI, MPO level and restored colon length dose dependently in DSS group. Histological scoring for colonic crypt damage and inflammation was significantly ( p<0.001 ) improved upon TQ treatment. TQ also inhibited the pro‐inflammatory cytokines (IL‐1β, IL‐6 and TNF‐α) response significantly ( p<0.01 ) both at protein and mRNA level. TQ also significantly inhibited COX‐2, iNOS, Nrf2 & KEAP1 protein and mRNA expression. TQ co‐treatment with LPS significantly ( p<0.01 ) decreased proinflammatory response in HT‐29 cells along with decrease in COX‐2, iNOS, Nrf2 & KEAP1 at both protein and mRNA level. Conclusion TQ appears to mitigate aberrant inflammatory response observed both in vivo and in vitro model of colon inflammation. This appears to be mediating through Nrf2/Keap1 system. Support or Funding Information This research work was supported by SURE+ 2018 grant of UAE University This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .