Protective effect of Gegen Qinlian Decoction on inflammation injured Caco-2 cells via the p38MAPK/MLCK pathway
OBJECTIVE To investigate the effect of Gegen Qinlian Decoction(GQD)on Tight Junction(TJ)protein in Lipopolysaccharides(LPS)-induced inflammatory model constructed from human colon epithelial cancer cell line Caco-2,and to explore its mechanisms involved in the treatment of ulcerative colitis(UC).METHODS Blank and drug-containing sera were extracted from SD male rats.Caco-2 cells were cultured and categorized into five groups:Control,LPS,GQD-L,GQD-M and GQD-H;except for the control one,other groups were cultured by 1 μg/ml LPS to establish the inflammation model.The cellular proliferation rates were detected by CCK-8 assay,the mRNA and protein expressions of Occludin,Claudin-1,interleukin(IL)-1β,IL-6,IL-10,tumor necrosis fac-tor-α(TNF-α),p-p38 mitogen-activated protein kinase(p-p38MAPK)/p38MAPK,myosin light-chain kinase(MLCK)and p-myosin light chain(MLC)were detected by quantitative real time polymerase chain reaction(RT-qPCR)and Western blot(WB)respectively,the densities of Occludin,Claudin-1,p-p38MAPK,MLCK and MLC were detected by immunofluorescence staining.RESULTS Compared with the control group,cell prolifera-tion was significantly inhibited in the LPS group,with low levels of Occludin and Claudin-1,and high levels of IL-1β,IL-6,TNF-α,low level of IL-10,with high levels of p-p38MAPK,p38MAPK,p-p38MAPK/p38MAPK,MLCK,MLC2 and p-MLC(P＜0.05).Compared with the LPS group,all GQD dose groups,especially GQD-M and GQD-H,showed a significant promotion of cell proliferation,with high levels of Occludin and Claudin-1,and low levels of IL-1β,IL-6,TNF-α,high level of IL-10,with low levels of p-p38MAPK,p38MAPK,p-p38MAPK/p38MAPK,MLCK,MLC2 and p-MLC(P＜0.05).CONCLUSION Gegen Qinlian Decoction may exert therapeutic effects on UC by inhibiting the overactivation of the p38MAPK/MLCK pathway,attenuating the inflammatory re-sponse,reversing intestinal epithelial TJ injury and protecting the intestinal mucosal barrier.
Gegen Qinlian DecoctionCaco-2 cellsUcerative Colitisp38 mitogen-activated protein kinaseMyo-sin light-chain kinaseTight junction proteinInflammatory factors
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