Effect Of Zinc ?2 Glycoprotein On JNK Inflammatory Signaling Pathway In HepG2 Intervention By LP

Objective: To observe the effects of over-expression of Zinc alpha2 glycoprotein on TNF-? and IL-1? gene expression in Hep G2 cells and explore the effects of ZAG ?FXR and Adiponectin gene expression in Hep G2 and Hepa1-6 cells by LPS intervention at different time.Methods:1. Plasmid vector p IRES2–h ZAG was transfected into Hep G2 cells for 48 hours by Lipofectamine 2000-mediated gene transaction at concentrations of 1,2,3,4,5?g/ml.The protein expression of ZAG was detected by Western blot.2.Plasmid vector p IRES2–h ZAG and empty vector were transfected into Hep G2 cells by Lipofectamine 2000-mediated gene transaction for 48 hours.Cells were collected to detect the protein expression of TNF-? and IL-1? gene in Hep G2 cells.3.Hep G2 and Hepa1-6 cells were treated with LPS(1?g/ml) intervention at different time, such as 0h,1/2h,1h,2h,6h,12 h,24h,48 h to detect the protein level of ZAG?FXR and Adiponectin gene by Western blot.Results:1.Compared with the control group,the protein expression of ZAG was significantly increased in p IRES2-h ZAG transfection group(4?g/ml).2.Compared with p IRES2-GFP group,the protein level of inflammatory factor TNF-? was decreased in p IRES2–h ZAG transfection group(P<0.05),Moreover, the protein level of inflammatory factor IL-1? was decreased in p IRES2–h ZAG transfection group(P<0.05).3.The protein level of ZAG was increased at 1h,and significantly decreased at 12 h,24h and 48 h in Hep G2 cells induced by LPS(P<0.01), the protein level of FXR was decreased at 12 h,24h and 48 h in Hep G2 cells induced by LPS(P<0.05), the protein level of Adiponectin was decreased at 6h,12 h,24h and 48 h in Hep G2 cells induced by LPS(P<0.05). Moreover, The protein level of ZAG was increased at 1h,and decreased at 6h,12 h,24h and 48 h in Hepa1-6 cells induced by LPS(P<0.05), the protein level of FXR was decreased at 12 h,24h and 48 h in Hepa1-6 cells induced by LPS(P<0.05), the protein level of Adiponectin was decreased at 6h,12 h,24h and 48 h in Hepa1-6 cells induced by LPS(P<0.05).Conclusion: 1. ZAG could reduce the expression of inflammatory factors TNF-? and IL-1? in Hep G2 cells.2. The protein expression of ZAG ?FXR and Adiponectin were decreased at 12 h in Hep G2 cells and Hepa1-6 cells by LPS-induced.Objective:To explore the mechanism of JNK signaling pathway mediate the effects of ZAG on inflammation in Hep G2 cells induced by LPS.Methods:1. Hep G2 cells and Hepa1-6 cells were treated with LPS(1?g/ml) for 12 hours in control group?p IRES2–GFP transfection group and p IRES2–h ZAG transfection group,These cells were collected after 12 hours to detect the protein level of ZAG?FXR and Adiponectin gene by Western blot.2. Plasmid vector p IRES2–h ZAG and empty vector were transfected into Hep G2 cells for 48 hours by Lipofectamine 2000-mediated gene transaction,Cells were treated with lipopolysaccharide(1?g/ml) and JNK inhibitor SP600125(10?M).The protein level of JNK?P-JNK?C-jun?P-cjun were detected by Western blot.Results: 1.Compared with the control group,the protein expression of ZAG was significantly decreased in LPS group and LPS+p IRES2–GFP group(P<0.01), compared with p IRES2–GFP group,the protein level of ZAG was increased in p IRES2–h ZAG group(P<0.05),Compared with the control group,the protein level of FXR and Adiponectin gene were decreased in LPS group and LPS+p IRES2–GFP group(P<0.05).2.Compared with p IRES2–GFP group,the protein expression of JNK had no significant change in p IRES2–h ZAG group(P>0.05), compared with control group,the protein level of P-JNK was increased by LPS(P<0.05),compared with the LPS group,the protein level of P-JNK was decreased in LPS+SP600125 group(P<0.05),Moreover,compared with p IRES2–GFP group,the protein expression of P-JNK was decreased by LPS+SP600125 in p IRES2–h ZAG group(P<0.05).3.Compared with control group,the protein level of C-jun and P-cjun were increased in p IRES2–GFP group by LPS(P<0.05),compared with the LPS group,the protein level of C-jun and P-cjun were decreased in LPS+SP600125 group(P<0.05),Moreover,compared with p IRES2–GFP group,the protein expression of C-jun and P-cjun were decreased by LPS+SP600125 in p IRES2–h ZAG group(P<0.05).Conclusion: 1.Over-expression of ZAG could reduce the protein level of P-JNK?C-jun and P-cjun gene,Moreover, ZAG could promote the effects of JNK inhibitor SP600125 to decrease the expression of P-JNK?C-jun and P-cjun gene.