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Aim To study the effects of naringenin on MCD diet-induced liver fibrosis and its related mechanisms.Methods LX2 cells were incubated with TGF-β1 for 24 h to establish the in vitro fibrosis model.LX2 cells were treated with NGN at the same time.Male C57BL/6 mice were fed with MCD diet for six weeks to induce liver fibrosis.100 mg·kg-1·d-1 NGN was administered by gavage simultaneously.The protein expressions of α-SMA, col1, TGF-β1, p-smad2 and p-smad3 were evaluated by Western blot.The mRNA expressions of α-SMA, col1 and col3 were detected by qRT-PCR.The degree of liver fibrosis was evaluated by Sirius red staining.Results Both in in vivo and in vitro experiments, compared with model group, the mRNA levels of α-SMA, col1 and col3 and protein levels of α-SMA, TGF-β1, p-smad2 and p-smad3 significantly decreased in NGN treatment group.The results of HE staining and Sirius red staining also indicated that NGN significantly decreased liver fibrosis induced by MCD diet.Conclusions Naringin can significantly inhibit liver fibrosis induced by MCD diet, which may be related to TGF-β1/Smad pathway.
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OBJECTIVE：To study the effects of matrine on proliferation and collagen synthesis of rat hepatic stellate cells CFSC-8B activated by acetaldehyde ，and to investigate its possible mechanism. METHODS ：CFSC-8B cells cultured in vitro were divided into blank control group ，model group ，positive control group （2.5 μmol/L colchicine）and matrine low ，medium and high concentration groups （30，60，120 μmol/L）. Except for blank control group ，other groups were activated with 200 μmol/L acetaldehyde for 24 h；medicine groups were intervened with relevant medicine for 24 h（blank control group and model group were intervened with equal volume blank medium ）. Survival rate of cell was detected by CCK- 8 assay. C ells were divided into blank control group ，model group ，positive control group （2.5 μmol/L colchicine），matrine medium and high concentration groups （60，120 μmol/L），then activated and treated with same method. Hydroxyprolin （Hyp）content in cell culture solution was tested by enzyme digestion. The contents of Col- Ⅰ and Col- Ⅲ in cell culture solution were determined by ELISA. mRNA expressionss of α-SMA，TGF-β1，TβR-І，TβR-Ⅱ，Smad3，Smad4 and Smad 7 in cells were detected by RT-PCR. The protein expressions of α-SMA，TGF-β1，TβR-Ⅰ，TβR- Ⅱ，Smad3，Smad4 and Samd 7 in cells were detected by Western blotting. RESULTS ：Compared with blank control group ，survival rate of cells in model group was increased significantly （P＜0.05）；the contents of Hyp ，Col-Ⅰ and Col- Ⅲ in cell culture solution ，mRNA and its protein expressions of α-SMA，TGF-β1，TβR-Ⅰ，TβR-Ⅱ，Smad3，Smad4 in cells were increased significantly in model group （P＜0.05），while the mRNA and protein expression of Smad 7 was decreased significantly（P＜0.05）. Compared with model group ，survival rate of cells ，the contents of Hyp ，Col-Ⅰ and Col- Ⅲ in cell culture solution，the mRNA and protein expressions of α-SMA and Smad 4 were decreased significantly in positive control group and matrine medium and high concentration groups （P＜0.05）， while the mRNA and protein expression of Smad 7 was WF-0099） increased significantly （P＜0.05）；the mRNA and proteinexpressions of TGF-β1，TβR-Ⅰ，TβR-Ⅱ and Smad 3 were decreased significantly in positive control group and matrine high concentration group （P＜0.05）. Compared with matrine medium concentration group ，all above indexes were improved significantly in matrine high concentration group （P＜0.05）. CONCLUSIONS ：Matrine can suppress the proliferation and collagen synthesis of CFSC- 8B cells activated by acetaldehyde ，with a centain concentrlation dependence ，the mechanism of which may be associated with regulating the conduction of TGFβ/Smad signal pathway.
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Objective:To investigate the intervention effect of panax notoginseng saponins (PNS) on epithelial-mesenchymal transition (EMT) of rat renal tubular epithelial cells (NRK-52E) induced by transforming growth factor-β1(TGF-β1), and analyze the mechanism based on the silent information regulation 2 homolog 1(SIRT1)/TGF-β1/Smad signaling pathway. Method:NRK-52E were cultured in DMEM medium with 10% fetal bovine serum, and divided into normal control group, TGF-β1 group (5 μg·L-1), resveratrol (RSV) group (50 mg·L-1), EX527 group (10 μmol·L-1), Panax notoginseng saponins (PNS) group (100 mg·L-1), and EX527+ PNS group (10 μmol·L-1+100 mg·L-1). Then cells were collected after drug intervention for 48 h. The expressions of α-SMA,E-cadherin,SIRT1,TGF-β1,Smad3,Smad4 mRNA in each group were detected by Real-time PCR. The protein expressions of α-SMA, E-cadherin,SIRT1 and TGF-β1 were detected by Western blot. Result:Compared with normal group, mRNA and protein expressions of α-SMA increased obviously(PPβ1 group. Compared with TGF-β1 group, mRNA and protein expressions of α-SMA decreased significantly(PPPβ1,Smad3,and Smad4 decreased(PConclusion:PNS can prevent the occurrence of EMT of renal tubular epithelial cells induced by TGF-β1, and the mechanism may be related to active SIRT1 to inhibit TGF-β1/Smad pathway.
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Objective To investigate the roles of ubiquitin-conjugating enzyme(UBC9)in HSCs activa-tion and liver fibrosis. Methods Western blot was used to analyze the expression of UBC9 under the stimulus of different concentrations of TGF-β1. The effective shRNA-targeting UBC9 gene was synthesized and HCSs were in-stantaneously transfected using lipofectamine method. Non-specific shRNA-transfected group cells and shRNA-tar-geting UBC9-transfected group cells were set up. The mRNA and protein levels of UBC9 were determined with Quantitative Real-Time PCR and Western blot. Western blot also used to examine the expression level of collagenⅠ,α-SMA and P-smad3 after transfection of UBC9 shRNA into HCSs and CCK-8 assay was used to detect cell proliferative capacity after transfection. Results UBC9 expression was significantly up-regulated in TGF-β1-treat-ed HSCs. Knockdown of UBC9 significantly inhibited TGF-β1-induced HSCs proliferation,as well as decreased the expression levels of a-SMA and collagen I. Furthermore ,knockdown of UBC9 attenuated the phosphorylation of Smad3 in the presence of TGF-β1. Conclusions UBC9 may function as a novel regulator to modulate HSC activa-tion,potentially by inhibiting the TGF-β1/Smad3 signaling pathway,which reveals novel mechanistic insights into the anti-fibrotic effect of UBC9.