Effect of total saponins from Panax japonicus on non-alcoholic steatohepatitis by regulating autophagy / 中国中药杂志

Non-alcoholic steatohepatitis(NASH) was induced by high-sugar and high-fat diet in mice to investigate the intervention effect of total saponins from Panax japonicus(TSPJ) and explore its possible mechanism. Mice were fed with high-sugar and high-fat diet to establish NASH model, and intervened with different doses of TSPJ(15, 45 mg·kg~(-1)). The animals were fed for 26 weeks. The histomorphology and pathological changes of liver tissues were observed by HE staining. The transcriptional expression levels of miR-199 a-5 p, autophagy related gene 5(ATG5) and inflammatory cytokines interleukin-6(IL-6), interleukin-1β(IL-1β) and tumor necrosis factor α(TNF-α) in mouse liver were measured by quantitative Real-time polymerase chain reaction(qRT-PCR). Western blot was used to detect the expression of autophagy-related proteins ATG5, P62/SQSTM1(P62), and microtubule-associated protein light chain 3(LC3)-I/Ⅱ proteins in mouse liver. The expression of P62 protein was detected by immunofluorescence staining. In order to verify the targeting regulation relationship between miR-199 a-5 p and ATG5, miR mimic/inhibitor NC and miR-199 a-5 p mimic/inhibitor were transfected into Hepa 1-6 cells, and the expression of ATG5 mRNA and protein was detected. pMIR-reportor ATG5-3'UTR luciferase reporter gene plasmid was constructed and co-transfected with miR mimic/inhibitor NC and miR-199 a-5 p mimic/inhibitor into Hepa 1-6 cells to detect luciferase activity. In vivo, HE staining in the model group showed typical fatty degeneration and inflammatory infiltration, with increased expression of miR-199 a-5 p and decreased expression of ATG5 mRNA and protein. The expression of autophagy-associated protein P62 increased significantly, the ratio of LC3Ⅱ/Ⅰ decreased, and the transcriptional expression of inflammatory factors increased significantly. After the intervention by TSPJ, the pathological performance of liver tissue was significantly improved, the expression of miR-199 a-5 p decreased and the expression of ATG5 mRNA and protein increased, the expression of autophagy-associated protein P62 decreased significantly, the ratio of LC3Ⅱ/Ⅰ increased, and the transcriptional expression of inflammatory cytokines IL-6, IL-1β and TNF-α decreased significantly. In vitro, it was found that the expression of ATG5 mRNA and protein and luciferase activity decreased significantly in miR-199 a-5 p overexpression cells, while after inhibition of miR-199 a-5 p expression, the expression level of ATG5 mRNA and protein and luciferase activity increased. The results showed that TSPJ can improve NASH in mice fed with high-sugar and high-fat diet, and its mechanism may be related to the regulation of miR-199 a-5 p/ATG5 signal pathway, the regulation of autophagy activity and the improvement of inflammatory response of NASH.

[Experimental study on improvement of alcoholic liver disease by polysaccharides from Balanophora henryi through targeting miR-122a].

The aim of this paper was to investigate the effect of total polysaccharide from Balanophora henryi(TBP) on alcoholic liver disease(ALD) and explore the possible mechanism. C57 BL/6 N mice were randomly divided into 4 groups: pair-feeding group, alcohol-feeding model group, model+TBP group and TBP drug control group. The Gao-binge method was used to prepare the chronic ALD model, and at the same time, 400 mg·kg~(-1) TBP was given for interventional therapy. After feeding for 6 weeks, the serum, liver and colon tissues were collected for detection. As compared with the pair-feeding group, the model group mice showed obvious fatty degeneration and a large number of infiltration of inflammatory cells in the liver, with increased serum ALT and AST levels. After TBP intervention, histopathological changes in liver tissues were significantly improved, with decreased lipid deposition, closer arrangement of hepatocytes, lower expression level of inflammatory factors, and reduced activity of serum ALT and AST, indicating that TBP had a significant improvement effect on ALD. The observation of colonic tissues in mice showed that TBP effectively maintained the integrity of intestinal tissue structure of mice with ALD, enhanced the expression of tight junction protein occludin and reduced miR-122 a expression level. More importantly, TBP significantly reduced serum lipopolysaccharide(LPS) level in model mice. These results indicated that TBP may improve ALD by maintaining and enhancing intestinal barrier function. In vitro experiments showed that TBP significantly inhibited the expression level of miR-122 a in Caco-2 cells exposed to ethanol. Overexpression of miR-122 a in Caco-2 cells induced the inhibition of occludin protein production, and the addition of TBP significantly interfered with the effect. These results suggested that TBP could improve ALD by maintaining the stability of intestinal barrier function and reducing LPS content into the liver, and the mechanism may be partially related to inhibiting miR-122 a expression.

Protective effects of total saponins of Panax japonicas on HepG2 cell apoptosis induced with palmitic acid / 中国中药杂志

This study aimed to investigate the molecular mechanism and protective effect of total saponins of Panax japonicas (TSPJ) on HepG2 cells apoptosis induced by palmitic acid (PA).The HepG2 cells were cultured , and divided into five groups: the control group, the model group, the high-dose group (50 mg·L⁻¹), the middle-dose group (25 mg·L⁻¹) and the low-dose group (12.5 mg·L⁻¹).The cells of the five groups were cultured continuously for 24 hours. The cell viability was measured with MTT. HepG2 cells apoptosis was detected by Hoechest staining and Annexin V-FITC/PI staining. The protein expressions of BCL-2, CHOP and TLR4 were measured with western blotting and flow cytometry analysis. The mRNA expressions of TNF-α, IL-1β, BCL-2, CHOP and GAPDH were measured with RT-PCR. The results suggested that compared with the control group, the number of HepG2 cells of the model group were reduced significantly (<0.01), while the number of apoptotic HepG2 cells were increased. Compared with the model group, the number of HepG2 cells of the high-dose group and the middle-dose group were increased significantly (<0.01), whereas the number of apoptotic HepG2 cells were reduced. Compared with the control group, TNF-α, IL-1β and CHOP mRNA expressions and CHOP and TLR4 protein expressions in the model group were significantly up-regulated (<0.01), while BCL-2 protein and mRNA expressions in the model group were significantly decreased (<0.01). Compared with the model group, TNF-α, IL-1β and CHOP mRNA expressions and CHOP and TLR4 protein expressions in the high-dose group were significantly decreased (<0.01), while BCL-2 protein and mRNA expressions in the high-dose group were significantly up-regulated (<0.01).In conclusion, TSPJ can reduce inflammation and apoptosis induced by palmitic acid, with a certain protective effect on liver cells.