Study on the mechanism of Yishen tongluo formula improving abnormal lipid metabolism based on SREBPs pathway / 中国药房

OBJECTIVE To explore the mechanism of Yishen tongluo formula （YSTLF） in improving abnormal lipid metabolism based on the sterol regulatory element binding proteins （SREBPs） pathway. METHODS Using C57BLKS/J （db/db） mice as model and C57BLKS/J （db/m） mice as normal control， the mechanism of 1， 2.5 and 5 g/kg YSTLF improving abnormal lipid metabolism of db/db mice was investigated by determining the liver coefficient， the contents of serum total cholesterol （TC）， triglyceride （TG）， low-density lipoprotein （LDL） and high-density lipoprotein （HDL）， observing steatosis and lipid accumulation in liver tissue of mice， detecting the protein expressions of SREBP-1 and SREBP-2 as well as mRNA transcription levels of Srebp- 1c， Srebp-2 and their downstream lipid metabolism-related target genes （Fasn， Acc1， Scd5， Fads1， Hmgcr， Dhcr24， Insig-1， Fdps） in liver tissue of mice. Using low-fat cultured human liver cancer cell HepG2 as an in vitro cell model for abnormal lipid metabolism， and 25-HC （SREBPs inhibitor， 10 μmol/L） as the control， the effects of 125， 250 and 500 μg/mL YSTLF on protein expressions of SREBP-1 and SREBP-2 as well as mRNA transcription of SREBP-1c， SREBP-2 and their downstream lipid metabolism-related target genes were investigated to verify the mechanism in vitro. RESULTS 1， 2.5， 5 g/kg YSTLF significantly reduced the levels of TC， TG and LDL， the percentage of lipid droplet-positive region in liver tissue and liver coefficient， significantly down-regulated protein expressions of Pre-SREBP-1， n-SREBP-1， Pre-SREBP-2 and n-SREBP-2， and mRNA transcription of Srebp-1c， Srebp-2 and their downstream target genes in liver tissue， while significantly increased HDL level， with statistical significance （P＜0.05 or P＜0.01）. In the cell experiment in vitro， the expressions of the above-mentioned proteins and genes in the cells treated with YSTLF at 125， 250 and 500 μg/mL for 24 hours were consistent with those in the animal experiment； there was no significant difference in the expressions of the above-mentioned proteins and genes between inhibitor control group and 250， 500 μg/mL YSTLF groups （P＞0.05）. CONCLUSIONS YSTLF can regulate the expression of transcription factor SREBPs， so as to inhibit the high expression of fatty acid and cholesterol synthesis-related genes， promote the degradation of TC and TG， improve the abnormality of lipid metabolism and inhibit lipid accumulation， thus playing the role of lipid-lowering.

Inhibition of ginsenoside Rb 1 on the epithelial-mesenchymal transition of renal tubular epithelial cells by regulating TGF-β1/Smad3 signaling pathway / 中国药房

OBJECTIVE To study the effects of ginsenoside Rb 1（G-Rb1）on epithelial-mesenchymal transition （EMT）of renal tubular epithelial cells and its potential mechanism. METHODS The growth factor β1（TGF-β1）10 ng/mL was used to induce EMT of human renal tubular epithelial cells HK- 2. The morphological changes of HK- 2 cells were observed after treated with 10， 20，30 μmol/L G-Rb1 for 48 h. The transcriptional activities of biovector SBE in human embryonic kidney cell HEK 293 were determined after 24 h treatment with 1.0，2.5，5.0，10，20，30 μmol/L G-Rb1. Effects of above concentration of G-Rb 1 on the viability of HK- 2 cells were determined after 24 h of treatment. mRNA expressions of α-smooth muscle actin （α-SMA），collagen Ⅰ （COL-Ⅰ）and fibronectin （FN）in HK- 2 cells were detected after treated with 10，20，30 μmol/L G-Rb1 for 24 h. The expressions of α-SMA，Smad3，p-Smad3，COL-Ⅰ，FN and E-cadherin were detected after treated with 10，20，30 μmol/L G-Rb1 for 24 h. RESULTS G-Rb1 of 10-30 μmol/L significantly inhibited TGF-β1-induced EMT in HK- 2 cells and the increase of transcriptional activities of biovector SBE induced by TGF-β1（P＜0.05），but had no effects on relative activities of HK- 2 cells（P＞0.05）. The protein and mRNA expressions of α-SMA，COL-Ⅰ and FN ， the protein expressions of Smad 3 and p-Smad 3 were significantly up-regulated induced by TGF-β1（P＜0.05），while the protein expression of E-cadherin was significantly down- regulated（P＜0.05）；G-Rb1 could effectively reverse aboveprotein or mRNA expressions. CONCLUSIONS G-Rb1 can protect renal tubular epithelial cells from EMT induced byxiezhishen TGF-β1 to a certain extent ，which may be related to inhibiting the activation of TGF-β1/Smad3 signaling pathway.

Mechanism of Zexie Decoction in improvement of nonalcoholic fatty liver disease based on LKB1/AMPK/PGC-1α pathway / 中国中药杂志

The present study investigated the pharmaceutical effect and underlying mechanism of Zexie Decoction(ZXD) on nonalcoholic fatty liver disease(NAFLD) in vitro and in vivo via the LKB1/AMPK/PGC-1α pathway based on palmitic acid(PA)-induced lipid accumulation model and high-fat diet(HFD)-induced NAFLD model in mice. As revealed by the MTT assay, ZXD had no effect on HepG2 activity, but dose-dependently down-regulated alanine aminotransferase(ALT) and aspartate aminotransferase(AST) in the liver cell medium induced by PA, and decreased the plasma levels of ALT and AST, and total cholesterol(TC) and triglyceride(TG) levels in the liver. Nile red staining showed PA-induced intracellular lipid accumulation, significantly increased lipid accumulation of hepatocytes induced by PA, suggesting that the lipid accumulation model in vitro was properly induced. ZXD could effectively improve the lipid accumulation of hepatocytes induced by PA. Oil red O staining also demonstrated that ZXD improved the lipid accumulation in the liver of HFD mice. JC-1 staining for mitochondrial membrane potential indicated that ZXD effectively reversed the decrease in mitochondrial membrane potential caused by hepatocyte injury induced by PA, activated PGC-1α, and up-regulated the expression of its target genes, such as ACADS, CPT-1α, CPT-1β, UCP-1, ACSL-1, and NRF-1. In addition, as revealed by the Western blot and immunohistochemistry, ZXD up-regulated the protein expression levels of LKB1, p-AMPK, p-ACC, and PGC-1α in vivo and in vitro. In conclusion, ZXD can improve NAFLD and its mechanism may be related to the regulation of the LKB1/AMPK/PGC-1α pathway.

Study on the Effect and Mechanism of Ganoderic Acid C 2 on Lipid Metabolism of Hepatocytes by Regulating S6K/SREBPs Signaling Pathway / 中国药房

OBJECTIVE：To stu dy in vitro lipid-lowering effect of ganoderic acid C 2（GAC2），and to investigate its potential mechanism on the basis of S 6K/SREBPs signaling pathway. METHODS ：Using human liver cells HL- 7702 as objects ，MTT assay was used to test relative cell viability after treated with low ，medium and high doses （5，10，20 μmol/L，hereinafter）of GAC 2. Using lovastatin as positive control ，ELISA method was used to detect the contents of TC and TG in cells after treated with low ， medium and high doses of GAC 2. Nile red staining was used to observe the accumulation of lipids in cells. After transfected SREBPs report gene plasmid ，using 25-HC as positive control ，relative viability of SREBPs luciferase in cells were determined by luciferase assay after treated with low ，medium and high doses of GAC 2. Using 25-HC as positive control ，real-time fluorescent quantitative PCR was used to measure the mRNA expression of SREBPs and their downstream genes in cells after treated with medium and high doses of GAC 2. Using SREBPs inhibitor （25-HC）and S 6K inhibitor （rapamycin）as control ，Western blotting assay was adopted to determine the expression of SREBP- 1 and SREBP- 2（in the case of n-SREBPs ），relative expression ratio of phosphorylated S 6K to S 6K（p-S6K/S6K ratio ）. AutoDock 4.0 and other softwares were used for molecular docking of S 6K and GAC2. RESULTS ：There was no significant effect of low ， 0.05）. Compared with blank control group ，the content of TC qq.com in lovastatin group and GAC 2 high-dose group as well as thecontent of TG in lovastatin group ， GAC2 medium- and 床应用。电话：0371-65962746。E-mail：whui3697@126.com high-dose groups were decreased significantly （P＜0.05 or P＜ 0.01）；the number of lipid droplets in the cells of all medication groups decreased. Compared with blank control group ，relative viability of SREBPs luciferase in 25-HC group ，GAC2 low-，medium- and high-dose groups were decreased significantly ；mRNA expression of HMGCS1，MVK，SCD，HMGCR gene in 25-HC group and GAC 2 medium-，high-dose groups ，mRNA expression of DHCR7 gene in 25-HC group ，mRNA expression of SREBP-2 gene in GAC- 2 high-dose group as well as mRNA expression of DHCR24 and MSMO2 gene in 25-HC group and GAC 2 high-dose group were all decreased significantly ；relative protein expression of n-SREBP- 1 in 25-HC group ，GAC2 low-，medium- and high-dose groups ，relative protein expression of n-SREBP- 2 in 25-HC group and GAC 2 high-dose group as well as p-S 6K/S6K ratio in rapamycin group and GAC 2 groups were decreased significantly （P＜0.05 or P＜0.01）. The molecular docking results showed that GAC 2 could bound to amino acid residues Arg 335，Arg330 and Ala332 of S 6K via hydrogen bond. CONCLUSIONS ：GAC2 can reduce the lipid level of HL- 7702 cells，which may be associated with inhibiting the expression of S 6K/SREBPs signaling pathway.