Gypenoside LVI improves hepatic LDL uptake by decreasing PCSK9 and upregulating LDLR expression.

BACKGROUNDS: Atherosclerotic Cardiovascular Disease (ASCVD) is defined as ischemic or endothelial dysfunction-various inflammatory diseases, which is mainly caused by excessive low-density lipoprotein cholesterol (LDL-C) in circulating blood. Gynostemma pentaphyllum is a traditional Chinese medicine, and total Gypenosides are used for the treatment of hyperlipidemia and to reduce circulating proprotein convertase subtilisin/kexin type 9 (PCSK9) level. However, which gypenoside involved in the modulation of PCSK9 expression is still unknown. PURPOSE: This study aimed to discover effective PCSK9 inhibitors from Gypenosides in accordance with the 2019 ESC/EAS guidelines. METHODS: HPLC was employed to determine major six components of Gypenosides. The inhibitory activity on secreted PCSK9 in HepG2 of six major compounds from Gypenosides were screened by ELISA. The level of low-density lipoprotein (LDL) receptor (LDLR) was determined by Flow cytometry and Immunofluorescence. The expression levels of PCSK9, LDLR and Sterol-regulatory element binding proteins-2 (SREBP-2) were analyzed by qPCR and Western blot. DiI-LDL was added to evaluated LDL uptake into HepG2. RESULTS: The results suggested that the mRNA and protein levels of PCSK9 were down-regulated by Gypenoside LVI and the LDLR degradation in lysosomes system was inhibited, thereby leading to an increasing in LDL uptake into HepG2 cells. Furthermore, Gypenoside LVI decreased PCSK9 expression induced by stains. Altogether, Gypenoside LVI enhances LDL uptake into HepG2 cells by increased LDLR level on cell-surface and suppressed PCSK9 expression. CONCLUSION: This indicates that Gypenoside LVI can be used as a useful supplement for statins in the treatment of hypercholesterolemia. This is firstly reported that monomeric compound of G. pentaphyllum planted in Hunan province has the effect of increasing LDL-C uptake in hepatocytes via inhibiting PCSK9 expression.

Four new dammarane triterpenoid glycosides from the leaves of Cyclocarya paliurus and their SIRT1 activation activities.

Four new C-11 monosaccharide attached dammarane triterpenoid glycosides cypaliurusides SV (1-4), along with nine known dammarane triterpenoid glycosides (5-13) were isolated from a CHCl3-soluble extract of the leaves of Cyclocarya paliurus. All characterized compounds were assayed for their cytotoxicities against HepG2 cells and 10 compounds were evaluated for the agonistic effects on sirtuin1 (SIRT1). The results showed that compounds 1, 5 and 6 were strongly cytotoxic in HepG2 cell line. Two dammarane triterpenoid glycosides 3 and 10 exhibited agonistic activities on SIRT1 with IC50 of 10 µM and 20 µM, respectively.

Triterpenic acids-enriched fraction from Cyclocarya paliurus attenuates insulin resistance and hepatic steatosis via PI3K/Akt/GSK3ß pathway.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) is the most prevalent form of chronic liver diseases. Cyclocarya paliurus (C. paliurus), an edible and medicinal plant in Chinese folk, has been demonstrated to ameliorate diabetes, obesity and lipid metabolism disorders. However, its effects on NAFLD and its potential molecular mechanism have not been clearly expounded. PURPOSE: The present study was designed to explore the therapeutic potential of triterpenic acids-enriched fraction from C. paliurus (CPT), as well as its underlying mechanism in vivo and in vitro models of NAFLD. METHODS: The metabolic effects and possible molecular mechanism of CPT were examined using HepG2 cells and primary hepatocytes (isolated from C57BL/6 J mice) models of fatty liver induced by palmitic acid (PA) and a high fat diet mouse model. RESULTS: In high fat diet-induced C57BL/6 J mice, CPT significantly reduced liver weight index, serum alanine transaminase (ALT), aspartate transaminase (AST), triacylglycerol (TG), total cholesterol (TC) and hepatic TG, TC levels. Moreover, CPT dramatically decreased the contents of blood glucose, insulin, and insulin resistance (HOMA-IR) index. Meanwhile, CPT significantly increased the tyrosine phosphorylation level of IRS and the uptake of 2-deoxyglucose (2DG) in PA-induced HepG2 cells and primary hepatocytes fatty liver models. Furthermore, in PA-induced HepG2 cells and primary hepatocytes, CPT significantly decreased the number of lipid droplets and intracellular TG content. In addition, mechanism investigation showed that CPT increased the phosphorylation of phosphoinositide 3-kinase (PI3K), protein kinase B (Akt) and glycogen synthase-3ß (GSK3ß) in vivo and in vitro models, which were abrogated by PI3K inhibitor LY294002 in vitro models. CONCLUSION: These findings indicate that CPT may exert the therapeutic effects on NAFLD via regulating PI3K/Akt/GSK3ß pathway.

Triterpenic acids-enriched fraction from Cyclocarya paliurus attenuates non-alcoholic fatty liver disease via improving oxidative stress and mitochondrial dysfunction.

The effects of triterpenic acids-enriched fraction from Cyclocarya paliurus (CPT) on nonalcoholic fatty liver disease (NAFLD) were investigated using in vivo and in vitro models. In high fat diet-induced Wister rats, CPT significantly increased superoxide dismutase (SOD) activity and glutathione/oxidized glutathione (GSH/GSSG) ratio, reduced malondialdehyde (MDA) and protein carbonyl (PCO) levels. Moreover, CPT restored mitochondrial membrane potential dysfunction, decreased cytochrome P450 enzyme 2E1 (CYP2E1) activity, improved nuclear factor erythroid 2-related factor 2 (Nrf2) and Nrf2-mediated antioxidant enzyme heme oxygenase1 (HO-1) expression. In free fatty acids-induced HepG2 cells, CPT dramatically decreased ROS content, increased mitochondrial NADH dehydrogenase (Complex I) and mitochondrial cytochrome C oxidase (Complex IV) levels. Furthermore, CPT could upregulate HO-1, quinine oxidoreductase 1 (NQO1) expression, and increase Nrf2 translocation from cytoplasm-to-nucleus. The results indicated CPT could protect mitochondria function and improve oxidative stress by activating Nrf2. Therefore, it can be inferred that CPT may be a potential agent against NAFLD.

Pentacyclic triterpenoids from Cyclocarya paliurus and their antioxidant activities in FFA-induced HepG2 steatosis cells.

Six undescribed pentacyclic triterpenoids including four triterpenoid aglycones, 1ß,2a,3ß,23-tetrahydroxyurs-12-en-28-ursolic acid, 2a,3a,6ß,19α,23-pentahydroxyurs-12-en-28-ursolic acid, 2α,3α,20ß,23-tetrahydroxyurs-12-en-28-ursolic acid and 1ß,2a,3ß,23-tetrahydroxyurs-12,20(30)-dien-28-ursolic acid, and two triterpenoid glucosides, 2a,3a,23-trihydroxy-12,20(30)-dien-28-ursolic acid 28-O-ß-d-glucopyranoside and 1-oxo-3ß,23-dihydroxyolean-12-en-28-oic acid 28-O-ß-d-xylopyranoside, along with 5 known triterpenoids were isolated from a CH3Cl-soluble extract of the leaves of Cyclocarya paliurus. Their structures were established on the basis of chemical and spectroscopic approaches. These compounds were assessed for their antioxidant effects on FFA-induced hepatic steatosis in HepG2 cells. The results revealed that three saponins and two aglycones markedly increased SOD activity and reduced MDA level.

The chloroform extract of Cyclocarya paliurus attenuates high-fat diet induced non-alcoholic hepatic steatosis in Sprague Dawley rats.

BACKGROUND: Hepatic steatosis (HS) is the early stage of nonalcoholic fatty liver disease which is caused by impaired hepatic lipid homeostasis. Cyclocarya paliurus, an herbal tea consumed in China, has been demonstrated to ameliorate abnormal lipid metabolism for the treatment of metabolic diseases. PURPOSE: We aimed to investigate the regulative effect of chloroform extract from Cyclocarya paliurus (ChE) on treatment of HS, as well as key factors involved in hepatic lipid metabolism. STUDY DESIGN: Sprague Dawley rats were fed with high-fat diet (HFD) for 6 weeks to induce HS and treated with or without ChE by gavage for 4 weeks. METHODS: The body weight, relative liver weight and liver fat content were measured. Serum and liver total cholesterol, triglyceride and non-esterified fatty acids, as well as hepatic malonaldehyde levels were accessed by biochemical methods. Serum and liver TNF-α levels were quantified by ELISA kit. Histologic analysis and 1H-MRS study were performed to evaluate HS level. RT-PCR and Western blot were also applied to observe the expression changes of key factors involved in hepatic lipid intake, synthesis, utilization and export. RESULTS: ChE significantly decreased the rats' body weight, serum lipid and TNF-α level. ChE also reduced their relative liver weight, liver fat content, hepatic oxidative products and TNF-α level. Hepatic steatosis in HFD-fed rats was effectively regressed after 2-weeks administration of ChE. Moreover, ChE treatment remarkably reduced HFD-induced high expression level of fatty acid synthesis genes (including sterol-regulatory element-binding protein 1, acetyl-CoA carboxylase 1 and fatty acid synthase). However, it had no effect on mRNA expression of some genes involved in lipid uptake, ß-oxidation and lipid outflow. CONCLUSION: ChE exerted a promising regression effect on HS due to a reduced level of serum non-esterified fatty acids which might lead to a decrease in the amount of lipid taken in by the liver, as well as owing to the inhibition of hepatic lipid de novo synthesis to reduce liver lipid production.

Nauclea officinalis inhibits inflammation in LPS-mediated RAW 264.7 macrophages by suppressing the NF-κB signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Nauclea officinalis has been traditionally used in China for the treatment of fever, pneumonia and enteritidis etc. This study aims to investigate effects of N. officinalis on the inflammatory response as well as the possible molecular mechanism in LPS-stimulated RAW 264.7 murine macrophage cells. MATERIALS AND METHODS: Anti-inflammatory activity of N. officinalis (10, 20, 50, and 100µg/mL) was investigated by using LPS-induced RAW 264.7 macrophages. The NO production was determined by assaying nitrite in culture supernatants with the Griess reagent. The levels of TNF-α, IL-6 and IL-1ß in culture media were measured with ELISA kits. Real time fluorescence quantitative PCR was detected for mRNA expression of iNOS, TNF-α, IL-6 and IL-1ß. Western blot assay was performed to illustrate the inhibitory effects of N. officinalis on phosphorylation of IκB-α and NF-κB p65. RESULTS: Treatment with N. officinalis (10-100µg/mL) dose-dependently inhibited the production as well as mRNA expression of NO, TNF-α, IL-6 and IL-1ß in RAW 264.7 macrophages. Western blot assay suggested that the mechanism of the anti-inflammatory effect was associated with the inhibition of phosphorylation of IκB-α and NF-κB p65. CONCLUSIONS: The results indicated that N. officinalis potentially inhibited the activation of upstream mediator NF-κB signaling pathway via suppressing phosphorylation of IκB-α and NF-κB p65 to inhibit LPS-stimulated inflammation.