Label-free proteomic analysis reveals the hepatoprotective mechanism of gypenosides in liver injury rats.

Chronic liver disease, a long-term condition resulting from various causes such as alcohol abuse, metabolic disorders, and viral hepatitis, is becoming a significant global health challenge. Gypenosides (GPs), derived from the traditional Chinese medicine Gynostemma pentaphyllum (Thunb.) Makino, exhibited hepatoprotective properties in recent years, yet the precise therapeutic mechanism remains unclear. In this study, label-free and parallel reaction monitoring (PRM) proteomics were used to elucidate the hepatoprotective mechanism of GPs in liver injury rats. Through label-free proteomics, we identified 2104 differentially expressed proteins (DEPs) associated with liver injury, along with 1974 DEPs related to the effects of GPs. Bioinformatics analysis revealed that GPs primarily restored metabolic processes involving valine, leucine, and isoleucine degradation, as well as propanoate and butanoate metabolism, and steroid hormone biosynthesis during liver injury. Subsequently, overlapping the two groups of DEPs identified 1508 proteins reversed following GPs treatment, with key targets further validated by PRM. Eight target proteins were identified for GPs treatment of liver injury, including Lgals3, Psat1, Phgdh, Cyp3a9, Cyp2c11, Cyp4a2, Glul, and Ces1d. These findings not only elucidated the hepatoprotective mechanism of GPs, but may also serve as potential therapeutic targets of chronic liver disease.

Gypenosides exert cardioprotective effects by promoting mitophagy and activating PI3K/Akt/GSK-3ß/Mcl-1 signaling.

Background: Gynostemma pentaphyllum (Thunb.) Makino, a well-known edible and medicinal plant, has anti-aging properties and is used to treataging-associated conditions such as diabetes, metabolic syndrome, and cardiovascular diseases. Gypenosides (GYPs) are the primary constituents of G. pentaphyllum. Increasing evidence indicates that GYPs are effective at preserving mitochondrial homeostasis and preventing heart failure (HF). This study aimed to uncover the cardioprotective mechanisms of GYPs related to mitochondrial regulation. Methods: The bioactive components in GYPs and the potential targets in treating HF were obtained and screened using the network pharmacology approach, followed by drug-disease target prediction and enrichment analyses. The pharmacological effects of GYPs in cardioprotection, mitochondrial function, mitochondrial quality control, and underlying mechanisms were further investigated in Doxorubicin (Dox)-stimulated H9c2 cardiomyocytes. Results: A total of 88 bioactive compounds of GYPs and their respective 71 drug-disease targets were identified. The hub targets covered MAPK, EGFR, PI3KCA, and Mcl-1. Enrichment analysis revealed that the pathways primarily contained PI3K/Akt, MAPK, and FoxO signalings, as well as calcium regulation, protein phosphorylation, apoptosis, and mitophagy process. In Dox-stimulated H9c2 rat cardiomyocytes, pretreatment with GYPs increased cell viability, enhanced cellular ATP content, restored basal oxygen consumption rate (OCR), and improved mitochondrial membrane potential (MMP). Furthermore, GYPs improved PINK1/parkin-mediated mitophagy without influencing mitochondrial fission/fusion proteins and the autophagic LC3 levels. Mechanistically, the phosphorylation of PI3K, Akt, GSK-3ß, and the protein level of Mcl-1 was upregulated by GYP treatment. Conclusion: Our findings reveal that GYPs exert cardioprotective effects by rescuing the defective mitophagy, and PI3K/Akt/GSK-3ß/Mcl-1 signaling is potentially involved in this process.

The potential molecular mechanism underlying gypenoside amelioration of atherosclerosis in ApoE-/- mice: A multi-omics investigation.

Gypenosides (Gyp) are bioactive components of Gynostemma pentaphyllum that have a variety of pharmacological properties. Extracts of G. pentaphyllum have been found to be effective in the reduction of blood sugar and lipids and prevention of atherosclerosis. Here, the functions of Gyp and the mechanisms underlying their effects on atherosclerosis were investigated. Mice were allocated to three groups, namely, the control (C57BL/6), atherosclerosis model (ApoE-/- mice with high-fat diet), and Gyp-treated groups. Differentially expressed mRNAs, miRNAs, circRNA, and differential metabolites among the groups were analyzed. The results showed that "Fatty acid metabolism", "Fatty acid elongation", "Cytokine-cytokine receptor interaction", and "PI3K-Akt signaling pathway", amongst others, were involved in treatment process. Differentially expressed genes, including Fabp1, Apoe, FADS1, ADH1, SYNPO2, and Lmod1were also identified. Mmu-miR-30a and mmu-miR-30e showed reduced expression in atherosclerosis models but were increased following Gyp treatment, suggesting involvement in the effects of Gyp. In addition, chr5:150604177-150608440 were found to interact with mmu-miR-30a and mmu-miR-30e to regulate their abundance. In terms of metabolomics, Gyp may regulate biological processes involving PGD2 and PGJ2, potentially alleviating atherosclerosis. In conclusion, Gyp appeared to have complex effects on atherosclerosis, most of which were positive. These results support the use of Gyp in the treatment of atherosclerosis.

[Comparison on anti-inflammatory activity of Gynostemma pentaphyllum processed with different methods].

The aim of this study is to investigate the effects of Gynostemma pentaphyllum dried with two different methods(air drying and heating) on inflammation in acute lung injury(ALI) mice in vivo and in vitro. Lipopolysaccharide(LPS) was sprayed into the airway of wild type C57BL/6J male mice to establish the model, and the drug was injected into the tail vein 24 h after modeling. Lung function, lung tissue wet/dry weight(W/D) ratio, the total protein concentration, interleukin 6(IL-6), IL-1ß, and tumor necrosis factor-α(TNF-α) in the bronchoalveolar lavage fluid(BALF), and pathological changes of the lung tissue were used to evaluate the effects of different gypenosides on ALI mice. The results showed that total gypenosides(YGGPs) and the gypenosides substituted with one or two glycosyl(GPs_(1-2)) in the air-dried sample improved the lung function, significantly lowered the levels of IL-1ß and TNF-α in BALF, and alleviated the lung inflammation of ALI mice. Moreover, GPs_(1-2) had a more significant effect on inhibiting NO release in RAW264.7 cells. This study showed that different drying methods affected the anti-inflammatory activity of G. pentaphyllum, and the rare saponins in the air-dried sample without heating had better anti-inflammatory activity.

Multi­omics­based analysis of the regulatory mechanism of gypenosides on bile acids in hypercholesterolemic mice.

Gynostemma pentaphyllum is a traditional medicine used by ethnic minorities in southwest China and gypenosides are currently recognized as essential components of the pharmacological substances of Gynostemma pentaphyllum, which are effective in regulating metabolic syndrome, especially in improving hepatic metabolic disorders. The present study randomly divided C57BL/6J male mice into the normal diet control group (ND), high-fat diet modeling group (HFD) and gypenosides group (GP). Liquid chromatography-mass spectrometry (UPLC-MS) was applied to quantify bile acids in the liver, bile and serum of mice in ND, HFD and GP groups. Liver proteins were extracted for trypsin hydrolysis and analyzed quantitatively using UPLC-MS + MS/MS (timsTOF Pro 2). Total mouse liver RNA was extracted from ND, HFD and GP groups respectively, cDNA sequencing libraries constructed and sequenced using BGISEQ-500 sequencing platform. The expression of key genes Fxr, Shp, Cyp7a1, Cyp8b1, and Abab11 was detected by RT-qPCR. The results showed that gypenosides accelerated free bile acid synthesis by promoting the expression of bile acid synthase CYP7A1 and CYP8B1 genes and proteins and accelerating the secretion of conjugated bile acids from the liver to the bile ducts. GP inhibited the bile acid transporters solute carrier organic anion transporter family member (SLCO) 1A1 and SLCO1A4, reducing the reabsorption of free bile acids and accelerating the excretion of free bile acids from the blood to the kidneys. It also promoted the metabolic enzyme CYP3A11, which accelerated the metabolism and clearance of bile acids, thus maintaining the balance of the bile acid internal environment.

Gypenosides suppress hepatocellular carcinoma cells by blocking cholesterol biosynthesis through inhibition of MVA pathway enzyme HMGCS1.

Hepatocellular carcinoma (HCC) is one of the most common malignant tumors with high morbidity and mortality. Targeting abnormal cholesterol metabolism is a potential therapeutic direction. Therefore, more natural drugs targeting cholesterol in HCC need to be developed. Gypenosides (Gyp), the major constituent of Gynostemma pentaphyllum, has been demonstrated to have pharmacological properties on anti-cancer, anti-obesity, and hepatoprotective. We investigated whether Gyp, isolated and purified by our lab, could inhibit HCC progression by inhibiting cholesterol synthesis. The present research showed that Gyp inhibited proliferation and migration, and induced apoptosis in Huh-7 and Hep3B cells. Metabolomics, transcriptomics, and target prediction all suggested that lipid metabolism and cholesterol biosynthesis were the mechanisms of Gyp. Gyp could limit the production of cholesterol and target HMGCS1, the cholesterol synthesis-related protein. Downregulation of HMGCS1 could suppress the progression and abnormal cholesterol metabolism of HCC. In terms of mechanism, Gyp suppressed mevalonate (MVA) pathway mediated cholesterol synthesis by inhibiting HMGCS1 transcription factor SREBP2. And the high expression of HMGCS1 in HCC human specimens was correlated with poor clinical prognosis. The data suggested that Gyp could be a promising cholesterol-lowering drug for the prevention and treatment of HCC. And targeting SREBP2-HMGCS1 axis in MVA pathway might be an effective HCC therapeutic strategy.

Response mechanism of growth and gypenosides content for Gynostemma longipes cultivated at two altitude habitats to fine root morphological characteristics.

Introduction: Fine roots are the critical functional organs of plants to absorb water and nutrients from the soil environment, while the relation between fine root morphological characteristics and yield & quality has received less attention for medicinal plants. Methods: Therefore, we investigated the relationship between fine root morphological characteristics and biomass & gypenosides content. We explored the primary environmental drivers of fine root indicators for Gynostemma longipes from three provenances cultivated at two altitude habitats. Results: At the end of the growing season, compared with the low-altitude habitat, the underground biomass of G. longipes in the high-altitude habitat increased significantly by 200%~290% for all three provenances. The response of gypenosides content to different altitude habitats varied with provenance and plant organs. The biomass of G. longipes strongly depended on the fine root characteristic indicators (P < 0.001), fine root length density, and fine root surface area. Our results also showed that the harvest yield of G. longipes could be effectively increased by promoting the growth of fine roots per unit leaf weight (P < 0.001, R2 = 0.63). Both fine root length density and fine root surface area had strong positive correlations with soil nutrient factors (R2 > 0.55) and a strong negative correlation with soil pH (R2 > 0.48). In a word, the growth of G. longipes is strongly controlled by the fine root morphological characteristics through the response of fine roots to soil nutrient factors and pH. Discussion: Our findings will help to deepen the understanding of the root ecophysiological basis driven by soil factors for the growth and secondary metabolites formation of G. longipes and other medicinal plants under changing habitat conditions. In future research, we should investigate how environmental factors drive plant morphological characteristics (e.g., fine roots) to affect the growth & quality of medicinal plants over a longer time scale.

Systematical characterization of gypenosides in Gynostemma pentaphyllum and the chemical composition variation of different origins.

Gynostemma pentaphyllum (Thunb.) Makino is an herbaceous plant of Cucurbitaceae family, which has been widely used as an herbal tea and traditional Chinese medicine. Since its saponins are similar to ginsenosides and have a wide range of activities, it has attracted wide interest. However, there are still a large number of unknown saponins that have not been isolated, especially some trace gypenosides. In the present study, a HILIC × RP offline two-dimensional liquid separation combined with a multimode data acquisition was developed for the systematical characterization of gypenosides. On top of the negative mode information, considering that saponins are prone to in-source fragmentations in positive ion mode, a precursor ion list data acquisition method was used for the targeted acquisition of multistage positive data. Reference herbal drug was taken as a golden sample to probe the chemical composition of G. pentaphyllum. The mixed sample of commercially available samples were also analyzed in parallel. Furthermore, the chemical compositions of commercially available samples from different sources were compared. In total, 1108 saponins were characterized, among which 588 were accurately characterized, with 574 identified in the reference herbal drug and 700 in the mixed commercially available samples. The commercially available samples showed great composition variation. These findings clarified the material basis and provided clues for quality control of G. pentaphyllum.

Gypenosides ameliorate high-fat diet-induced nonalcoholic fatty liver disease in mice by regulating lipid metabolism.

Gypenosides (GP), extracted from the traditional Chinese herb Gynostemma pentaphyllum (Thunb.) Makino, have been used to treat metabolic disorders, including lipid metabolism disorders and diabetes. Although recent studies have confirmed their beneficial effects in nonalcoholic fatty liver disease (NAFLD), the underlying therapeutic mechanism remains unclear. In this study, we explored the protective mechanism of GP against NAFLD in mice and provided new insights into the prevention and treatment of NAFLD. Male C57BL6/J mice were divided into three experimental groups: normal diet, high-fat diet (HFD), and GP groups. The mice were fed an HFD for 16 weeks to establish an NAFLD model and then treated with GP for 22 weeks. The transcriptome and proteome of the mice livers were profiled using RNA sequencing and high-resolution mass spectrometry, respectively. The results showed that GP decreased serum lipid levels, liver index, and liver fat accumulation in mice. Principal component and heatmap analyses indicated that GP significantly modulated the changes in the expression of genes associated with HFD-induced NAFLD. The 164 differentially expressed genes recovered using GP were enriched in fatty acid and steroid metabolism pathways. Further results showed that GP reduced fatty acid synthesis by downregulating the expression of Srebf1, Fasn, Acss2, Acly, Acaca, Fads1, and Elovl6; modulated glycerolipid metabolism by inducing the expression of Mgll; promoted fatty acid transportation and degradation by inducing the expression of Slc27a1, Cpt1a, and Ehhadh; and reduced hepatic cholesterol synthesis by downregulating the expression of Tm7sf2, Ebp, Sc5d, Lss, Fdft1, Cyp51, Nsdhl, Pmvk, Mvd, Fdps, and Dhcr7. The proteomic data further indicated that GP decreased the protein expression levels of ACACA, ACLY, ACSS2, TM7SF2, EBP, FDFT1, NSDHL, PMVK, MVD, FDPS, and DHCR7 and increased those of MGLL, SLC27A1, and EHHADH. In conclusion, GP can regulate the key genes involved in hepatic lipid metabolism in NAFLD mice, providing initial evidence for the mechanisms underlying the therapeutic effect of GP in NAFLD.

Gypenosides suppress fibrosis of the renal NRK-49F cells by targeting miR-378a-5p through the PI3K/AKT signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: The incidence of renal fibrosis caused by chronic kidney disease is increasing year by year. Preventing the activation and conversion of kidney-intrinsic fibroblasts to a myofibroblast phenotype is an important target for blocking the development of renal interstitial fibrosis. Our team established a stable renal interstitial fibrosis cell model in the early stage, and the screening results showed that GPs has good anti-fibrosis potential. At this stage, only a few literatures have reported its anti-fibrosis effect, and the mechanism of action is still unclear. AIM OF THE STUDY: The massive synthesis and secretion of extracellular-matrix (ECM) components by activated fibroblasts in the kidneys causes irreversible renal interstitial fibrosis. Gypenosides (GPs) have been shown to decelerate this process, in which micro RNAs (miRNAs) play an important regulatory role. This study aimed to evaluate the mechanism underlying the suppressive effect of GPs on renal fibrosis. MATERIALS AND METHODS: This study used TGF-ß1-stimulated NRK-49F renal cells as an in-vitro model of renal interstitial fibrosis. First, the concentration range of GPs that significantly affects the cytoactive was determined. Then, the anti-fibrotic effects of various concentrations of GPs in the in-vitro model were assessed via immunofluorescence, western blotting, and quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Non-coding-RNA sequencing combined with bioinformatics was used to predict the mechanistic basis of the anti-fibrotic effect of GPs, and qRT-PCR was used to verify the sequencing results and bioinformatic predictions. The identified relationships of the anti-fibrotic effect of GPs with miR-378a-5p and the PI3K/AKT signaling were evaluated using a miR-NC mimic and the PI3K inhibitor LY294002 as controls, respectively. RESULTS: TGF-ß1 stimulation up-regulated α-SMA, COL1, and COL3 in NRK-49F cells, and this effect was suppressed by GPs. Additionally, TGF-ß1 stimulation significantly changed the expression levels of 151 miRNAs, and GPs significantly suppressed the effect of TGF-ß1 on the levels of 18 of these miRNAs. Among them, miR-3588 and miR-378a-5p were down-regulated, and miR-135b-5p and miR-3068-5p were up-regulated upon TGF-ß1 induction. Of these miRNAs, miR-378a-5p was predicted to target the mRNAs of numerous proteins mainly enriched in the PI3K/AKT signaling pathway. The miRNA transfection experiments with the miR-NC mimic and PI3K inhibitor as controls showed that miR-378a-5p overexpression could suppress the TGF-ß1-induced up-regulation of α-SMA, COL1, PI3K, and AKT, including the phosphorylated form (p-AKT). CONCLUSION: GPs inhibit the PI3K/AKT signaling by up-regulating miR-378a-5p in TGF-ß1-stimulated NRK-49F cells and thereby reduce their massive secretion of ECM components. Given that this in-vitro model of renal interstitial fibrosis closely mimics the in-vivo pathogenesis, our results most likely apply to the in-vivo conditions.

Liver lipidomics analysis reveals the anti-obesity and lipid-lowering effects of gypnosides from heat-processed Gynostemma pentaphyllum in high-fat diet fed mice.

BACKGROUND: In traditional Chinese medicine, Gynostemma pentaphyllum (G. pentaphyllum) is widely used to treat conditions associated with hyperlipidemia, and its therapeutic potential has been demonstrated in numerous studies. However, the mechanism of lipid metabolism in hyperlipidemic by G. pentaphyllum, especially heat-processed G. pentaphyllum is not yet clear. PURPOSE: The aim of this study was to investigate the therapeutic mechanism of gypenosides from heat-processed G. pentaphyllum (HGyp) in hyperlipidemic mice by means of a lipidomics. METHODS: The content of the major components of HGyp was determined by ultra-performance liquid chromatography-electrospray ionization ion trap mass spectrometry (UPLC-ESI-MS). An animal model of hyperlipidaemia was constructed using C57BL/6J mice fed with high-fat diet. HGyp was also administered at doses of 50, 100 and 200 mg/kg, all for 12 weeks. Serum parameters were measured, histological sections were prepared and liver lipidome analysis using UPLC-MS coupled with multivariate statistical analysis. Quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting were used to analyze the genes and proteins associated with lipid lowering in HGyp. RESULTS: HGyp reduced body weight, serum total cholesterol (TC), triglyceride (TG) and low-density lipoprotein (LDL) and hepatic lipid accumulation in hyperlipidemic obese mice. To explore specific changes in lipid metabolism in relation to HGyp administration, lipid analysis of the liver was performed. Orthogonal partial least squares discriminant analysis (OPLS-DA) score plots showed that HGyp altered lipid metabolism in HFD mice. In particular, fatty acids (FA), triglycerides (DG), TG and ceramides (CER) were significantly altered. Eleven lipids were identified as potential lipid biomarkers, namely TG (18:2/20:5/18:2), TG (18:2/18:3/20:4), DG (18:3/20:0/0:0), Cer (d18:1/19:0), Cer (d16:1/23:0), Ceramide (d18:1/9Z-18:1), PS (19:0/18:3), PS (20:2/0:0), LysoPC (22:5), LysoPE (0:0/18:0), PE (24:0/16:1). Western blot and qRT-PCR analysis showed that these metabolic improvements played a role by down-regulating genes and proteins related to fat production (SREBP1, ACC1, SCD1), up-regulating genes and proteins related to lipid oxidation (CPTA1, PPARα) and lipid transport decomposition in the bile acid pathway (LXRα, PPARγ, FXR, BSEP). CONCLUSION: The lipid-lowering effect of gypenosides from heat-processed G. pentaphyllum is regulate lipid homeostasis and metabolism.

Comparison on anti-inflammatory activity of Gynostemma pentaphyllum processed with different methods / 中国中药杂志

The aim of this study is to investigate the effects of Gynostemma pentaphyllum dried with two different methods(air drying and heating) on inflammation in acute lung injury(ALI) mice in vivo and in vitro. Lipopolysaccharide(LPS) was sprayed into the airway of wild type C57BL/6J male mice to establish the model, and the drug was injected into the tail vein 24 h after modeling. Lung function, lung tissue wet/dry weight(W/D) ratio, the total protein concentration, interleukin 6(IL-6), IL-1β, and tumor necrosis factor-α(TNF-α) in the bronchoalveolar lavage fluid(BALF), and pathological changes of the lung tissue were used to evaluate the effects of different gypenosides on ALI mice. The results showed that total gypenosides(YGGPs) and the gypenosides substituted with one or two glycosyl(GPs_(1-2)) in the air-dried sample improved the lung function, significantly lowered the levels of IL-1β and TNF-α in BALF, and alleviated the lung inflammation of ALI mice. Moreover, GPs_(1-2) had a more significant effect on inhibiting NO release in RAW264.7 cells. This study showed that different drying methods affected the anti-inflammatory activity of G. pentaphyllum, and the rare saponins in the air-dried sample without heating had better anti-inflammatory activity.

Effects of gypenosides on proliferation and apoptosis of human gastric cancer cells and its mechanism / 中国药理学通报

Aim To explore the effect of gypenosides on proliferation and apoptosis of human gastric cancer cells SGC-7901 and AGS and its mechanism. Methods Different concentrations of gypenosides were cultured with human gastric cancer cells SGC-7901 and AGS. Cell viability assay was used to detect cell proliferation activity, and the IC

Gypenosides ameliorate ductular reaction and liver fibrosis via inhibition of hedgehog signaling.

Backgroud and aims: Ductular reaction (DR) is a common pathological change and thought to have a key role in the pathogenesis and progression of liver fibrosis. Our previous study reported Gypenosides (GPs) ameliorated liver fibrosis, however, the anti-fibrotic mechanisms of GPs are still unclear. Methods: Liver fibrosis was induced in rats by carbon tetrachloride combining with 2-acerylaminofluorene (CCl4/2-AAF), and Mdr2 knockout (Mdr2 -/-) mice to evaluate the anti-fibrotic role of GPs. In vitro, WB-F344 cells, a hepatic progenitor cells (HPCs) line, with or without Gli1 overexpressing lentiviral vectors, were induced by sodium butyrate (SB) to validate the mechanism of GPs and NPLC0393, the main ingredient of GPs. Results: Both in CCl4/2-AAF-treated rats and Mdr2 -/- mice, GPs obviously reduced the deposition of collagen and hydroxyproline content, inhibited the activation of hepatic stellate cells and inflammatory cell infiltration. Notably, GPs reduced the expressions of Epcam, CK19, CK7, Dhh, Smo, Ptch2, Gli1 and Gli2. Furthermore, CK19+ cells co-expressed Gli1, while the number of CK19+/Gli1+ cells was decreased by GPs. In vitro, GPs and NPLC0393 inhibited the differentiation of WB-F344 cells toward a biliary phenotype. Mechanistically, GPs and NPLC0393 protected against DR by inhibiting hedgehog signaling, which was supported by the results that DR, triggered directly by Gli1 overexpressing lentiviral vector was blocked by administration with GPs or NPLC0393. Conclusion: GPs attenuated DR and liver fibrosis by inhibiting hedgehog signaling, which provided more evidences and a novel mechanism of anti-fibrotic effect of GPs.

Comprehensive serum metabolomics and network analysis to reveal the mechanism of gypenosides in treating lung cancer and enhancing the pharmacological effects of cisplatin.

Gypenosides (GYP) exerted anticancer activity against various cancers. However, the mechanism of GYP against lung cancer (LC) in vivo remains unclear. This study aims to reveal the potential mechanism of GYP against LC and enhancing cisplatin efficacy using a comprehensive analysis of metabolomics, network analysis. Pharmacodynamic results showed that GYP inhibited tumor growth, reduced tumor volume and tumor weight, and alleviated pathological symptoms in Lewis tumor-bearing mice, and GYP could enhance the anti-LC effects of cisplatin. Using serum metabolomics methods, 53 metabolites were found to be significantly altered in the model group, and the levels of 23 biomarkers were significantly restored after GYP treatment. GYP-related metabolic pathways involved six pathways, including alpha-linolenic acid metabolism, glutathione metabolism, sphingolipid metabolism, glycerophospholipid metabolism, tryptophan metabolism, and primary bile acid biosynthesis. 57 genes associated with differential metabolites of GYP recovery and 7 genes of 11 saponins of GYP against LC were screened by network analysis, the STRING database was used to find the association between 57 genes and 7 genes, and a compound-intersection gene-metabolite related gene-metabolite-pathway network was constructed, and STAT3, MAPK14, EGFR and TYMS might be the crucial targets of GYP against LC. Western blot results showed that GYP restored the levels of STA3, MAPK14, EGFR, and TYMS in the model group, and GYP also restored the levels of STAT3 and MAPK14 in the cisplatin group, indicating that GYP might exert anti-LC effects and enhance the pharmacological effects of cisplatin through MAPK14/STAT3 signaling pathway. Our method revealed the effect and mechanism of GYP on LC and the pharmacological effects of GYP-enhanced chemotherapeutic agent cisplatin, which provided some reference for the development of anti-cancer drugs.

Gypenosides (GPs) alleviates hypoxia-induced injury in PC12 cells and enhances tolerance to anoxia in C57BL/6 mice.

Oxygen is a necessary substance for life activities, but reduced oxygen utilization due to high altitude exposure and respiratory dysfunction diseases could lead to pathological changes in the organisms. Herein gypenosides, the active ingredients in the food and medicine resource plant Gynostemma pentaphyllum (Thunb.) Makino were found to alleviate hypoxia-induced injury in PC12 cells. Moreover, hypoxia induced an increase in Ca2+ and reactive oxygen species content, and such patterns were both significantly reduced by gypenosides treatment. At the same time, gypenosides significantly blocked the decrease of both NO content and mitochondrial membrane potential caused by hypoxia. Furthermore, gypenosides gavage treatment significantly prolonged the survival time of C57BL/6 mice in confinement up to 24.3% and enhanced the locomotor ability of mice. Therefore, gypenosides have good neuroprotective effects and hypoxia tolerance activity and have the prospect of being developed as a preventive and therapeutic drug for hypoxia-related diseases. PRACTICAL APPLICATIONS: Gypenosides can enhance tolerance of cells and mice to hypoxia and have the potential to be developed into hypoxia-resistant health food and drugs.

[Preparation and in vitro evaluation of quercetin nanosuspension stabilized by gypenosides].

This study was designed to explore the potential of gypenosides as a novel natural stabilizer for the production of nanosuspensions. The gypenosides-stabilized quercetin nanosuspensions(QUE-NS) were prepared using the high-speed shearing and high-pressure homogenization method with quercetin as a model drug, followed by their in vitro evaluation.Based on the measured mean particle size and polydispersity index(PDI) of QUE-NS,the single factor experiment was conducted to optimize the preparation process parameters.The freeze-drying method was used to transform QUE-NS into freeze-dried powders, whose storage stability and saturation solubility were then studied.Moreover, the effects of pH and ionic strength on the physical stability of the nanosuspension system were examined.According to the results, the optimized process parameters were listed as follows: shear rate 13 000 r·min~(-1),shear time 2 min, homogenization pressure 100 MPa, and homogenization frequency 12 times.The mean particle size of QUE-NS prepared under the optimum process conditions was(461.9±2.4) nm, and the PDI was 0.059±0.016.During the two months of storage at room temperature, the freeze-dried QUE-NS powders remained stable.The saturation solubility of freeze-dried QUE-NS powders was proved higher than those of quercetin and the physical mixture.The results of stability testing demonstrated that QUE-NS stabilized with gypenosides exhibited good stability within the pH range of 6 to 8,while coalescence was prone to occur in the presence of salt.Overall, gypenosides is expected to become a new natural stabilizer for the preparation of nanosuspensions.

Chemical Characterization and Atherosclerosis Alleviation Effects of Gypenosides from Gynostemma pentaphyllum through Ameliorating Endothelial Dysfunction via the PCSK9/LOX-1 Pathway.

Dietary saponins have the potential to ameliorate atherosclerosis (AS). Gypenosides of Gynostemma pentaphyllum (GPs) have been used as functional foods to exhibit antiatherosclerotic activity. The present study aimed to explore the protective effect, underlying mechanism and active substances of GPs on AS in vivo and in vitro. Results demonstrated GPs administration reduced the serum concentrations of TC and LDL-C, upregulated the plasma HDL-C content, inhibited the secretion of ICAM-1, VCAM-1, and MCP-1, and alleviated vascular lesions in VitD3 plus high cholesterol diet-induced AS rats as well as reduced adhesion factors levels in ox-LDL-stimulated HUVECs, which was potentially associated with suppressing PCSK9/LOX-1 pathway. Further activity-guided phytochemical investigation of GPs led to the identification of five new dammarane-type glycosides (1-5) and ten known analogs (6-15). Bioassay evaluation showed compounds 1, 6, 7, 12, 13, and 14 observably reduced the expressions of PCSK9 and LOX-1, as well as the secretion of adhesion factors in injured HUVECs. Molecular docking experiments suggested that the active saponins of GPs might bind to the allosteric pocket of PCSK9 located at the catalytic and C-terminal domains, and 2α-OH-protopanaxadiol-type gypenosides might exert a higher affinity for an allosteric binding site on PCSK9 by hydrogen-bond interaction with ARG-458. These findings provide new insights into the potential nutraceutical application of GPs and their bioactive compounds in the prevention and discovery of novel therapeutic strategies for AS.

Gypenosides ameliorate high-fat diet-induced non-alcoholic steatohepatitis via farnesoid X receptor activation.

Background: Gypenosides (Gyps), the major botanical component of Gynostemma pentaphyllum, was found to up-regulate the farnesoid X receptor (FXR) in a mouse model of non-alcoholic steatohepatitis (NASH). However, the exact role of FXR and underlying mechanisms in Gyps-mediated effects on NASH remain to be elucidated. Purpose: This study investigated whether Gyps attenuates NASH through directly activating FXR in high-fat diet (HFD)-induced NASH, and delineated the molecular pathways involved. Study design: A mouse model of HFD-induced NSAH was used to examine effects of Gyps on NASH with obeticholic acid (OCA) as a positive control, and the role of FXR in its mechanism of action was investigated in wild-type (WT) and FXR knockout (KO) mice. Methods: WT or FXR KO mice were randomly assigned into four groups: normal diet (ND) group as negative control, HFD group, HFD + Gyps group, or HFD + OCA group. Results: Treatment with Gyps and OCA significantly improved liver histopathological abnormalities in HFD-induced NASH, reduced the non-alcoholic fatty liver disease (NAFLD) activity score (NAS), and lowered hepatic triglyceride (TG) content compared with the HFD group. In agreement with these liver tissue changes, biochemical tests of blood samples revealed that alanine aminotransferase (ALT), aspartate aminotransferase (AST), TG, total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), fasting blood glucose (FBG), and fasting insulin (FINS) levels were significantly lower in the HFD + Gyps vs. HFD group. Furthermore, Gyps and OCA treatment significantly up-regulated hepatic FXR, small heterodimer partner (SHP), carnitine palmitoyltransferase 1A (CPT1A), and lipoprotein lipase (LPL) expression, and significantly down-regulated sterol-regulatory element binding protein 1 (SREBP1), fatty acid synthetase (FASN), and stearoyl-CoA desaturase 1 (SCD1) protein levels compared with the HFD group in WT mice but not in FXR KO mice. Notably, Gyps- and OCA-mediated pharmacological effects were significantly abrogated by depletion of the FXR gene in FXR KO mice. Conclusion: Gyps ameliorated HFD-induced NASH through the direct activation of FXR and FXR-dependent signaling pathways.

Gynostemma pentaphyllum and Gypenoside-IV Ameliorate Metabolic Disorder and Gut Microbiota in Diet-Induced-Obese Mice.

Gynostemma pentaphyllum (G. pentaphyllum) is a perennial liana herb of the Cucurbitaceae family which has both nutraceutical and pharmacological functions. The objective of the current study was to investigate the preventative effects of G. pentaphyllum and Gypenoside-IV (GP-IV, a saponin monomer in G. pentaphyllum) on metabolic symptoms in high fat diet induced obese (DIO) mice with gut microbiota dysbiosis. G. pentaphyllum water extract (GPWE, 150 mg/kg•d- 1) and GP-IV (50 mg/kg•d- 1) were orally administered to DIO mice by gavage for 10 weeks. The results showed that both GPWE and GP-IV prevented obesity development by decreasing body weight gain, reducing fat mass/body weight ratio and inhibiting adipocyte hypertrophy. GPWE and GP-IV also improved lipid profile and glucose tolerance effectively. Moreover, GPWE and GP-IV treatments partly restored gut microbiota in DIO mice. Typically, GPWE and GP-IV reduced Firmicutes to Bacteroidetes ratio, increased the abundance of certain health-promoting bacteria and reduced the abundance of microbiota that were associated with metabolic disorders. We conclude that GPWE and GP-IV can ameliorate metabolic symptoms possibly via modulating gut microbiota in DIO mice.