Activating PGC-1α-mediated signaling cascades in the aorta contributes to the amelioration of vascular senescence and atherosclerosis by 2,3,4',5-tetrahydroxystilbene-2-O-ß-d-glycoside.

BACKGROUND: 2,3,4',5-tetrahydroxystilbene-2-O-ß-d-glycoside (TSG), the main active polyphenolic component of Polygonum multiflorum, possesses many pharmacological activities. Its anti-aging effect influences a variety of tissues with diverse mechanisms. However, the effectiveness and exact mechanisms of TSG against vascular senescence in atherosclerosis remain unclear. The present study is aimed to investigate the effects of TSG against vascular senescence in atherosclerosis both in vivo and in vitro, and the possible underlying mechanisms focusing on aortic peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α)-mediated signaling cascades which have never been studied. METHODS: In vivo, 12-mo-old male LDLr-/- mice were randomly separated into control, high-fat diet (HFD), and TSG -treatment groups. At the end of the 12 weeks, the blood samples and aorta tissues of mice were collected for further analysis. In vitro, to mimic the condition of endothelial senescence in hyperlipidemic mice, human aortic endothelial cells (HAECs) were incubated with oxidized low-density lipoprotein (ox-LDL) to induce senescence. RESULTS: TSG administration improved lipid profiles, ameliorated HFD-exacerbated vascular senescence and atherosclerosis. The protective effect of TSG via inhibiting telomere malfunction, oxidative stress, and mitochondrial damage was found both in vivo and in vitro. Notably, TSG administration increased aortic PGC-1α mRNA and protein expression along with the regulation of its targeted genes TERT, NRF1, TFAM, Mn-SOD, and catalase. Further, by using PGC-1α siRNA in ox-LDL-treated HAECs, it is proved that TSG reduced endothelial senescence, telomere malfunction, oxidative stress, and mitochondrial damage at least partly through activating the PGC-1α pathway. CONCLUSIONS: These results provide new evidence for TSG in the treatment of atherosclerosis and the activation of aortic PGC-1α is involved in its beneficial effects.

Naringenin alleviates nonalcoholic steatohepatitis in middle-aged Apoe-/-mice: role of SIRT1.

BACKGROUND: Naringenin is naturally isolated from citrus fruits possessing many pharmacological activities. However, little is known about the effect of naringenin on nonalcoholic steatohepatitis (NASH) in the model of metabolic syndrome. PURPOSE: The present study is aimed to investigate the effect of naringenin on NASH in 12-mo-old male ApoE-/- mice and its possible underlying mechanism. METHODS: In vivo, 12-mo-old male ApoE-/- mice were administrated with naringenin by intragastric gavage for 12 weeks. At the end of experiment, the blood samples and liver tissues were collected. Metabolic parameters in serum, levels of triglyceride, cholesterol and hydroxyproline, activities of antioxidant enzymes, and content of inflammatory cytokines (TNF-α and IL-6) in liver were examined by corresponding assay kits. Pathological changes in liver were observed by hematoxylin-eosin, oil red O, masson's trichrome, picro-sirius red and senescence ß-galactosidase staining. Dihydroethidium was used for detection of reactive oxygen species (ROS). In vitro, AML-12 cells were treated with oleic acid in the presence or absence of naringenin for 24 h. Transfection of SIRT1 siRNA was also conducted in vitro. Lipid accumulation, cellular ROS generation, malondialdehyde content, antioxidant enzyme activities and secretion levels of TNF-α and IL-6 were examined. Both in vivo and in vitro, gene expressions were detected by real-time PCR or western blot. RESULTS: Naringenin administration improved metabolic parameters, suppressed hepatic steatosis, regulated expression of genes involved in lipid metabolism (FASN, SCD1, PPARα and CPT1α), reduced hepatic fibrosis and cell senescence, inhibited hepatic inflammation as evidenced by the decreased macrophage recruitment and content of TNF-α and IL-6, and reduced hepatic oxidative stress by suppressing ROS generation and normalizing activities of antioxidant enzymes. Notably, naringenin administration increased hepatic SIRT1 protein expression and activity along with the increased deacetylation of liver kinase B1 (LKB1), PGC1α and NF-κB. In vitro study, the benefits of naringenin on lipid accumulation, oxidative stress and inflammation were diminished by SIRT1 siRNA transfection. CONCLUSIONS: These results indicate that naringenin administration may be a potential curative therapy for NASH treatment and the activation of hepatic SIRT1-mediated signaling cascades is involved in its beneficial effects.

Polydatin attenuates atherosclerosis in apolipoprotein E-deficient mice: Role of reverse cholesterol transport.

BACKGROUND: Polydatin has been recently shown to possess extensive cardiovascular pharmacological activities. However, its protective effect against atherosclerosis in vivo remains poorly understood. The aim of the present study was to evaluate the potential effects of polydatin on high fat diet (HFD)-induced atherosclerosis using ApoE-/- mice, and explore the underlying mechanisms involved, especially focusing on reverse cholesterol transport (RCT) regulation. METHODS: after 12 weeks treatment, serum samples, mouse aorta, liver, peritoneal macrophages were collected to determine lipid profiles, atherosclerotic lesions, hepatic steatosis, foam cell formation and expression of related molecules. RAW264.7 macrophages were used to study cholesterol efflux. RESULTS: Polydatin improved serum lipid profiles, attenuated atherosclerosis and hepatic steatosis. Furthermore, polydatin may facilitate RCT by stimulating cholesterol efflux through ATP-binding cassette transporters (ABC) A1, ABCG1 and scavenger receptor class B type I (SR-BI) in macrophages, increasing serum levels of high density lipoprotein and apolipoprotein A-I, promoting of SR-BI-mediated cholesterol uptake of liver, increasing secretion of cholesterol into bile by ABCG5/ABCG8 and improving cholesterol metabolism by CYP7A1 pathway. Polydatin also regulated the protein expressions of hepatic fatty acid synthase and peroxisome proliferator-activated receptor-α. Additionally, polydatin reduced hepatic and aortic reactive oxygen species generation, normalized activities of antioxidant enzymes and increased protein expressions of NADPH-oxidase (NOX) 2 and NOX4 in liver. Polydatin also prevented hepatic and aortic inflammation as evidenced by the reduced macrophage infiltration and mRNA expressions of tumor necrosis factor-α and interleukin-6 in both aorta and liver. CONCLUSION: These findings indicated that polydatin can inhibit atherosclerosis through enhancement of overall RCT. In addition, anti-oxidative and anti-inflammatory effect of polydatin may also contribute to its inhibitory effects on atherosclerosis.

2, 3, 4', 5-tetrahydroxystilbene-2-0-ß-d Glycoside Attenuates Age- and Diet-Associated Non-Alcoholic Steatohepatitis and Atherosclerosis in LDL Receptor Knockout Mice and Its Possible Mechanisms.

The compound, 2,3,5,4'-tetrahydroxystilbene-2-O-ß-d-glucoside (TSG), a primary bioactive polyphenolic component of Polygonum multiflorum exerts numerous pharmacological activities. However, its protective effect against non-alcoholic steatohepatitis (NASH), in the context of metabolic syndrome, remains poorly understood. The aim of the present study is to evaluate the effects of TSG treatment on middle-aged (12-mo-old) male LDLr-/- mice, which were fed a high fat diet for 12 weeks to induce metabolic syndrome and NASH. At the end of the experiment, the blood samples of mice were collected for determination of metabolic parameters. Liver and aorta tissues were collected for analysis, such as histology, immunofluorescence, hepatic lipid content, real-time PCR, and western blot. Our data show that TSG treatment improved the different aspects of NASH (steatosis, inflammation, and fibrosis) and atherosclerosis, as well as some of the metabolic basal characteristics. These modulatory effects of TSG are mediated, at least in part, through regulating key regulators of lipid metabolism (SREBP1c, PPARα and their target genes, ABCG5 and CYP7A1), inflammation (CD68, TNF-α, IL-6 and ICAM), fibrosis (α-SMA and TNFß) and oxidative stress (NADPH-oxidase 2/4, CYP2E1 and antioxidant enzymes). These results suggest that TSG may be a promising candidate for preventing and treating the progression of NASH.

Inhibitory effects of 2,3,4',5-tetrahydroxystilbene-2-O-ß-D-glucoside on angiotensin II-induced proliferation of vascular smooth muscle cells.

OBJECTIVE: To investigate the effect of 2,3,4',5-tetrahydroxystilbene-2-O-ß-D-glucoside (TSG), an active component extracted from the root of Polygonum multiflorum, on angiotensin II (Ang II)-induced proliferation of cultured rat vascular smooth muscle cells (VSMCs) and to identify the potential mechanism. METHODS: Cell proliferation and cell cycle were determined by cell counting, 5-bromo-2'-deoxyuridine incorporation assay, proliferating cell nuclear antigen protein expression and flow cytometry. Levels of phosphorylated extracellular signal-regulated kinase 1/2 (ERK1/2), mitogenic extracellular kinase 1/2 (MEK1/2) and Src in VSMCs were measured by Western blot. The expression of c-fos, c-jun and c-myc mRNA were measured by reverse transcription polymerase chain reaction (RT-PCR). Intracellular reactive oxygen species (ROS) was measured by fluorescence assay. RESULTS: TSG significantly inhibited Ang II-induced VSMCs proliferation and arrested cells in the G /S checkpoint (P<0.05 or P<0.01). TSG decreased the levels of phosphorylated ERK1/2, MEK1/2 and Src in VSMCs (P<0.05 or P<0.01). TSG also suppressed c-fos, c-jun and c-myc mRNA expression <0.05 or P<0.01). In addition, the intracellular ROS was reduced by TSG (P<0.01). CONCLUSIONS: TSG inhibited Ang II-induced VSMCs proliferation. Its antiproliferative effect might be associated with down-regulation of intracellular ROS, followed by the suppression of the Src-MEK1/2-ERK1/2 signal pathway, and hence, blocking cell cycle progression.

The effect of 2,3,4',5-tetrahydroxystilbene-2-O-ß-D-glucoside on pressure overload-induced cardiac remodeling in rats and its possible mechanism.

The aim of the present study was to investigate the effects of 2,3,4',5-tetrahydroxystilbene-2-O-beta-D-glucoside, an active component extracted from Polygonum multiflorum, on pressure overload-induced cardiac remodeling in rats. A rat model with cardiac remodeling was induced by abdominal aortic banding. 2,3,4',5-Tetrahydroxystilbene-2-O-beta-D-glucoside (30, 60, 120 mg/kg/day) was administered 3 days after abdominal aortic banding and continued for 30 days. The abdominal aortic banding-treated rats had significantly elevated blood pressure, left ventricular hypertrophy, and myocardial fibrosis. Left ventricular hypertrophy was characterized by an increase in the ratios of the heart and left ventricular weights to body weight, and increased myocyte cross-sectional areas, hypertrophic ventricular septum, and left ventricular posterior wall. The accumulation of myocardial interstitial perivascular collagen and elevated cardiac hydroxyproline content indicated myocardial fibrosis. The pathological changes above were attenuated by 2,3,4',5-tetrahydroxystilbene-2-O-beta-D-glucoside. Additionally, it markedly reduced collagen I and III expressions and regulated matrix metalloproteinase-2,9 and inhibitors of metalloproteinase expressions, as markers of myocardial fibrosis. Furthermore, we explored the underlying mechanisms for such effects of 2,3,4',5-tetrahydroxystilbene-2-O-beta-D-glucoside. The results showed that it significantly reduced myocardium angiotensin II, enhanced the activities of superoxide dismutase and glutathione peroxidase in serum and myocardial tissue, as well as inhibited protein expression of transforming growth factor-ß1 and phosphorylation of extracellular signal-regulated kinase 1/2 and p38 mitogen-activated protein kinase in the myocardial tissue. Our results suggest that 2,3,4',5-tetrahydroxystilbene-2-O-beta-D-glucoside could prevent cardiac remodeling induced by pressure overload in rats. The underlying mechanisms may be related to a decreasing angiotensin II level, an antioxidant effect of the tested compound, suppression of transforming growth factor-ß1 expression, and inhibition of extracellular signal-regulated kinase 1/2 and p38 mitogen-activated protein kinase activation.

Role of mitochondrial permeability transition in human hepatocellular carcinoma Hep-G2 cell death induced by rhein.

Rhein, a compound found as a glucoside in the root of rhubarb, is currently a subject of interest for its antitumor properties. The apoptosis of tumor cell lines induced by rhein was observed, and the involvement of mitochondria was established; however, the role of mitochondrial permeability transition (MPT) remains unknown. Here we report that MPT plays an important role in the apoptosis of human hepatocellular carcinoma Hep-G2 cells induced by rhein. After adding rhein to the isolated hepatic mitochondria, swelling effects and the leakage of Ca(2+) were observed. These alterations were suppressed by cyclosporin A (CsA), an MPT inhibitor. Furthermore, in Hep-G2 cells, the decrease of ATP production, the loss of mitochondrial transmembrane potential (MTP), the release of cytochrome c (Cyto c), and the activation of caspase 3 were also observed. These toxic effects of rhein can also be attenuated by CsA as well. Moreover, TUNEL assay confirmed that in the presence of CsA, rhein-induced apoptosis was largely inhibited. These results suggest that MPT plays a critical role in the pathogenesis of Hep-G2 cell injury induced by rhein, and imply that MPT may contribute to the anti-cancer activity of rhein.

2,3,4',5-tetrahydroxystilbene-2-O-ß-D-glucoside inhibits proliferation of vascular smooth muscle cells: involvement of NO/cGMP/PKG pathway.

The proliferation of vascular smooth muscle cells (VSMCs) induced by injury to the intima of arteries is an important etiologic factor in vascular proliferative disorders such as atherosclerosis and restenosis. 2,3,4',5-Tetrahydroxystilbene-2-O-ß-D-glucoside (TSG), an active component extracted from Polygonum multiflorum, has been found to have an antiatherosclerotic effect. The aim of this study was to investigate the effects of TSG on platelet derived growth factor (PDGF)-BB induced VSMCs proliferation and to explore the possible mechanisms of such effects. Pretreatment of VSMCs with TSG significantly inhibited PDGF-BB-induced cell proliferation in a concentration-dependent but not time-dependent manner. In addition, flow cytometry analysis of the DNA content revealed blocking of the PDGF-BB-inducible cell cycle progression by TSG. On the contrary, an inhibitory effect of TSG on VSMCs proliferation and expression of cell cycle regulators were markedly attenuated by addition of an nitric oxide (NO) synthase inhibitor, a soluble guanylate cyclase inhibitor and a cyclic GMP (cGMP)-dependent protein kinase (PKG) inhibitor: N(G)-nitro-L-arginine methyl ester (L-NAME), 1H-[1,2,4] oxadiazolo [4,3-α] quinoxalin-1-one (ODQ) and KT5823, respectively. It was also demonstrated that TSG enhanced NO and cGMP formation through up-regulating endothelial NO synthase expression in VSMCs. The findings indicate that TSG inhibited VSMCs proliferation induced by PDGF-BB may involve the NO/cGMP/PKG signal pathway.

2,3,4',5-Tetrahydroxystilbene-2-O-ß-d-glucoside inhibits platelet-derived growth factor-induced proliferation of vascular smooth muscle cells by regulating the cell cycle.

1. 2,3,4',5-Tetrahydroxystilbene-2-O-ß-d-glucoside (TSG) has been shown to have an anti-atherosclerotic effect. Vascular smooth muscle cell (VSMC) proliferation contributes to the pathobiology of atherosclerosis. The aim of the present study was to investigate the effects of TSG on platelet-derived growth factor (PDGF)-BB-induced VSMC proliferation and to explore the molecular mechanisms underlying the effects. 2. Cultured rat VSMC were pretreated with TSG (l-50 µmol/L) for 1 h, followed by exposure to PDGF-BB (10 ng/mL) for 24 h, after which cell proliferation and cell cycle stages were examined. The expression of protein cell cycle regulators, including retinoblastoma (Rb), cyclin D1/E, cyclin-dependent kinase (CDK) 2/4, CDK inhibitors p21 and p27 and proliferative cell nuclear antigen (PCNA), was examined. Activation of extracellular signal-regulated kinase (ERK) 1/2 was evaluated to elucidate the possible upstream mechanism by which TSG affects cell cycle regulators. 3. The results showed that TSG dose-dependently inhibited PDGF-BB-induced VSMC proliferation, possibly by blocking the progression of the cell cycle from the G(1) to S phase. In addition, TSG significantly inhibited PDGF-BB-induced phosphorylation of Rb and the expression of cyclin D1, CDK4, cyclin E, CDK2 and PCNA. In addition, TSG suppressed PDGF-BB-induced downregulation of p27 and upregulation of p21, as well as PDGF-BB-induced activation of ERK1/2. 4. Together, the findings of the present study provide the first evidence that TSG can inhibit PDGF-BB-stimulated VSMC proliferation via cell cycle arrest in association with modulation of the expression of cell cycle regulators, which may be mediated, at least in part, by suppression of ERK1/2 activation.

Effects of 2,3,4',5-tetrahydroxystilbene 2-O-beta-D-glucoside on vascular endothelial dysfunction in atherogenic-diet rats.

2,3,4',5-Tetrahydroxystilbene 2- O-beta- D-glucoside (TSG), an active component extracted from Polygonum multiflorum, has been found to have an anti-atherosclerotic effect. The aim of this study was to investigate whether the TSG could prevent the development of atherosclerosis through influencing endothelial function in atherogenic-diet rats and to explore the possible mechanisms. Vascular endothelial dysfunction was assessed using isolated aortic ring preparation, transmission electron microscopy of the aorta, and levels of nitrate/nitrite (NOx) in serum and aorta. Endothelial nitric oxide (NO) synthase (eNOS) and inducible NO synthase (iNOS) mRNA and protein expression were also measured. After 12 weeks treatment, TSG improved acetylcholine-induced endothelium-dependent relaxation, prevented intimal remodeling, inhibited the decreased NOx content in serum and aorta in atherogenic-diet rats. Furthermore, the observed decreased eNOS mRNA and protein expression and increased iNOS mRNA and protein expression in atherogenic-diet rats were attenuated by TSG treatment. These results suggest that TSG could restore vascular endothelial function, which may be related to its ability to prevent changes of eNOS and iNOS expression, leading to preservation of NO bioactivity.

[The expression of matrix metalloproteinase-2,9 on atherosclerosis in experimental rats by treatment of 2,3,4',5-tetrahydroxystilbene -2-0-beta-D glucoside].

AIM: To observe the changes of MMP-2, 9 level on atherosclerosis in experimental rats by treatment of 2,3,4',5-tetrahydroxystilbene-2-0-beta-D glucoside (TSG) and to investigate the mechanism of TSG in stabilizing plaque and anti-atherosclerosis. METHODS: The atherosclerosis model of rat was made by feeding high grease food and injecting VitD3. Sixty male SD rats were randomly divided into six groups: control, Simvastatin, model and TSG 120 mg x kg(-1) x d(-1), TSG 60 mg x kg(-1) x d(-1) and TSG 30 mg x kg(-1) x d(-1). After 12 weeks, several aorta were randomly tested, model and TSG 120 mg x kg(-1) x d(-1), TSG 60 mg kg(-1) x d(-1) and TSG 30 mg x kg(-1) x d(-1). After 12 weeks, several aorta were randomly tested, model made was successful when we found plaque. And after six weeks treating, the mRNA expressions of MMP-2 and MMP-9 were measured by RT-PCR. The activities of MMP-2 and MMP-9 were measured by Western blot. The levels of CRP, IL-6 and TNF-alpha in serum were measured in biochemical method. RESULTS: Data of the study demonstrated that the level of TNF-alpha, IL-6, CRP, MMP-2 and MMP-9 were remarkably decreased by TSG60, 120 mg x kg(-1) x d(-1) groups, which showed a dose-dependent effect. CONCLUSION: TSG has the effect of anti-atherogenic and stabilizing plaque on the experimental rats with atherosclerosis, which are induced by the high cholesterol feeding and VitD3 injecting. The effect of TSG seems to be closely involved in regulating the expressions of MMP-2 and MMP-9, and inhibiting inflammation.

Cardioprotective effects of Astragali Radix against isoproterenol-induced myocardial injury in rats and its possible mechanism.

The purpose of the present study was to investigate the effects of the Chinese medical herb Astragali Radix on myocardial injury in vivo and its possible mechanisms. Myocardial injury in rats was induced by the subcutaneous injection of a high dose of isoproterenol for 10 days, and the therapeutic effects of Astragali Radix were observed. Cardiac hemodynamics, heart coefficient and marker enzymes in serum showed that Astragali Radix prevented isoproterenol-induced myocardial damage. Astragali Radix also improved the antioxidant status by decreasing the lipid peroxidative product malondialdehyde and increasing the activity of the antioxidant enzyme superoxide dismutase. The observed depressions in sarcoplasmic reticulum Ca2+-ATPase mRNA and protein expression as well as Ser(16)-phosphorylated phospholamban protein expression in isoproterenol-treated rats were attenuated by Astragali Radix treatment. Moreover, treatment with Astragali Radix showed higher myocardial cAMP content compared with the isoproterenol-alone group. These results suggest that the antioxidant property and partial prevention of changes in protein and gene expression of cardiac sarcoplasmic reticulum Ca2+ regulatory proteins which may be mediated through the cAMP pathway could help to explain the beneficial effects of Astragali Radix on myocardial injury in vivo.

Modification of alterations in cardiac function and sarcoplasmic reticulum by astragaloside IV in myocardial injury in vivo.

Astragaloside IV, the primary pure saponin isolated from Astragalus membranaceus has been found to have potent cardioprotective effects. In this study, we aim to investigate if the beneficial effects of astragaloside IV on cardiac function are associated with improvement in sarcoplasmic reticulum Ca(2+)-pump function in myocardial injury in vivo. Myocardial injury in rats was induced by subcutaneous injection of a high dose of isoproterenol, and the therapeutic effect of astragaloside IV was observed. Isoproterenol-treated rats showed widespread subendocardial necrosis, a rise in serum lactate dehydrogenase and creatine kinase, formation of lipid oxide product malondialdehyde and inhibition of left ventricular diastolic and systolic function, which suggested severe myocardial injury and acute heart failure. Moreover, sarcoplasmic reticulum Ca(2+)-uptake ability and Ca(2+)-ATPase (SERCA2a) activity were significantly reduced. And the level of SERCA2a mRNA and protein expression was also markedly decreased, associated with a decrease in Ser(16)-phosphorylated phospholamban protein expression, while total phospholamban level was unchanged in the isoproterenol-treated group compared with controls. However, these biochemical and hemodynamic changes in the acute failing hearts were prevented by treatment of isoproterenol-induced rats with astragaloside IV. Likewise, the observed reductions in sarcoplasmic reticulum Ca(2+)-pump function as well as in SERCA2a mRNA and protein levels and the phosphorylation level of phospholamban in the injured hearts were attenuated by astragaloside IV treatment. These results suggest that the beneficial effect of astragaloside IV on isoproterenol-induced myocardial injury may be due to its ability to prevent changes of SERCA2a and Ser(16)-phosphorylated phospholamban protein expression and, thus, may prevent the depression in sarcoplasmic reticulum Ca(2+) transport and improve cardiac function.