Hongxin Wang

Astragaloside IV attenuates inflammatory cytokines by inhibiting TLR4/NF-кB signaling pathway in isoproterenol-induced myocardial hypertrophy.

ETHNOPHARMACOLOGICAL RELEVANCE Astragaloside IV(As IV) is one of the main effective components isolated from the traditional Chinese medical herb Astragalus membranaceus. The protective effect of Astragalus membranaceus on myocardial hypertrophy has been extensively proved. To test the hypothesis that Astragaloside IV can ameliorate the myocardial hypertrophy and inflammatory effect induced by β-adrenergic hyperactivity, we carried out in vivo and in vitro experiments. MATERIAL AND METHODS In in vivo study, the isoproterenol(Iso) (5mg.kg-1.d-1) was used as a model of myocardial hypertrophy by intraperitoneal injection. SD rats were randomly assigned to following six groups: A:the control；B: Iso group；C: Iso plus As IV 20mg.kg-1.d-1；D: Iso plus As IV 40mg.kg-1.d-1；E: Iso plus As IV 80mg.kg-1.d-1；F: Iso plus Propranolol 40mg.kg-1.d-1. In in vitro study, cultured neonatal rat cardiomyocytes were pretreated with As IV(3, 10, 30μmol.L-1), Propranolol(2μmol.L-1) and BAY11-7082(5μmol.L-1) for 30minutes, and then incubated with Iso(10μmol.L-1) for 48 hours. For the rats in each group, the heart mass index (HMI) and the left ventricular mass index (LVMI) were measured. To measure the transverse diameter of left ventricular myocardial cells (TDM), the hematoxylin-eosin (HE) staining method was applied. In addition, the volume and the total protein content of cardiomyocytes were measured, the mRNA expression of ANP and TLR4 were quantified by RT-PCR, the protein expression of TLR4, IκBα and p65 were quantified by Western blot, and the level of TNF-α and IL-6 were measured by ELISA. RESULTS In vivo: Comparing the Iso group to the control, the HMI, LVMI, TDM were significantly increased; the protein expression of TLR4 and p65 were increased, while the IκBα were decreased; the expression of ANP, TLR4 mRNA, and TNF-α, IL-6 in serum were significantly increased. These changes could be partly prevented by As IV and Pro. In vitro: the over-expression of the cell size, total protein content could remarkably down-regulated by As IV and Pro, and the results of RT-PCR, Western blot and ELISA were similar to those of in vivo. CONCLUSIONS The results of these studies indicate that Astragaloside IV has good protective effect on myocardial hypertrophy induced by isoproterenol. More specifically, the cardioprotection is related to inhibiting the TLR4/NF-кB signaling pathway and the attenuating inflammatory effect. CHEMICAL COMPOUNDS STUDIED IN THIS ARTICLE Astragaloside IV (PubChem CID:122690); BAY 11-7082 (PubChem CID:5353431); Propranolol (PubChem CID:62882); Isoproterenol (PubChem CID: 5806).

Astragaloside IV Attenuates Injury Caused by Myocardial Ischemia/Reperfusion in Rats via Regulation of Toll‐Like Receptor 4/Nuclear Factor‐κB Signaling Pathway

Myocardial ischemia/reperfusion (MI/R) injury, in which inflammatory response and cell apoptosis play a vital role, is frequently encountered in clinical practice. Astragaloside IV (AsIV), a small molecular saponin of Astragalus membranaceus, has been shown to confer protective effects against many cardiovascular diseases. The present study was aimed to investigate the antiinflammatory and antiapoptotic effects and the possible mechanism of AsIV on MI/R injury in rats. Rats were randomly divided into sham operation group, MI/R group and groups with combinations of MI/R and different doses of AsIV. The results showed that the expressions of myocardial toll‐like receptor 4 (TLR4) and nuclear factor‐κB (NF‐κB) were significantly increased, and apoptosis of cardiomyocytes was induced in MI/R group compared with that in sham operation group. Administration of AsIV attenuated MI/R injury, downregulated the expressions of TLR4 and NF‐κB and inhibited cell apoptosis as evidenced by decreased terminal deoxynucleotidyl transferase dUTP nick end labeling positive cells, B‐cell lymphoma‐2 associated X protein and caspase‐3 expressions and increased B‐cell lymphoma‐2 expression compared with that in MI/R group. In addition, AsIV treatment reduced levels of inflammatory cytokines induced by MI/R injury. In conclusion, our results demonstrated that AsIV downregulates TLR4/NF‐κB signaling pathway and inhibits cell apoptosis, subsequently attenuating MI/R injury in rats. Copyright

Esculetin, a coumarin derivative, exerts in vitro and in vivo antiproliferative activity against hepatocellular carcinoma by initiating a mitochondrial-dependent apoptosis pathway

This study investigated the in vitro and in vivo antiproliferative activity of esculetin against hepatocellular carcinoma, and clarified its potential molecular mechanisms. Cell viability was determined by the MTT (tetrazolium) colorimetric assay. In vivo antitumor activity of esculetin was evaluated in a hepatocellular carcinoma mouse model. Seventy-five C57BL/6J mice were implanted with Hepa1-6 cells and randomized into five groups (n=15 each) given daily intraperitoneal injections of vehicle (physiological saline), esculetin (200, 400, or 700 mg·kg-1·day-1), or 5-Fu (200 mg·kg-1·day-1) for 15 days. Esculetin significantly decreased tumor growth in mice bearing Hepa1-6 cells. Tumor weight was decreased by 20.33, 40.37, and 55.42% with increasing doses of esculetin. Esculetin significantly inhibited proliferation of HCC cells in a concentration- and time-dependent manner and with an IC50 value of 2.24 mM. It blocked the cell cycle at S phase and induced apoptosis in SMMC-7721 cells with significant elevation of caspase-3 and caspase-9 activity, but did not affect caspase-8 activity. Moreover, esculetin treatment resulted in the collapse of mitochondrial membrane potential in vitro and in vivo accompanied by increased Bax expression and decreased Bcl-2 expression at both transcriptional and translational levels. Thus, esculetin exerted in vitro and in vivo antiproliferative activity in hepatocellular carcinoma, and its mechanisms involved initiation of a mitochondrial-mediated, caspase-dependent apoptosis pathway.

Role of endogenous opioid peptides in the infarct size-limiting effect of adaptation to chronic continuous hypoxia.

AIMS The objective of this study was to examine the involvement of endogenous opioid peptides and opioid receptor (OR) subtypes in the cardioprotective effect of adaptation to chronic hypoxia in rats. MAIN METHODS Rats were exposed to continuous normobaric hypoxia (CNH; 12% oxygen) for 3 weeks. Myocardial ischemia was induced by 20-min coronary artery occlusion followed by 3-h reperfusion in anesthetized open-chest animals. Various OR antagonists were administered to rats prior to ischemia. The size of myocardial infarction and the incidence of ischemic ventricular arrhythmias were assessed. Myocardial and plasma concentrations of opioid peptides (met-enkephalin, β-endorphin, and endomorphins) were determined. KEY FINDINGS Adaptation to CNH significantly increased myocardial and plasma concentrations of opioids, potentiated their further elevation by ischemia/reperfusion, and reduced myocardial infarct size, but it did not affect the incidence of ischemic arrhythmias. The infarct size-limiting effect of CNH was abolished by OR antagonists naltrexone (non-selective), naloxone methiodide (non-selective peripherally acting), TIPP[ψ] (δ-OR), naltriben (δ2-OR), or CTAP (μ-OR), while BNTX (δ1-OR) and nor-binaltorphimine (κ-OR) had no effect. SIGNIFICANCE The results suggest that the infarct size-limiting effect afforded by adaptation to CNH is mediated by activation of peripheral δ2- and μ-ORs by elevated levels of endogenous opioid peptides.

Astragaloside IV Protects against Isoproterenol-Induced Cardiac Hypertrophy by Regulating NF-κB/PGC-1α Signaling Mediated Energy Biosynthesis

We previously reported that Astragaloside IV (ASIV), a major active constituent of Astragalus membranaceus (Fisch) Bge protects against cardiac hypertrophy in rats induced by isoproterenol (Iso), however the mechanism underlying the protection remains unknown. Dysfunction of cardiac energy biosynthesis contributes to the hypertrophy and Nuclear Factor κB (NF-κB)/Peroxisome Proliferator-Activated Receptor-γ Coactivator 1α (PGC-1α) signaling gets involved in the dysfunction. The present study was designed to investigate the mechanism by which ASIV improves the cardiac hypertrophy with focuses on the NF-κB/PGC-1α signaling mediated energy biosynthesis. Sprague-Dawley (SD) rats or Neonatal Rat Ventricular Myocytes (NRVMs) were treated with Iso alone or in combination with ASIV. The results showed that combination with ASIV significantly attenuated the pathological changes, reduced the ratios of heart weight/body weight and Left ventricular weight/body weight, improved the cardiac hemodynamics, down-regulated mRNA expression of Atrial Natriuretic Peptide (ANP) and Brain Natriuretic Peptide (BNP), increased the ratio of ATP/AMP, and decreased the content of Free Fat Acid (FFA) in heart tissue of rats compared with Iso alone. In addition, pretreatment with ASIV significantly decreased the surface area and protein content, down-regulated mRNA expression of ANP and BNP, increased the ratio of ATP/AMP, and decreased the content of FFA in NRVMs compared with Iso alone. Furthermore, ASIV increased the protein expression of ATP5D, subunit of ATP synthase and PGC-1α, inhibited translocation of p65, subunit of NF-κB into nuclear fraction in both rats and NRVMs compared with Iso alone. Parthenolide (Par), the specific inhibitor of p65, exerted similar effects as ASIV in NRVMs. Knockdown of p65 with siRNA decreased the surface areas and increased PGC-1α expression of NRVMs compared with Iso alone. The results suggested that ASIV protects against Iso-induced cardiac hypertrophy through regulating NF-κB/PGC-1α signaling mediated energy biosynthesis.

Astragaloside IV protects against cardiac hypertrophy via inhibiting the Ca2+/CaN signaling pathway.

Astragaloside IV is widely used for the treatment of cardiovascular diseases in China. However, its role in cardiac hypertrophy remains unclear. In this study, we aim to determine the protective effects of astragaloside IV on myocardial hypertrophy induced by lipopolysaccharide and to identify their precise molecular and cellular mechanisms. Cell size, reorganization of actin filaments, and ANP and BNP mRNA expression were used as indices of hypertrophy; CaN and GATA-4 expression and the distribution of NFAT-3 in both cytoplasm and nucleus were determined by Western blot analysis; Ca2+ transient in Fura-2/AM-loaded cells was measured by Till image system. Our data demonstrated that lipopolysaccharide challenge induced cardiac hypertrophy, increased resting Ca2+ transient level, promoted activation of CaN and GATA-4, and enhanced nuclear translocation of NFAT-3. Administration of astragaloside IV (16, 32, and 64 µM) 1 h prior to lipopolysaccharide stimulation dose-dependently attenuated cardiac hypertrophy induced by lipopolysaccharide. Further studies demonstrated that astragaloside IV inhibited the increment of the resting intracellular free Ca2+, and its effect was similar to verapamil. Moreover, astragaloside IV also inhibited the activation of CaN and GATA-4, and the nuclear translocation of NFAT-3 induced by lipopolysaccharide. In conclusion, our results revealed that astragaloside IV had the potential to protect against cardiac hypertrophy through Ca2+-mediated CaN signaling pathways.

Astragalus polysaccharide attenuates isoproterenol-induced cardiac hypertrophy by regulating TNF-α/PGC-1α signaling mediated energy biosynthesis.

We previously reported that Astragalus polysaccharide (APS) extracted from Chinese medicine Astragalus membranaceus (Fisch.) Bge, attenuates hypertrophy of neonatal rat ventricular myocytes (NRVMs) induced by isoproterenol (Iso). The present study was designed to investigate the effects and the possible mechanism of APS on Iso-induced hypertrophy in rats and NRVMs with focus on tumor necrosis factor α (TNF-α)/peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α) signaling mediated energy biosynthesis. 36-Week old rats were randomly divided into 3 groups: (1) Control, rats received vehicle; (2) Iso, rats received isoproterenol injections; (3) Iso+APS, rats received isoproterenol injections and APS. NRVMs were divided into similar groups as rats. The results showed that combination of APS with Iso significantly attenuated the pathological changes, reduced the ratios of heart weight/body weight (HW/BW) and left ventricular weight/BW (LVW/BW), improved the cardiac hemodynamics, down-regulated mRNA and protein expression of atrial natriuretic peptide (ANP), increased the ratios of ATP/ADP and ATP/AMP, and decreased the content of free fatty acid (FFA) in heart tissue of rats compared with Iso alone. In addition, pretreatment with APS significantly decreased the surface area and protein content, down-regulated mRNA and protein expression of ANP, increased the ratios of ATP/ADP and ATP/AMP, and decreased the content of FFA in NRVMs compared with Iso alone. Furthermore, APS increased the protein expressions of ATP5D, the σ subunit of ATP synthase, PGC-1α and pyruvate dehydrogenase kinase 4 (PDK4) in tissue and NRVMs respectively and inhibited the production of TNF-α in serum and culture medium compared with Iso alone. The results suggested that APS attenuates Iso-induced cardiac hypertrophy through regulating TNF-α/PGC-1α signaling mediated energy biosynthesis.

Astragaloside IV improves the isoproterenol-induced vascular dysfunction via attenuating eNOS uncoupling-mediated oxidative stress and inhibiting ROS-NF-κB pathways.

OBJECTIVE Oxidative stress and inflammation are regarded as two important triggers of endothelial dysfunction and play pivotal role in progression of vascular damage associated with cardiac hypertrophy. Our previous studies demonstrated that astragaloside IV (AsIV) could protect against cardiac hypertrophy in rats induced by isoproterenol (Iso), but its effects on the aorta are not known. In present study, we aimed to assess the effects of AsIV on Isoinduced vascular dysfunction. METHODS Sprague-Dawley (SD) rats were treated with Iso (10mg/kg/d) alone or in combination with AsIV (50mg/kg/d). RESULTS Compared with Isotreated alone, AsIV significantly reduced the ratios of heart weight/body weight and left ventricular weight/body weight. AsIV ameliorated the increased vasoconstriction response to phenylephrine induced by Iso and suppressed superoxide anion generation in rat aorta, increased endothelial nitric oxide synthase (eNOS) dimer/monomer ratio and its critical cofactor tetrahydrobiopterin (BH4) content in aorta as well as the NO production in the serum, reduced the plasmatic peroxynitrite (ONOO-). Moreover, in contrast with Isotreatment alone, AsIV decreased the ratio of nuclear-to-cytosolic protein expression of the NF-κB p65 subunit while enhanced its inhibited protein expression of IκB-α, down-regulated mRNA expression of IL-1β, IL-6 and TNF-α of the aorta. CONCLUSIONS The present study suggested that AsIV protects against Isoinduced vascular dysfunction probably via attenuating eNOS uncoupling-mediated oxidative stress and inhibiting ROS-NF-κB pathways.

Astragaloside IV attenuates apoptosis of hypertrophic cardiomyocyte through inhibiting oxidative stress and calpain-1 activation.

Calpain-1 activation and oxidative stress are two critical factors contributing to apoptosis of hypertrophic cardiomyocyte. Astragaloside IV (ASIV) exhibits protective effect against various heart diseases. The present study was designed to investigate whether the inhibitory effect of ASIV on isoproterenol (ISO)-induced apoptosis of hypertrophic cardiomyocyte was associated with the anti-oxidation and calpain-1 inhibition. Hypertrophy, apoptosis, mitochondrial oxidative stress and calpain-1 expression were measured in the heart tissue of Sprague-Dawley (SD) rats and H9C2 cells treated with ISO alone or combination with ASIV. The results showed that ASIV attenuated apoptotic rate, increased Bcl-2 expression, decreased Bax expression, ameliorated the integrity of mitochondrial structure and improved mitochondrial membrane potential (MMP). Moreover, ASIV combination reduced both calpain-1 protein expression and calpain activity, down-regulated mitochondrial NOX4 (mito-NOX4) expression, increased activity of mitochondrial superoxide dismutase (mito-SOD) and mitochondrial catalase (mito-CAT) compared to ISO treated alone. The results suggested that ASIV exerted anti-apoptosis effect on ISO-induced hypertrophic cardiomyocyte by attenuating oxidative stress and calpain-1 activation.

Inhibition of TNF-α-mediated NF-κB activation by Ginsenoside Rg1 contributes the attenuation of cardiac hypertrophy induced by abdominal aorta coarctation.

Abstract: Ginsenoside Rg1 (Rg1), a protopanaxadiol saponin extracted from Chinese medicine Panax ginseng C.A. Meyer, has been demonstrated to inhibit the cardiac hypertrophy. However, the molecular mechanisms underlying the inhibition remain poorly understood. Activation of nuclear factor-kappa B (NF-&kgr;B) mediated by tumor necrosis factor &agr; (TNF-&agr;) gets involved in the cardiac hypertrophy. This study is designed to investigate the effects and the potential mechanism of Rg1 on the abdominal aorta coarctation (AAC)-induced cardiac hypertrophy with focus on TNF-&agr;/NF-&kgr;B signaling pathway. The results showed that oral administration of Rg1 dose-dependently improved the pathological changes, decreased the ratios of left ventricular weight/body weight (LVW/BW) and heart weight/BW (HW/BW), corrected the dysfunction of the cardiac hemodynamics by decreasing the left ventricular systolic pressure and left ventricular end-diastolic pressure and increasing the maximal rate of left ventricular systolic and diastolic pressure (±dp/dtmax) compared with the AAC alone. Rg1 also downregulated the atrial natriuretic peptide mRNA expression and decreased the mRNA and protein expression of TNF-&agr; in the heart tissue of rats compared with the AAC alone. In addition, Rg1 and BAY, the specific inhibitor of NF-&kgr;B, decreased the protein content and downregulated the mRNA expression of atrial natriuretic peptide in neonatal rat ventricular myocytes treated with TNF-&agr;. Furthermore, Rg1 increased the protein expression of p65, the subunit of NF-&kgr;B, in cytoplasm and decreased the expression p65 in nucleus of the heart tissue of rats undergoing the AAC and of neonatal rat ventricular myocytes treated with TNF-&agr;. The results suggested that Rg1 attenuates the AAC-induced cardiac hypertrophy through inhibition of TNF-&agr;/NF-&kgr;B signaling pathway.