Ginsenoside Rg1 and Rb1, in combination with salvianolic acid B, play different roles in myocardial infarction in rats

Background: The herb pair of Salvia miltiorrhiza and Panax notoginseng has widely been used for improving coronary and cerebral circulation in China. However, the exact contribution of the major active components of S. miltiorrhiza and P. notoginseng to cardioprotection is far from clear. In the present study, three representative ingredients, salvianolic acid B (SalB) from S. miltiorrhiza and ginsenoside Rg1 (Rg1) and ginsenoside Rb1 (Rb1) from P. notoginseng, were selected to elucidate the mechanism of the herb pair at the ingredient level. Methods: The purity of SalB, Rg1, and Rb1 was >99%, as detected by high-performance liquid chromatography. Acute myocardial infarction was introduced by ligation of the left anterior descending coronary artery near the main pulmonary artery. Cardiac contractility was detected through a Mikro-tipped catheter, and cardiac infarct size was determined using triphenyltetrazolium chloride stain. Results: The combination of SalB and Rg1, and not the combination of SalB and Rb1, improved heart contractility in rats with myocardial infarction. The different contributions of Rg1 and Rb1, in combination with SalB, to cardioprotection provides further direction to optimize and modernize the herbal medicines containing S. miltiorrhiza and P. notoginseng. Conclusion: The combination of SalB and Rg1 may provide potential protection against myocardial infarction

The Safety Evaluation of Salvianolic Acid B and Ginsenoside Rg1 Combination on Mice

Our previous study indicated that the combination of salvianolic acid B (SalB) and ginsenoside Rg1 (Rg1), the main components of Salvia miltiorrhizae and Panax notoginseng, improves myocardium structure and ventricular function in rats with ischemia/reperfusion injury. The present study aimed to determine the safety of the combined SalB and Rg1 (SalB-Rg1) in mice. The safety of SalB-Rg1 was evaluated through acute toxicity and repeated-dose toxicity. In the acute toxicity study, the up and down procedure was carried out firstly, and then, the Bliss method was applied. In the toxicity study for seven-day repeated treatment of SalB-Rg1, forty Kunming mice were randomly divided into four groups. The intravenous median lethal dose (LD50) of the SalB-Rg1 combination was 1747 mg/kg using the Bliss method. For both the acute toxicity study and the seven-day repeated toxicity study, SalB-Rg1 did not induce significant abnormality on brain, heart, kidney, liver and lung structure at any dose based on H&E stain. There were no significant changes related to the SalB-Rg1 toxicity detected on biochemical parameters for two kinds of toxicity studies. The LD50 in mice was 1747 mg/kg, which was more than one hundred times higher than the effective dose. Both studies of acute toxicity and seven-day repeated dose toxicity indicated the safety of the SalB-Rg1 combination

Impact of Autophagy Inhibition at Different Stages on Cytotoxic Effect of Autophagy Inducer in Glioblastoma Cells

Background/Aims: Glioblastoma multiforme (GBM) is the most malignant primary brain tumor with a poor prognosis. Combination treatment of autophagy inducer and autophagy inhibitor may be a feasible solution to improve the therapeutic effects. However, the correlation between them is unclear. The purpose of this study was to investigate the effect of autophagy inhibition at different stages on cytotoxicity of autophagy inducers in glioblastoma cells. Methods: Autophagy inhibition at early stage was achieved by 3-methyladenine (3-MA) or Beclin 1 shRNA. Autophagy inhibition at late stage was achieved by chloroquine (CQ) or Rab7 shRNA. Cell viability was assessed by MTT assay. Autophagy was measured using transmission electron microscopy and western blot. Apoptosis was measured using western blot and flow-cytometry. Results: Inhibition of early steps of autophagy by 3-MA or Beclin 1 knockdown decreased the toxic effect of arsenic trioxide (ATO) in GBM cell lines. In contrast, blockade of autophagy flux at late stage by CQ or Rab7 knockdown enhanced the cytotoxicity of ATO, and caused accumulation of degradative autophagic vacuoles and robust apoptosis. Moreover, depletion of Beclin 1 abolished the synergistic effect of ATO and CQ by reducing autophagy and apoptosis. Combination of CQ with other autophagy inducers also induced synergistic apoptotic cell death. Conclusion: These results suggest that inhibition of late process of autophagy, not initial step, increases the cytotoxic effect of autophagy inducers via autophagy and apoptosis, which may contribute to GBM chemotherapy

Combined Salvianolic Acid B and Ginsenoside Rg1 Exerts Cardioprotection against Ischemia/Reperfusion Injury in Rats

<div><p>Lack of pharmacological strategies in clinics restricts the patient prognosis with myocardial ischemia/reperfusion (I/R) injury. The aim of this study was to evaluate the cardioprotection of combined salvianolic acid B (SalB) and ginsenoside Rg1 (Rg1) against myocardial I/R injury and further investigate the underlying mechanism. I/R injury was induced by coronary artery ligation for Wistar male rats and hypoxia/reoxygenation injury was induced on H9c2 cells. Firstly, the best ratio between SalB and Rg1was set as 2:5 based on their effects on heart function detected by hemodynamic measurement. Then SalB-Rg1 (2:5) was found to maintain mitochondrial membrane potential and resist apoptosis and necrosis in H9c2 cell with hypoxia/reoxygenation injury. Companying with same dose of SalB or Rg1 only, SalB-Rg1 showed more significant effects on down-regulation of myocardial infarct size, maintenance of myocardium structure, improvement on cardiac function, decrease of cytokine secretion including TNF-α, IL-1β, RANTES and sVCAM-1. Finally, the SalB-Rg1 improved the viability of cardiac myocytes other than cardiac fibroblasts in rats with I/R injury using flow cytometry. Our results revealed that SalB-Rg1 was a promising strategy to prevent myocardial I/R injury.</p></div

The ratio for SalB to Rg1 was set as 2:5 based on ventricular function.

<p>I/R injury was induced by 40 minutes coronary artery ligation followed by 1 hour reperfusion. Ventricular function was evaluated by hemodynamic parameters including maximum rate of pressure development for contraction (+dP/dt_max), maximum rate of pressure development for relaxation (-dP/dt_min), left ventricular systolic pressure (LVSP) and end-diastolic pressure (EDP). All the values were expressed as mean ± S.E. *p<0.05, **p<0.01, ***p<0.001 versus Sham; #p<0.05 versus I/R; n≥6 for every group.</p

SalB-Rg1 (2:5) improved cardiac function.

<p>10mg/kg SalB-Rg1 significantly improved ventricular function by evaluation of hemodynamic parameters including +dP/dt_max,-dP/dt_min, EDP and LVSP. No improvement on cardiac function was found with the treatment of SalB or Rg1 at 10 mg/kg only. All the values are expressed as mean ± S.E. *p<0.05, **p<0.01versus Sham; #p<0.05, ##p<0.01 versus I/R; n≥6 for every group.</p

SalB-Rg1 protected H9c2 cells against apoptosis and necrosis.

<p>(A) The protection of SalB-Rg1 against H9c2 apoptosis was detected by mitochondrial depolarization using JC-1 stain. (B) Quantification of the ratio for JC-1 multimer to JC-1 monomer fluorescence intensity (red:green). (C) The apoptosis or necrosis of H9c2 cell was evaluated using Annexin V-FITC/PI Apoptosis Detection Kit. Apoptotic cells were stained green (Annexin V-FITC) and necrosis cells were stained red (PI). At least three time experiments were repeated and the representative figures were shown.</p