Panax Notoginseng Saponins Protect Cardiac Myocytes Against Endoplasmic Reticulum Stress and Associated Apoptosis Through Mediation of Intracellular Calcium Homeostasis

Abstract

Endoplasmic reticulum (ER) stress has been demonstrated to play important roles in the pathogenesis of various cardiovascular diseases. The ER stress pathway is therefore a promising therapeutic target in cardiovascular disease. Although Panax notoginseng saponins (PNS) are one of the patent medicines that are traditionally used to treat cardiovascular disorders, their effects on ER stress in cardiac myocytes remain unexploited so far. This study investigates the effects of PNS on ER stress and its associated cell apoptosis along with the related mechanism in cardiac myocytes. PNS compounds were identified via high-performance liquid chromatograph (HPLC) assay. PNS-pretreated H9c2 cells, HL-1 cells, and primary cultured neonatal rat cardiomyocytes were stimulated with thapsigargin (TG) to induce ER stress response and apoptosis. ER stress response was tested by immunofluorescence or immunoblot of the ER protein chaperones—calnexin, binding immunoglobulin protein (BiP) and the C/EBP homologous protein (CHOP). Cell viability was tested by methyl thiazolyl tetrazolium (MTT) assay. Cell apoptosis was detected by immunoblot of Cleaved caspase-3 and flow cytometry analysis of Annexin V/propidium iodide (PI) staining. Cytosolic, mitochondrial, and ER calcium dynamics were investigated by calcium imaging. Moreover, a ryanodine receptor type-2 (RyR2) overexpression stable cell line was generated to verify the mechanism of RyR2 involved in PNS in the inhibition of ER stress and cell apoptosis. We demonstrate here that PNS protected cardiac myocytes from ER stress response and associated cell death in a concentration-dependent manner. Importantly, PNS reduced the elevation of cytosolic calcium, mitochondria calcium, as well as ER calcium in response to either TG or histamine treatment. PNS protection in ER stress was regulated by RyR2 expression. In summary, PNS protection against TG-induced ER stress response and its associated cell apoptosis in cardiac myocytes is calcium dependent. Through the regulation of ER calcium release mediated by RyR2, a novel mechanism for PNS in the prevention of cardiovascular diseases is thereby identified.

<p>Effects of Co-Administration of Icariin and <em>Panax notoginseng</em> Saponins on Intestinal Microbiota and Hippocampal Protein Expression in a Mouse Model of Alzheimer’s Disease</p>

“…PNS is the main active component of Panax notoginseng, which was able to improve cardiovascular system function and regulates immunity. 22,23 In this study, AD mice treated with ICA and PNS successfully completed MWM testing, and the learning and memory of mice in the middle dose group were improved to a certain extent. These results are consistent with previous publications regarding the combined application of ICA and PNS to treat memory impairment.…”

Inhibition of Autophagy Prevents Panax Notoginseng Saponins (PNS) Protection on Cardiac Myocytes Against Endoplasmic Reticulum (ER) Stress-Induced Mitochondrial Injury, Ca2+ Homeostasis and Associated Apoptosis

“…TG (content ≥98%) was purchased from Sigma (T9033, Sigma-Aldrich, United States). High performance liquid chromatograph (HPLC) of the main PNS components has shown in the previous report ( Chen et al, 2019 ). For drug treatment, cells were pretreated with 40 μ g/ml PNS for 12 h and then added 1 μm TG for 12 h.…”

“…The experimental protocol for animals was approved by the Ethics Committee for Scientific Research and Clinical Trials of the Affiliated Hospital of Zhengzhou University. The detailed isolation and purification of cardiomyocyte was referred to the previous protocols ( Chen et al, 2019 ). Briefly, cardiomyocytes were isolated using a neonatal cardiomyocytes isolation system (LK003303, Worthington Biochemical, United States).…”

“…After the fibroblasts and endothelial cells were extracted, the cardiomyocytes were then seeded onto cell culture plates pre-coated with 10 μg/ml of fibronectin and cultured in DMEM-F12 media containing 10 mM HEPES, 10% FBS, penicillin–streptomycin (100 U/ml–100 μg/ml), and 0.1 μmol/ml BrdU to inhibit non-myocyte cell proliferation. The cardiomyocytes were treated with PNS or TG after 72 h of tissue culture as indicated ( Chen et al, 2019 ).…”

“…Panax notoginseng saponin (PNS) extracted from Panax notoginseng is commonly used for treatment of cardiovascular disorders. Our previous study has shown that PNS protects against thapsigargin (TG)-induced ER stress response and its associated cell apoptosis in cardiac myocytes through the regulation of ER calcium release mediated by RyR 2 ( Chen et al, 2019 ). There is an evidence showing that PNS treatment enhanced HIF-1 α , which in turn increased autophagy in renal tissue ( Liang et al, 2017 ).…”