Abstract
Purpose: To investigate the protective effect of polysaccharide (BRP) from the root of Bupleurum Chinese DC. and Bupleurum scorzonerifolium Willd. on cardiomyocyte cells.
 Methods: Response surface methodology (RSM) based on Box-Behnken Design (BBD) was performed to optimize the extraction conditions for BRP. The effect of BRP on cardiomyocyte cell apoptosis was evaluated in H9c2 cells treated with hydrogen peroxide (H2O2). Cell viability was determined by CCK-8 assay, while oxidative stress levels in H9c2 cells, including lactate dehydrogenase (LDH), superoxide dismutase (SOD), catalase (CAT) and creatine kinase (CK) were determined using commercial kits following the manufacture’s instruction. mRNA expressions (caspase-3, caspase-8, caspase-9 and Fas) were determined by quantitative real time-polymerase chain reaction (RT-qPCR).
 Results: The obtained optimal extraction conditions for BRP was as follows: extraction time (1.43 h), ratio of water to the raw material (30 mL/g) and extraction times (2 times). BRP (200, 400, 600 and 800 μg/mL) significantly increased the cell viability of H2O2 induced H9c2 cells (p < 0.05, p < 0.01, p < 0.01, p < 0.01, respectively). BRP (200, 400 and 800 μg/mL) significantly decreased LDH and CK levels (p < 0.01, p < 0.01, p < 0.01, respectively). However, BRP increased levels of SOD (200, 400 and 800 μg/mL, p < 0.05) and CAT (400 and 800 μg/mL, p < 0.05) in H9c2 cells. BRP significantly downregulated mRNA expressions of Caspase-3, Caspase-8, Caspase-9 and Fas (200, 400 and 800 μg/mL, p < 0.01) in H9c2 cells induced by H2O2.
 Conclusion: BRP protects cardiomyocyte against apoptosis via inhibition of oxidative stress and mitochondria-mediated intrinsic apoptosis, and thus, may be potential therapeutic agent for the management of cardiovascular diseases.
 Keywords: Bupleurum Chinese, Bupleurum scorzonerifolium Willd., Polysaccharide, Cardiomyocyte, Apoptosis, H9c2 cell, Biochemical parameters
Highlights
Incidence of cardiovascular disease is increasing every year, and myocardial infarction has become a leading cause of death worldwide [1]
The F-test was used to test the statistical significance of the regression equation, and the effects of independent variables on the extraction yield were checked for adequacy and fitness by analysis of variance (ANOVA)
The average Bupleuri Radix polysaccharide (BRP) yield from the actual experiments was 6.08%, demonstrating the Response surface methodology (RSM) model validity
Summary
Incidence of cardiovascular disease is increasing every year, and myocardial infarction has become a leading cause of death worldwide [1]. Phytochemical studies have revealed the presence of saponins, volatile oils, polysaccharides, flavonoids, and sterols in BR [5]. It is well-known that polysaccharides are important biological macromolecules isolated from natural products [6]. Polysaccharides derived from naturally occurring medicinal plants have been evaluated as drug candidates for treating various diseases due to their widespectrum of therapeutic activities, relative low toxicities, and minor side-effects [7,8]. No reports exist on the protective effect of Bupleuri Radix polysaccharide (BRP) on cardiomyocyte cells and its possible mechanisms of action on myocardial cell preservation. We aimed to evaluate the protective effect of BRP on the H2O2-induced H9c2 cell model
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