Baicalin Protects the Cardiomyocytes from ER Stress-Induced Apoptosis: Inhibition of CHOP through Induction of Endothelial Nitric Oxide Synthase

Mingzhi Shen,Lei Gao,Ke Cheng,Yong Xu,Lin Wang,Li Fan,Liangliang Shen,Chao Zeng,Guodong Yang,Bo Wang,Xiaowang Guo,Xiaoming Wang,Xiumin Li,Yuesheng Xia,Haichang Wang,Xin-Liang Ma

doi:10.1371/journal.pone.0088389

Abstract

Baicalin, the main active ingredient of the Scutellaria root, exerts anti-oxidant and anti-apoptotic effects in cardiovascular diseases. However, the therapeutic mechanism of baicalin remains unknown. Cultured neonatal rat cardiomyocytes were pre-treated with baicalin (0–50 µM) for 24 h, and subsequently treated with tunicamycin (100 ng/ml). Cell viability was detected by MTT assay, and cell damage was determined by LDH release and TUNEL assay. The expression of CHOP, JNK, caspase-3, eNOS was analyzed by western blot. NO was measured by DAF-FM staining. As a result, treatment with baicalin significantly reduced apoptosis induced by ER stress inducer tunicamycin in cardiomyocytes. Molecularly, baicalin ameliorated tunicamycin-induced ER stress by downregulation of CHOP. In addition, baicalin inverted tunicamycin-induced decreases of eNOS mRNA and protein levels, phospho eNOS and NO production through CHOP pathway. However, the protective effects of baicalin were significantly decreased in cardiomyocytes treated with L-NAME, which suppressed activation of nitric oxide synthase. In conclusion, our results implicate that baicalin could protect cardiomyocytes from ER stress-induced apoptosis via CHOP/eNOS/NO pathway, and suggest the therapeutic values of baicalin against ER stress-associated cardiomyocyte apoptosis.

Highlights

The endoplasmic reticulum (ER) is recognized as an organelle that participates in the folding of secretory and membrane proteins [1,2]
Compared with Control group, tunicamycin for 24 h had no effect on cell viability or Lactate Dehydrogenase (LDH) release, whereas cell viability decreased, and LDH release increased after 48- to 96-h exposure to tunicamycin (Figure 2A and 2B)
When cardiomyocytes were treated with baicalin (25 mM) alone, there were no obvious effect on survival cell number (Figure 3A), LDH release (Figure 3B)

Summary

Introduction

The endoplasmic reticulum (ER) is recognized as an organelle that participates in the folding of secretory and membrane proteins [1,2]. ER stress has been shown to participate in the pathogenesis of a wide variety of cardiovascular diseases such as ischemia reperfusion heart disease [5,6], atherosclerosis [7], hypertension [8], myocardial infarction [9], heart failure [2], and its inhibition seems to be a promising therapeutic target. In response to ER stress, there is significant expression of ER chaperone such as glucose regulated protein 78 kD (GRP 78) [10]. When ER stress is severe and/or prolonged, apoptotic processes are initiated by transcriptional induction of C/EBP homologous protein (CHOP), or the phosphorylation of JNK, and/or caspase-12–dependent pathways [2]. NO production from eNOS has been shown to protect cardiomyocytes from apoptosis [11,12]

Methods

Results

Conclusion