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Salvianolic acid A attenuates TNF-α- and d-GalN-induced ER stress-mediated and mitochondrial-dependent apoptosis by modulating Bax/Bcl-2 ratio and calcium release in hepatocyte LO2 cells

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Abstract

Salvianolic acid (Sal A) is a water-soluble compound extracted from Radix Salvia miltiorrhiza (danshen), which has been widely used to treat acute hepatitis and hepatic damage in traditional Chinese medicine. The aim of the present study was to delineate the antiapoptotic signaling pathways involved in Sal A’s hepato-protective action in hepatocyte LO2 cells and to further elucidate the mechanism by which Sal A elicits the antiapoptotic effects on hepatocytes. Here, the study showed that Sal A had antiapoptotic effects on the TNF-α/d-GalN-treated LO2 cells. Moreover, Western blotting demonstrated that the levels of p-eIF2α, ATF4, GRP78, CHOP and caspase-4 were markedly decreased in Sal A group. Additionally, the decrease of the cell mitochondrial membrane permeability and increase of ΔΨm were detected in Sal A-treated cells by high-content screening (HCS) analysis. And the levels of cleaved-caspase-9, cleaved-caspase-3, apoptosis-inducing factor (AIF), Apaf-1, and Cytc (cyto) were downregulated, while Cytc (mito) was upregulated by Sal A via Western blotting. Furthermore, the decreased levels of Bax/Bcl-2 ratio and calcium release were measured in Sal A-treated cells. In summary, Sal A attenuates TNF-α- and d-GalN-induced both ER stress and mitochondrial-dependent apoptosis by suppression of Bax/Bcl-2 ratio and prevention of calcium release, which support the notion that Sal A could be developed into a novel hepatic protectant.

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Abbreviations

ER:

Endoplasmic reticulum

UPR:

Unfolded protein response

TNF-α:

Tumor necrosis factor alpha

d-GalN:

d-Galactosamine

TRAIL:

TNF-related apoptosis-inducing ligand

GRP78:

ER chaperone 78-kDa glucose-regulated protein

IRE1α:

Inositol-requiring enzyme 1α

PERK:

PKR-like ER kinase

ATF6:

Activating transcription factor 6

eIF2α:

Eukaryotic initiation factor 2α

CHOP:

C/EBP homologous protein

GADD153:

Growth-arrest and DNA damage-inducible gene 153

CCK-8:

Cell counting kit-8

FITC:

Fluorescein isothiocyanate

PI:

Propidium iodide

ΔΨm:

Mitochondrial membrane potential

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Acknowledgments

This work was supported by grants from National Natural Sciences Foundation of China (81072777, 81273638). This work was also financially supported in part by the grants from a project funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD). In addition, this program was granted for scientific innovation research of college graduate in Jiangsu province (CXZZ13_0624).

Author contributions

Xiao-jing Yan, Ze-qun Jiang, and Wei-ping Chen designed research; Xiao-jing Yan, Ze-qun Jiang, Lei Bi, and Ye Yang performed research; Xiao-jing Yan wrote this manuscript; Ze-qun Jiang and Wei-ping Chen reviewed and edited this manuscript. All authors read and approved the final manuscript.

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The authors declare no conflict of interest.

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    1. Department of Preclinical Medicine, Nanjing University of Chinese Medicine, 138 Xianlin Rd., Nanjing, 210023, China

      Xiaojing Yan, Zequn Jiang, Lei Bi, Ye Yang & Weiping Chen

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    1. Xiaojing Yan
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    Xiaojing Yan and Zequn Jiang contributed equally to this work.

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    Yan, X., Jiang, Z., Bi, L. et al. Salvianolic acid A attenuates TNF-α- and d-GalN-induced ER stress-mediated and mitochondrial-dependent apoptosis by modulating Bax/Bcl-2 ratio and calcium release in hepatocyte LO2 cells. Naunyn-Schmiedeberg's Arch Pharmacol 388, 817–830 (2015). https://doi.org/10.1007/s00210-015-1116-3

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