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Apoptotic events induced by high glucose in human hepatoma HepG2 cells involve endoplasmic reticulum stress and MAPK’s activation

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Abstract

To investigate whether endoplasmic reticulum (ER) stress participates in the induction of apoptosis in HepG2 cells exposed to high glucose and explore its probable mechanism. A series of experiments were performed following HepG2 cells treated with different concentrations of glucose for 48 h. The apoptosis was detected by means of Hoechst staining and flow cytometry. Caspase-3 activity assay was performed by measuring the pNA (p-nitroaniline) to indirectly reveal the catalytic activity of caspase-3. The expression levels of apoptosis-, ER stress-associated proteins and MAPKs were analyzed by western blot. To further characterize the molecular mechanisms, the effects of antioxidant alpha-lipoic acid (ALA) and specific inhibitors for JNK and p38 (SP600125 and SB203580, respectively) were examined by Hoechst staining, immunofluorescence, and western blot. After HepG2 cells were incubated with high glucose for 48 h, both Hoechst staining and flow cytometry analyses unveiled the apoptosis of HepG2 cells. Caspase-3 activity assay revealed that the activity of caspase-3 was enhanced. Western blot showed an enhancement of pro-caspase-9 degradation, a reduction of Bcl-2/Bax ratio, a decrease in GRP78 expression, and increases in CHOP and p47/phox levels. In addition, western blot analysis presented that phosphorylation of p38 and JNK was triggered and that the expression of ASK1 was elevated. In the case of the contributions of oxidative stress and the MAPK signaling pathways, all ALA, SP600125 and SB203580 were able to largely rescue high glucose-induced apoptosis. High glucose induced the apoptosis in HepG2 cells through the activation of ASK1-p38/JNK pathway mediated by ER stress and oxidative stress.

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Acknowledgements

This study was supported by the National Natural Science Foundation of China (No. 81470568).

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Authors and Affiliations

  1. Laboratory of Molecular Biology and Department of Biochemistry, Anhui Medical University, Hefei, 230032, Anhui, People’s Republic of China

    Qiaoling Jiang, Yujun Yuan, Jiali Zhou, Yan Wu, Qing Zhou & Yuan Wang

  2. Key Laboratory of Gene Research of Anhui Province, Hefei, 230032, Anhui, People’s Republic of China

    Qing Zhou, Shuyu Gui & Yuan Wang

  3. Clinical Laboratory, Affiliated Hospital of Jiujiang University, Jiujiang, 332000, Jiangxi, People’s Republic of China

    Yujun Yuan

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  1. Qiaoling Jiang
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  2. Yujun Yuan
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  3. Jiali Zhou
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  4. Yan Wu
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  5. Qing Zhou
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  6. Shuyu Gui
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  7. Yuan Wang
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Correspondence to Yuan Wang.

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Qiaoling Jiang and Yujun Yuan contributed equally to this work.

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Jiang, Q., Yuan, Y., Zhou, J. et al. Apoptotic events induced by high glucose in human hepatoma HepG2 cells involve endoplasmic reticulum stress and MAPK’s activation. Mol Cell Biochem 399, 113–122 (2015). https://doi.org/10.1007/s11010-014-2238-5

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  • DOI: https://doi.org/10.1007/s11010-014-2238-5

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