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Saturated Free Fatty Acid Sodium Palmitate-Induced Lipoapoptosis by Targeting Glycogen Synthase Kinase-3β Activation in Human Liver Cells

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Abstract

Background

Elevated serum saturated fatty acid levels and hepatocyte lipoapoptosis are features of nonalcoholic fatty liver disease (NAFLD).

Aim

The purpose of this study was to investigate saturated fatty acid induction of lipoapoptosis in human liver cells and the underlying mechanisms.

Methods

Human liver L02 and HepG2 cells were treated with sodium palmitate, a saturated fatty acid, for up to 48 h with or without lithium chloride, a glycogen synthase kinase-3β (GSK-3β) inhibitor, or GSK-3β shRNA transfection. Transmission electron microscopy was used to detect morphological changes, flow cytometry was used to detect apoptosis, a colorimetric assay was used to detect caspase-3 activity, and western blot analysis was used to detect protein expression.

Results

The data showed that sodium palmitate was able to induce lipoapoptosis in L02 and HepG2 cells. Western blot analysis showed that sodium palmitate activated GSK-3β protein, which was indicated by dephosphorylation of GSK-3β at Ser-9. However, inhibition of GSK-3β activity with lithium chloride treatment or knockdown of GSK-3β expression with shRNA suppressed sodium palmitate-induced lipoapoptosis in L02 and HepG2 cells. On a molecular level, inhibition of GSK-3β expression or activity suppressed sodium palmitate-induced c-Jun-N-terminal kinase (JNK) phosphorylation and Bax upregulation, whereas GSK-3β inhibition did not affect endoplasmic reticulum stress-induced activation of unfolded protein response.

Conclusions

The present data demonstrated that saturated fatty acid sodium palmitate-induced lipoapoptosis in human liver L02 and HepG2 cells was regulated by GSK-3β activation, which led to JNK activation and Bax upregulation. This finding indicates that GSK-3β inhibition may be a potential therapeutic target to control NAFLD.

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Abbreviations

NAFLD:

Nonalcoholic fatty liver disease

NASH:

Nonalcoholic steatohepatitis

FFA:

Free fatty acid

ER:

Endoplasmic reticulum

UPR:

Unfolded protein response

PERK:

Protein kinase RNA-like ER kinase

IRE1:

Inositol-requiring protein 1

GRP78:

Glucose-regulated protein 78

GSK-3:

Glycogen synthase kinase-3

JNK:

c-Jun-N-terminal kinase

Bax:

Bcl-2-associated X protein

BSA:

Bovine serum albumin

7-AAD:

7-amino actinomycin D

ATF6:

Activating transcription factor 6

ATF4:

Activating transcription factor 4

CHOP:

C/EBP-homologous protein

XBP-1:

X-box-binding protein 1

shRNA:

Short hairpin RNA

PI3K:

Phosphatidylinositide-3-OH kinase

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Acknowledgments

We sincerely thank the staff of the Department of Gastroenterology and Hepatology at the Second Affiliated Hospital of Chongqing Medical University for their constant and unselfish help. We thank Ms. Yingxia Xiang and Mr. Jing Wang for their consistent and illuminating technical assistance. We also thank Medjaden Bioscience, Hong Kong, China for assistance in preparation of this manuscript. This study was supported in part by grants from the Natural Science Foundation of China (#81270494 and #30871160) and the Project of Medical Science and Technology of Chongqing (#2012-1-033).

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Authors and Affiliations

  1. Department of Gastroenterology and Hepatology, The 2nd Affiliated Hospital of Chongqing Medical University, No. 74 Linjiang Road, Chongqing, 400010, China

    Jie Cao, Xiao-Xia Feng, Bo Ning, Zhao-Xia Yang & Wei Shen

  2. Department of Intensive-Care Medicine, The Central Hospital of Jiangjin, Chongqing, China

    Long Yao

  3. Department of Gastroenterology and Hepatology, The Yongchuan Hospital, Chongqing Medical University, Chongqing, China

    Dian-Liang Fang

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  1. Jie Cao
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Correspondence to Wei Shen.

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Cao, J., Feng, XX., Yao, L. et al. Saturated Free Fatty Acid Sodium Palmitate-Induced Lipoapoptosis by Targeting Glycogen Synthase Kinase-3β Activation in Human Liver Cells. Dig Dis Sci 59, 346–357 (2014). https://doi.org/10.1007/s10620-013-2896-2

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