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Hypoxic preconditioning protects microvascular endothelial cells against hypoxia/reoxygenation injury by attenuating endoplasmic reticulum stress

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Abstract

Endothelial cells (ECs) are directly exposed to hypoxia and contribute to injury during myocardial ischemia/reperfusion. Hypoxic preconditioning (HPC) protects ECs against hypoxia injury. This study aimed to explore whether HPC attenuates hypoxia/reoxygenation (H/R) injury by suppressing excessive endoplasmic reticulum stress (ERS) in cultured microvascular ECs (MVECs) from rat heart. MVECs injury was measured by lactate dehydrogenase (LDH) leakage, cytoskeleton destruction, and apoptosis. Expression of glucose regulating protein 78 (GRP78) and C/EBP homologous protein (CHOP), activation of caspase-12 (pro-apoptosis factors) and phosphorylation of p38 mitogen-activated protein kinase (p38 MAPK) were detected by western blot analysis. HPC attenuated H/R-induced LDH leakage, cytoskeleton destruction, and cell apoptosis, as shown by flow cytometry, Bax/Bcl-2 ratio, caspase-3 activation and terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling. HPC suppressed H/R-induced ERS, as shown by a decrease in expression of GRP78 and CHOP, and caspase-12 activation. HPC enhanced p38 MAPK phosphorylation but decreased that of protein kinase R-like ER kinase (PERK, upstream regulator of CHOP). SB202190 (an inhibitor of p38 MAPK) abolished HPC-induced cytoprotection, downregulation of GRP78 and CHOP, and activation of caspase-12, as well as PERK phosphorylation. HPC may protect MVECs against H/R injury by suppressing CHOP-dependent apoptosis through p38 MAPK mediated downregulation of PERK activation.

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Abbreviations

CHOP:

CCAAT/enhancer-binding protein-homologous protein

ER:

Endoplasmic reticulum

ERS:

Endoplasmic reticulum stress

GRP78:

Glucose regulating protein 78

HPC:

Hypoxic preconditioning

H/R:

Hypoxia/reoxygenation

IPC:

Ischemic preconditioning

I/R:

Ischemia/reperfusion

LDH:

Lactate dehydrogenase

MVECs:

Microvascular endothelial cells

p38 MAPK:

p38 mitogen-activated protein kinase

PDIA3:

Protein disulfide isomerase associated 3

PERK:

Protein kinase R-like ER kinase

TG:

Thapsigargin

2-DE:

Two-dimensional gel electrophoresis

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Acknowledgments

This study was supported by the National Natural Science Foundation of China (No. 81070186) and a grant from the National Basic Research Program of China (No. 2011CB944004).

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Authors and Affiliations

  1. Department of Out-patient, Chinese PLA General Hospital, Beijing, 100853, China

    Xu-Dong Wu & Xiu-Qin Zhu

  2. Department of Pathophysiology, Chinese PLA General Hospital, Beijing, 100853, China

    Zhen-Ying Zhang, Sheng Sun, Yu-Zhen Li, Xiao-Reng Wang, Wei-Hong Li & Xiu-Hua Liu

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  1. Xu-Dong Wu
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Corresponding authors

Correspondence to Xu-Dong Wu or Xiu-Hua Liu.

Additional information

Xu-Dong Wu and Zhen-Ying Zhang contributed equally to this study.

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Wu, XD., Zhang, ZY., Sun, S. et al. Hypoxic preconditioning protects microvascular endothelial cells against hypoxia/reoxygenation injury by attenuating endoplasmic reticulum stress. Apoptosis 18, 85–98 (2013). https://doi.org/10.1007/s10495-012-0766-6

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  • DOI: https://doi.org/10.1007/s10495-012-0766-6

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