LGR4 silence aggravates ischemic injury by modulating mitochondrial function and oxidative stress via ERK signaling pathway in H9c2 cells

Abstract

It is reported that LGR4 (leucine-rich repeat domain containing G protein-coupled receptor 4) plays a crucial role in the physiological function of many organs. However, few data are available on the function and mechanism of LGR4 in myocardial ischemia–reperfusion (I/R) injury. The aim of this study was to explore the function and mechanism of LGR4 in I/R injury. We incubated H9c2 cells in simulating ischemia buffer and then re-incubated them in normal culture medium to establish a model of I/R injury in vitro. The expression of LGR4 was evaluated by RT-PCR and western blot. Besides, the cell apoptosis was evaluated by flow cytometric analysis and the content of ROS, SOD, MDA, LDH, CK, ATP, cyt c were detected by special commercial kits. The expression of mitochondrial function-related proteins were detected by western blot. Then, the roles of ERK signaling pathway was determined with TBHQ (ERK activator) treatment. Our data have demonstrated that I/R boosted the expression of LGR4 in H9c2 cells. Knockdown of LGR4 increased the apoptosis rate of H9c2 cells and led to excessed oxidant stress and impaired mitochondrial function by increasing the levels of ROS, MDA, LDH, CK and cyt c and inhibiting SOD activity, ATP production. In addition, LGR4 silence inhibited the activation of ERK pathway. And TBHQ partially reversed the effects of LGR4 knockdown on H9c2 cells. To conclude, our study indicated that LGR4 regulated mitochondrial dysfunction and oxidative stress by ERK signaling pathways, which provides a potential cardiac protective target against I/R.

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Data availability

The data used to support the findings of this study are available from the corresponding author upon request.

Abbreviations

MI:

Myocardial ischemia

I/R:

Ischemia/reperfusion

LGR4:

Leucine-rich repeat domain containing G protein-coupled receptor 4

GPCR48:

G-protein-coupled receptor-48

DMEM:

Dulbecco’s modified Eagle’s medium

FBS:

Fetal bovine serum

ISCH:

Ischemic heart disease

NF:

Non-failing hearts

ROS:

Reactive oxygen species

MAPKs:

Mitogen-activated protein kinases

ERK:

Extracellular regulated protein kinases

mRNA:

Messenger RNA

MTT:

Methylthiazolyldiphenyl-tetrazolium bromide

SDS-PAGE:

Sodium dodecyl sulphate polyacrylamide gel electrophoresis

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Funding

This study was supported by the National Natural Science Foundation of China(81870330).

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Contributions

Tao Chen and Xiaozhen Zhuo designed the study, analyzed and interpreted the data. Tao Chen, Xiangrui Qiao, Lele Cheng, Mengping Liu and Yangyang Deng conducted the experiments and drafted the manuscript. Tao Chen and Xiangrui Qiao reviewed the literature. Xiaozhen Zhuo revised the manuscript.

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Correspondence to Xiaozhen Zhuo.

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Chen, T., Qiao, X., Cheng, L. et al. LGR4 silence aggravates ischemic injury by modulating mitochondrial function and oxidative stress via ERK signaling pathway in H9c2 cells. J Mol Histol (2021). https://doi.org/10.1007/s10735-021-09957-1

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Keywords

  • LGR4
  • Ischemic myocardial injury
  • Mitochondrial function
  • Oxidative stress
  • ERK signaling pathway