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Glucagon-like peptide-1 attenuates endoplasmic reticulum stress–induced apoptosis in H9c2 cardiomyocytes during hypoxia/reoxygenation through the GLP-1R/PI3K/Akt pathways

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Abstract

Endoplasmic reticulum (ER) stress–induced apoptosis is a major cause of myocardial ischemia/reperfusion (I/R) injury. Emerging evidence indicates that glucagon-like peptide-1 (GLP-1) has potential cardioprotective effects. However, the precise mechanisms underlying the involvement of GLP-1 in I/R injury remain largely unknown. In the present study, we aimed to determine whether GLP-1 attenuates hypoxia/reoxygenation (H/R) injury in cardiomyocytes and to further elucidate the underlying signaling pathway. The results indicate that GLP-1 reversed the increased apoptotic ratio, the increased lactate dehydrogenase (LDH) levels, the reduced cell viability, the increased Caspase-3 activity, and the increased Bax/Bcl-2 ratio caused by H/R. Importantly, GLP-1 significantly decreased the expression of H/R-induced ER stress proteins (GRP78, CHOP) and Caspase-12. In addition, we found that GLP-1 increased the expression of p-Akt in H9c2 cells with H/R injuries, and that the protective action of GLP-1 against H/R-induced injury was blocked by the GLP-1 receptor (GLP-1R) inhibitor Exendin9-39 and the PI3K inhibitor LY294002. Exendin9-39 and LY294002 also blocked the downregulation of ER stress protein expression by GLP-1, after H/R injury. Therefore, we have shown that GLP-1 exerts its cardioprotective effects by alleviating ER stress–induced apoptosis due to H/R injury and that these effects are most likely associated with the activation of GLP-1R/PI3K/Akt signaling pathway.

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Acknowledgments

We thank professor Lei Yang (Basic Medical Science College, Jiujiang University, Jiujiang, Jiangxi Province, China) for providing intellectual content of critical importance to the work described.

Funding

This research study was funded by the National Natural Science Foundation of China (Grant No. 81660152) and the Natural Science Foundation of Jiangxi Province (Grant No.20181BAB205004).

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Author notes

  1. Gaopeng Guan and Jun Zhang contributed equally to this work.

Authors and Affiliations

  1. Department of Cardiology, Affiliated Hospital of Jiujiang University, Jiujiang, 332000, Jiangxi Province, China

    Gaopeng Guan, Jun Zhang, Shengyuan Liu & Xiang Gu

  2. Key Laboratory of System Bio-medicine of Jiangxi Province, Jiujiang University, Jiujiang, 332000, Jiangxi, China

    Gaopeng Guan, Shengyuan Liu, Wenyin Huang, Ying Gong & Xiang Gu

  3. The First Affiliated Hospital of Nanchang University, Nanchang, 330006, Jiangxi Province, China

    Gaopeng Guan

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  1. Gaopeng Guan
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Contributions

X.G. designed the study. G.G., J.Z., and S.L. performed the experiments and collected the data. W.H. and Y.G. analyzed and interpreted the experimental data. X.G. and G.G. prepared the manuscript.

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Correspondence to Xiang Gu.

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Supplementary Fig. S1

Determination of the cytotoxicity of H/R to H9c2 cells over various time intervals. The cells were treated with different durations of hypoxia, followed by reoxygenation for 6 h, and then processed for the CCK8 assay. Statistical analysis is shown on the bar graphs. Data are presented as the mean ± SEM of three independent experiments. *P < 0.05 versus control. (PNG 13 kb)

High Resolution Image (TIF 47 kb)

Supplementary Fig. S2

GLP-1R is expressed in H9c2 cells. GLP-1R expression was measured by western blot analysis in normal H9c2 cells. (PNG 26 kb)

High Resolution Image (TIF 82 kb)

Supplementary Fig. S3

Exendin9–39 abolished the effects of GLP-1 on cAMP levels, phosphorylation of CREB, and phosphorylation of ERK1/2 in H/R-treated H9c2 cells. H9c2 cells were treated with Exendin9–39 before GLP-1 treatment in H/R-injury and then processed for the cAMP levels assay (A). Western blot analysis of p-CREB, CREB, p-ERK1/2, and ERK1/2 in H/R-injured H9c2 cells treated with Exendin9–39 before GLP-1 treatment (B). The analysis of band intensities is presented as the relative ratio of p-CREB to CREB and the relative ratio of p-ERK1/2 to ERK1/2 (C). G: GLP-1; E: Exendin9-39. Statistical analysis is shown on the bar graphs. Data are presented as the mean ± SEM of three independent experiments. *P < 0.05 versus control, #P < 0.05 versus H/R, &P < 0.05 versus H/R+GLP-1. (PNG 126 kb)

High Resolution Image (TIF 319 kb)

Supplementary Table S1

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Guan, G., Zhang, J., Liu, S. et al. Glucagon-like peptide-1 attenuates endoplasmic reticulum stress–induced apoptosis in H9c2 cardiomyocytes during hypoxia/reoxygenation through the GLP-1R/PI3K/Akt pathways. Naunyn-Schmiedeberg's Arch Pharmacol 392, 715–722 (2019). https://doi.org/10.1007/s00210-019-01625-2

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