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A GLP-1 analog lowers ER stress and enhances protein folding to ameliorate homocysteine-induced endothelial dysfunction


Hyperhomocysteinemia (HHcy) is an independent risk factor for cardiovascular diseases and increases mortality in type 2 diabetic patients. HHcy induces endoplasmic reticulum (ER) stress and oxidative stress to impair endothelial function. The glucagon-like peptide 1 (GLP-1) analog exendin-4 attenuates endothelial ER stress, but the detailed vasoprotective mechanism remains elusive. The present study investigated the beneficial effects of exendin-4 against HHcy-induced endothelial dysfunction. Exendin-4 pretreatment reversed homocysteine-induced impairment of endothelium-dependent relaxations in C57BL/6 mouse aortae ex vivo. Four weeks subcutaneous injection of exendin-4 restored the impaired endothelial function in both aortae and mesenteric arteries isolated from mice with diet-induced HHcy. Exendin-4 treatment lowered superoxide anion accumulation in the mouse aortae both ex vivo and in vivo. Exendin-4 decreased the expression of ER stress markers (e.g., ATF4, spliced XBP1, and phosphorylated eIF2α) in human umbilical vein endothelial cells (HUVECs), and this change was reversed by cotreatment with compound C (CC) (AMPK inhibitor). Exendin-4 induced phosphorylation of AMPK and endothelial nitric oxide synthase in HUVECs and arteries. Exendin-4 increased the expression of endoplasmic reticulum oxidoreductase (ERO1α), an important ER chaperone in endothelial cells, and this effect was mediated by AMPK activation. Experiments using siRNA-mediated knockdown or adenoviral overexpression revealed that ERO1α mediated the inhibitory effects of exendin-4 on ER stress and superoxide anion production, thus ameliorating HHcy-induced endothelial dysfunction. The present results demonstrate that exendin-4 reduces HHcy-induced ER stress and improves endothelial function through AMPK-dependent ERO1α upregulation in endothelial cells and arteries. AMPK activation promotes the protein folding machinery in endothelial cells to suppress ER stress.

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Fig. 1: Vascular function of mouse aortae upon acute Hcy exposure.
Fig. 2: Vascular function of the conduit and small arteries of the mice with diet-induced HHcy.
Fig. 3: ROS level of mouse aortae ex vivo and in vivo.
Fig. 4: Ex4-activated AMPK and eNOS in HUVECs and aortae from the mice with diet-induced HHcy.
Fig. 5: ER stress-induced oxidative stress in endothelial cells.
Fig. 6: ER stress and ER chaperone of endothelial cells.
Fig. 7: siRNA-mediated ERO1α knockdown in HUVECs.
Fig. 8: ERO1α overexpression in HUVECs and mouse aortae.
Fig. 9: The endothelial benefits mediated by exendin-4.


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This work was supported by Health and Medical Research Fund [Grant numbers 05162906 and 05161746], Early Career Scheme [Grant number 24122318], the National Natural Science Foundation of China [Grant numbers 91739103 and 91939302], and Hong Kong Research Grants Council [Grant numbers 14112919 and C4024-16W].

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CKC designed the study, performed experiments, analyzed the data, and wrote the manuscript. CWL and JYL performed some of the experiments. WCsC, CFN, and RCWM were involved in regular discussion and revised the manuscript. YH and XYT designed the study, analyzed the data, wrote the manuscript, and provided grant support.

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Correspondence to Xiao Yu Tian or Yu Huang.

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The authors declare no competing interests.

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Cheng, C.K., Luo, JY., Lau, C.W. et al. A GLP-1 analog lowers ER stress and enhances protein folding to ameliorate homocysteine-induced endothelial dysfunction. Acta Pharmacol Sin (2021).

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  • homocysteine
  • GLP-1 analog
  • exendin-4
  • AMPK
  • ER stress
  • ER chaperone
  • oxidative stress
  • endothelial dysfunction


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