Inhibition of the mitochondrial unfolded protein response by acetylcholine alleviated hypoxia/reoxygenation-induced apoptosis of endothelial cells.
The mitochondrial unfolded protein response (UPR(mt)) is involved in numerous diseases that have the common feature of mitochondrial dysfunction. However, its pathophysiological relevance in the context of hypoxia/reoxygenation (H/R) in endothelial cells remains elusive. Previous studies have demonstrated that acetylcholine (ACh) protects against cardiomyocyte injury by suppressing generation of mitochondrial reactive oxygen species (mtROS). This study aimed to explore the role of UPR(mt) in endothelial cells during H/R and to clarify the beneficial effects of ACh. Our results demonstrated that H/R triggered UPR(mt) in endothelial cells, as evidenced by the elevation of heat shock protein 60 and LON protease 1 protein levels, and resulted in release of mitochondrial pro-apoptotic proteins, including cytochrome C, Omi/high temperature requirement protein A 2 and second mitochondrial activator of caspases/direct inhibitor of apoptosis-binding protein with low PI, from the mitochondria to cytosol. ACh administration markedly decreased UPR(mt) by inhibiting mtROS and alleviating the mitonuclear protein imbalance. Consequently, ACh alleviated the release of pro-apoptotic proteins and restored mitochondrial ultrastructure and function, thereby reducing the number of terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling (TUNEL)-positive cells. Intriguingly, 4-diphenylacetoxy-N-methylpiperidine methiodide, a type-3 muscarinic ACh receptor (M3AChR) inhibitor, abolished the ACh-elicited attenuation of UPR(mt) and TUNEL positive cells, indicating that the salutary effects of ACh were likely mediated by M3AChR in endothelial cells. In conclusion, our studies demonstrated that UPR(mt) might be essential for triggering the mitochondrion-associated apoptotic pathway during H/R. ACh markedly suppressed UPR(mt) by inhibiting mtROS and alleviating the mitonuclear protein imbalance, presumably through M3AChR.
a Department of Pharmacology , School of Basic Medical Sciences , Xian Jiaotong University Health Science Center , Xi'an , P.R. China.
Man
M
Xu
a Department of Pharmacology , School of Basic Medical Sciences , Xian Jiaotong University Health Science Center , Xi'an , P.R. China.
Xueyuan
X
Bi
a Department of Pharmacology , School of Basic Medical Sciences , Xian Jiaotong University Health Science Center , Xi'an , P.R. China.
Xi
X
He
a Department of Pharmacology , School of Basic Medical Sciences , Xian Jiaotong University Health Science Center , Xi'an , P.R. China.
Xiaojiang
X
Yu
a Department of Pharmacology , School of Basic Medical Sciences , Xian Jiaotong University Health Science Center , Xi'an , P.R. China.
Ming
M
Zhao
a Department of Pharmacology , School of Basic Medical Sciences , Xian Jiaotong University Health Science Center , Xi'an , P.R. China.
Weijin
W
Zang
D049069:ATP-Dependent Proteases / Q000378:metabolism; D000109:Acetylcholine / Q000494:pharmacology*; D000111:Acetylcysteine / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D015687:Cell Hypoxia; D018834:Chaperonin 60 / Q000378:metabolism; D045304:Cytochromes c / Q000378:metabolism; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D018613:Microscopy, Confocal; D046529:Microscopy, Electron, Transmission; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D043587:Receptor, Muscarinic M3 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D056811:Unfolded Protein Response / Q000187:drug effects*; D015854:Up-Regulation / Q000187:drug effects
D018834:Chaperonin 60; D024101:Mitochondrial Proteins; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; D043587:Receptor, Muscarinic M3; D045304:Cytochromes c; D049069:ATP-Dependent Proteases; C000595973:LONP1 protein, human; D000109:Acetylcholine; D000111:Acetylcysteine
M3AChR; acetylcholine; apoptosis; endothelial cells; ischemia/reperfusion injury; mitochondrial reactive oxygen species; mitochondrial unfolded protein response