Inhibition of the mitochondrial unfolded protein response by acetylcholine alleviated hypoxia/reoxygenation-induced apoptosis of endothelial cells

Abstract

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.

The heartbreak of depression: ‘Psycho-cardiac’ coupling in myocardial infarction

“…Interestingly, more recent work in cultured cell models of hypoxia/reoxygenation injury supports beneficial effects of acetylcholine on autophagy [150], and mitochondrial biogenesis [151] and quality control via mitophagy [152]. Similarly, vagal nerve stimulation may limit myocardial injury by inhibiting mitochondrial dysfunction and associated apoptosis [139], though this study found protection was limited to intra-ischemic intervention while post-ischemic stimulation was Additionally, M3 agonism with choline is cardioprotective [146,156], and the M3 receptor is implicated in the anti-apoptotic effects of acetylcholine [154] and in vagally mediated protection [133]. Finally, nicotinic receptors may also participate, with the study of Calvillo et al supporting involvement of the nicotinic pathway and inflammatory modulation in cardioprotection via vagal stimulation [137].…”

“…Other work supports vagal control of mitochondrial dynamics, likely promoting stresstolerance and limiting cell death[153]. Inhibition of the mitochondrial unfolded protein response may additionally contribute to anti-apoptotic effects of acetylcholine, potentially via inhibition of mitochondrial oxidant generation[154]. Studies thus collectively reveal important roles for vagal activity in dynamic maintenance of mitochondrial phenotype and function.…”

Mitophagy and Mitochondrial Quality Control Mechanisms in the Heart

“…Little information about the impact of URPmt is described in the context of heart diseases. Acute ischemia/hypoxia-reperfusion induces UPRmt in C. elegans [39] or endothelial cells [40] and can be a protective response. However, loss of the mitochondrial aspartyl tRNA-synthetase encoding gene DARS2 in mice results in dysregulation of mitochondrial translation and strong cardiomyopathy associated with an increase of UPRmt markers [41].…”

Pharmacological Modulation of Vagal Nerve Activity in Cardiovascular Diseases

“…Studies in our laboratory have demonstrated that ACh prevents angiotensin II-induced cardiomyocyte injury through down-regulation of the angiotensin II type 1 receptor and inhibition of ROS-mediated p38 MAPK activation, as well as regulation of Bcl-2, Bax, and caspase-3 expression [ 48 , 79 – 81 ]. Recently, we demonstrated that ACh markedly suppresses the mitochondrial unfolded protein response and endoplasmic reticulum stress, which alleviate I/R injury [ 82 , 83 ]. ACh may also modulate the abnormal inter-organelle crosstalk in cardiovascular diseases.…”