Abstract
Current evidence indicates that coronary microcirculation is a key target for protecting against cardiac ischemia–reperfusion (I/R) injury. Mitochondrial calcium uniporter (MCU) complex activation and mitochondrial calcium ([Ca2+]m) overload are underlying mechanisms involved in cardiovascular disease. Histidine triad nucleotide-binding 2 (HINT2) has been reported to modulate [Ca2+]m via the MCU complex, and our previous work demonstrated that HINT2 improved cardiomyocyte survival and preserved heart function in mice with cardiac ischemia. This study aimed to explore the benefits of HINT2 on cardiac microcirculation in I/R injury with a focus on mitochondria, the MCU complex, and [Ca2+]m overload in endothelial cells. The present work demonstrated that HINT2 overexpression significantly reduced the no-reflow area and improved microvascular perfusion in I/R-injured mouse hearts, potentially by promoting endothelial nitric oxide synthase (eNOS) expression and phosphorylation. Microvascular barrier function was compromised by reperfusion injury, but was repaired by HINT2 overexpression via inhibiting VE-Cadherin phosphorylation at Tyr731 and enhancing the VE-Cadherin/β-Catenin interaction. In addition, HINT2 overexpression inhibited the inflammatory response by suppressing vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1). Mitochondrial fission occurred in cardiac microvascular endothelial cells (CMECs) subjected to oxygen–glucose deprivation/reoxygenation (OGD/R) injury and resulted in mitochondrial dysfunction and mitochondrion-dependent apoptosis, the effects of which were largely relieved by HINT2 overexpression. Additional experiments confirmed that [Ca2+]m overload was an initiating factor for mitochondrial fission and that HINT2 suppressed [Ca2+]m overload via modulation of the MCU complex through directly interacting with MCU in CMECs. Regaining [Ca2+]m overload by spermine, an MCU agonist, abolished all the protective effects of HINT2 on OGD/R-injured CMECs and I/R-injured cardiac microcirculation. In conclusion, the present report demonstrated that HINT2 overexpression inhibited MCU complex-mitochondrial calcium overload-mitochondrial fission and apoptosis pathway, and thereby attenuated cardiac microvascular ischemia–reperfusion injury.
Highlights
The reperfusion strategy is the most widely acknowledged treatment for acute myocardial infarction (MI) [22]
Fluorescence staining of Flag indicated the transfection efficiency of AAV9 was > 95% in the microcirculation (Fig. S1a), and Histidine triad nucleotide-binding 2 (HINT2) signal was higher in the Flag-positive area in AAV9-HINT2-infected hearts (Fig. S1b)
HINT2 was not transfected into cardiomyocytes, as no obvious colocalization between Flag and cTnT was observed (Fig. S1a), and the expression of HINT2 was equal in cardiomyocytes isolated from the AAV9-negative control (NC) and AAV9-HINT2 groups (Fig. S1d)
Summary
The reperfusion strategy is the most widely acknowledged treatment for acute myocardial infarction (MI) [22]. The mechanisms of I/R injury to coronary microvascular impairment range from microembolization, impaired. Basic Research in Cardiology (2021) 116:65 vasomotion, capillary plugging and rupture, to leukocyte infiltration, which are partly different from those contributing to cardiomyocyte injury [18, 19]. These pathological changes further progress to cardiac edema and even coronary no-reflow phenomena, thereby discounting the benefits of reperfusion therapy [18]. Considering that coronary microvascular dysfunction has been recognized as a determinant for patients’ adverse outcomes, it is highly necessary to better understand the underlying mechanisms and search for effective therapeutic targets [8, 22]
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