Pinacidil ameliorates cardiac microvascular ischemia-reperfusion injury by inhibiting chaperone-mediated autophagy of calreticulin

  • Basic Res Cardiol. 2024 Jan 2. doi: 10.1007/s00395-023-01028-8.
Muyin Liu  #  1  2  3 ,  Su Li  #  1  2  3 ,  Ming Yin  #  1  2  3 ,  Youran Li  1  2  3 ,  Jinxiang Chen  1  2  3 ,  Yuqiong Chen  4 ,  You Zhou  1  2  3 ,  Qiyu Li  1  2  3 ,  Fei Xu  5 ,  Chunfeng Dai  1 ,  Yan Xia  1  2  3 ,  Ao Chen  1  2  3 ,  Danbo Lu  1  2  3 ,  Zhangwei Chen  6  7  8 ,  Juying Qian  9  10  11 ,  Junbo Ge  12  13  14
Affiliations
  • 1. Department of Cardiology, Zhongshan Hospital, Fudan University, Shanghai, 200032, China.
  • 2. Shanghai Institute of Cardiovascular Diseases, Shanghai, 200032, China.
  • 3. National Clinical Research Center for Interventional Medicine, Shanghai, 200032, China.
  • 4. Gusu School, The Affiliated Suzhou Hospital of Nanjing Medical University, Suzhou Municipal Hospital, Nanjing Medical University, Nanjing, China.
  • 5. Department of Cardiology, West China Hospital, Sichuan University, Chengdu, People's Republic of China.
  • 6. Department of Cardiology, Zhongshan Hospital, Fudan University, Shanghai, 200032, China. [email protected].
  • 7. Shanghai Institute of Cardiovascular Diseases, Shanghai, 200032, China. [email protected].
  • 8. National Clinical Research Center for Interventional Medicine, Shanghai, 200032, China. [email protected].
  • 9. Department of Cardiology, Zhongshan Hospital, Fudan University, Shanghai, 200032, China. [email protected].
  • 10. Shanghai Institute of Cardiovascular Diseases, Shanghai, 200032, China. [email protected].
  • 11. National Clinical Research Center for Interventional Medicine, Shanghai, 200032, China. [email protected].
  • 12. Department of Cardiology, Zhongshan Hospital, Fudan University, Shanghai, 200032, China. [email protected].
  • 13. Shanghai Institute of Cardiovascular Diseases, Shanghai, 200032, China. [email protected].
  • 14. National Clinical Research Center for Interventional Medicine, Shanghai, 200032, China. [email protected].
  • # Contributed equally.
PMID: 38168863 DOI: 10.1007/s00395-023-01028-8
Abstract

Calcium overload is the key trigger in cardiac microvascular ischemia-reperfusion (I/R) injury, and calreticulin (CRT) is a calcium buffering protein located in the endoplasmic reticulum (ER). Additionally, the role of pinacidil, an antihypertensive drug, in protecting cardiac microcirculation against I/R injury has not been investigated. Hence, this study aimed to explore the benefits of pinacidil on cardiac microvascular I/R injury with a focus on endothelial calcium homeostasis and CRT signaling. Cardiac vascular perfusion and no-reflow area were assessed using FITC-lectin perfusion assay and Thioflavin-S staining. Endothelial calcium homeostasis, CRT-IP3Rs-MCU signaling expression, and Apoptosis were assessed by real-time calcium signal reporter GCaMP8, western blotting, and fluorescence staining. Drug affinity-responsive target stability (DARTS) assay was adopted to detect proteins that directly bind to pinacidil. The present study found pinacidil treatment improved capillary density and perfusion, reduced no-reflow and infraction areas, and improved cardiac function and hemodynamics after I/R injury. These benefits were attributed to the ability of pinacidil to alleviate calcium overload and mitochondria-dependent Apoptosis in cardiac microvascular endothelial cells (CMECs). Moreover, the DARTS assay showed that pinacidil directly binds to HSP90, through which it inhibits chaperone-mediated Autophagy (CMA) degradation of CRT. CRT overexpression inhibited IP3Rs and MCU expression, reduced mitochondrial calcium inflow and mitochondrial injury, and suppressed endothelial Apoptosis. Importantly, endothelial-specific overexpression of CRT shared similar benefits with pinacidil on cardiovascular protection against I/R injury. In conclusion, our data indicate that pinacidil attenuated microvascular I/R injury potentially through improving CRT degradation and endothelial calcium overload.

Keywords
Calcium overload; Calreticulin; Cardiac microvascular ischemia–reperfusion injury; Chaperone-mediated autophagy; Mitochondrial injury; Pinacidil.
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