Roflumilast Reduces Myocardial Ischemia Reperfusion Injury In vivo and In vitro by Activating the AMPK Signaling Pathway

Overview

Journal Exp Ther Med

Specialty Pathology

Date 2023 May 25

PMID 37229319

Authors

Bo Liao

Zhongyuan Han

Affiliations

Soon will be listed here.

Abstract

Myocardial tissue cell damage induced by myocardial ischemia/reperfusion (MI/R) notably elevates the mortality rate, increases the complications of patients with myocardial infarction and decreases reperfusion benefit in patients suffering from acute myocardial infarction. Roflumilast protect against cardiotoxicity. Therefore, the present study aimed to investigate the effect of roflumilast on MI/R injury and the underlying mechanisms. To simulate MI/R and , the rat model of MI/R was established and H9C2 cells were subjected to hypoxia/reoxygenation (H/R) induction, respectively. The myocardial infarction areas were observed by 2,3,5-triphenyltetrazolium chloride staining. The myocardial enzyme levels in serum and levels of inflammatory cytokines and oxidative stress markers in cardiac tissue were assessed by corresponding assay kits. The cardiac damage was observed by hematoxylin and eosin staining. The mitochondrial membrane potential in cardiac tissue and H9C2 cells was detected using the JC-1 staining kit. The viability and apoptosis of H9C2 cells were detected by Cell Counting Kit-8 and TUNEL assay, respectively. The levels of inflammatory cytokines, oxidative stress markers and ATP in H/R-induced H9C2 cells were analyzed by corresponding assay kits. Western blotting was used for the estimation of AMP-activated protein kinase (AMPK) signaling pathway-, apoptosis- and mitochondrial regulation-associated protein levels. The mPTP opening was detected using a calcein-loading/cobalt chloride-quenching system. The results indicated that roflumilast decreased MI/R-induced myocardial infarction by alleviating myocardial injury and mitochondrial damage through the activation of the AMPK signaling pathway. In addition, roflumilast mitigated viability damage, alleviated oxidative stress, attenuated the inflammatory response and decreased mitochondrial damage in H/R-induced H9C2 cells by activating the AMPK signaling pathway. However, compound C, an inhibitor of the AMPK signaling pathway, reversed the effect of roflumilast on H/R-induced H9C2 cells. In conclusion, roflumilast alleviated myocardial infarction in MI/R rats and attenuated H/R-induced oxidative stress, inflammatory response and mitochondrial damage in H9C2 cells by activating the AMPK signaling pathway.

References

Kosutova P, Mikolka P, Kolomaznik M, Balentova S, Adamkov M, Calkovska A . Reduction of lung inflammation, oxidative stress and apoptosis by the PDE4 inhibitor roflumilast in experimental model of acute lung injury. Physiol Res. 2019; 67(Suppl 4):S645-S654. DOI: 10.33549/physiolres.934047. View

Weinberger T, Schulz C . Myocardial infarction: a critical role of macrophages in cardiac remodeling. Front Physiol. 2015; 6:107. PMC: 4387471. DOI: 10.3389/fphys.2015.00107. View

Yeh C, Chen T, Wang Y, Lin Y, Fang S . AMP-activated protein kinase activation during cardioplegia-induced hypoxia/reoxygenation injury attenuates cardiomyocytic apoptosis via reduction of endoplasmic reticulum stress. Mediators Inflamm. 2011; 2010:130636. PMC: 3034973. DOI: 10.1155/2010/130636. View

Grahame Hardie D . AMPK: positive and negative regulation, and its role in whole-body energy homeostasis. Curr Opin Cell Biol. 2014; 33:1-7. DOI: 10.1016/j.ceb.2014.09.004. View

Ong S, Hausenloy D . Mitochondrial Dynamics as a Therapeutic Target for Treating Cardiac Diseases. Handb Exp Pharmacol. 2016; 240:251-279. DOI: 10.1007/164_2016_7. View

Farah A, Barbagelata A . Unmet goals in the treatment of Acute Myocardial Infarction: Review. F1000Res. 2017; 6. PMC: 5532798. DOI: 10.12688/f1000research.10553.1. View

Ansari M, Ganaie M, Ur Rehman N, Alharthy K, Khan T, Imam F . Protective role of Roflumilast against cadmium-induced cardiotoxicity through inhibition of oxidative stress and NF-κB signaling in rats. Saudi Pharm J. 2019; 27(5):673-681. PMC: 6598217. DOI: 10.1016/j.jsps.2019.04.002. View

Xu W, Zhang J, Xiao J . Roflumilast Suppresses Adipogenic Differentiation AMPK Mediated Pathway. Front Endocrinol (Lausanne). 2021; 12:662451. PMC: 8215703. DOI: 10.3389/fendo.2021.662451. View

Xu X, Liao L, Hu B, Jiang H, Tan M . Roflumilast, a Phosphodiesterases-4 (PDE4) Inhibitor, Alleviates Sepsis‑induced Acute Kidney Injury. Med Sci Monit. 2020; 26:e921319. PMC: 7268890. DOI: 10.12659/MSM.921319. View

10.

Kyung S, Kim Y, Son E, Jeong S, Park J . The Phosphodiesterase 4 Inhibitor Roflumilast Protects against Cigarette Smoke Extract-Induced Mitophagy-Dependent Cell Death in Epithelial Cells. Tuberc Respir Dis (Seoul). 2018; 81(2):138-147. PMC: 5874143. DOI: 10.4046/trd.2017.0115. View

11.

Kim A, Miller E, Wright T, Li J, Qi D, Atsina K . A small molecule AMPK activator protects the heart against ischemia-reperfusion injury. J Mol Cell Cardiol. 2011; 51(1):24-32. PMC: 4005884. DOI: 10.1016/j.yjmcc.2011.03.003. View

12.

Tahrir F, Langford D, Amini S, Mohseni Ahooyi T, Khalili K . Mitochondrial quality control in cardiac cells: Mechanisms and role in cardiac cell injury and disease. J Cell Physiol. 2018; 234(6):8122-8133. PMC: 6395499. DOI: 10.1002/jcp.27597. View

13.

Zhou H, Zhang Y, Hu S, Shi C, Zhu P, Ma Q . Melatonin protects cardiac microvasculature against ischemia/reperfusion injury via suppression of mitochondrial fission-VDAC1-HK2-mPTP-mitophagy axis. J Pineal Res. 2017; 63(1). PMC: 5518188. DOI: 10.1111/jpi.12413. View

14.

Janjua S, Fortescue R, Poole P . Phosphodiesterase-4 inhibitors for chronic obstructive pulmonary disease. Cochrane Database Syst Rev. 2020; 5:CD002309. PMC: 7193764. DOI: 10.1002/14651858.CD002309.pub6. View

15.

Tian L, Cao W, Yue R, Yuan Y, Guo X, Qin D . Pretreatment with Tilianin improves mitochondrial energy metabolism and oxidative stress in rats with myocardial ischemia/reperfusion injury via AMPK/SIRT1/PGC-1 alpha signaling pathway. J Pharmacol Sci. 2019; 139(4):352-360. DOI: 10.1016/j.jphs.2019.02.008. View

16.

Perrelli M, Pagliaro P, Penna C . Ischemia/reperfusion injury and cardioprotective mechanisms: Role of mitochondria and reactive oxygen species. World J Cardiol. 2011; 3(6):186-200. PMC: 3139040. DOI: 10.4330/wjc.v3.i6.186. View

17.

Wang X, Zhang J, Li L, Zhao X . Beneficial effects of metformin on primary cardiomyocytes via activation of adenosine monophosphate-activated protein kinase. Chin Med J (Engl). 2011; 124(12):1876-84. View

18.

Xu B, Xu J, Cai N, Li M, Liu L, Qin Y . Roflumilast prevents ischemic stroke-induced neuronal damage by restricting GSK3β-mediated oxidative stress and IRE1α/TRAF2/JNK pathway. Free Radic Biol Med. 2020; 163:281-296. DOI: 10.1016/j.freeradbiomed.2020.12.018. View

19.

Li Y, Liu X . Novel insights into the role of mitochondrial fusion and fission in cardiomyocyte apoptosis induced by ischemia/reperfusion. J Cell Physiol. 2018; 233(8):5589-5597. DOI: 10.1002/jcp.26522. View

20.

Sasaki H, Asanuma H, Fujita M, Takahama H, Wakeno M, Ito S . Metformin prevents progression of heart failure in dogs: role of AMP-activated protein kinase. Circulation. 2009; 119(19):2568-77. DOI: 10.1161/CIRCULATIONAHA.108.798561. View