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Pretreatment with cilnidipine attenuates hypoxia/reoxygenation injury in HL-1 cardiomyocytes through enhanced NO production and action potential shortening

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Abstract

Myocardial ischemia/reperfusion injury worsens in the absence of nitric oxide synthase (NOS). Cilnidipine, a Ca2+ channel blocker, has been reported to activate endothelial NOS (eNOS) and increases nitric oxide (NO) in vascular endothelial cells. We examined whether pretreatment with cilnidipine could attenuate cardiac cell deaths including apoptosis caused by hypoxia/reoxygenation (H/R) injury. HL-1 mouse atrial myocytes as well as H9c2 rat ventricular cells were exposed to H/R, and cell viability was evaluated by an autoanalyzer and flow cytometry; eNOS expression, NO production, and electrophysiological properties were also evaluated by western blotting, colorimetry, and patch clamping, respectively, in the absence and presence of cilnidipine. Cilnidipine enhanced phosphorylation of eNOS and NO production in a concentration-dependent manner, which was abolished by siRNAs against eNOS or an Hsp90 inhibitor, geldanamycin. Pretreatment with cilnidipine attenuated cell deaths including apoptosis during H/R; this effect was reproduced by an NO donor and a xanthine oxidase inhibitor. The NOS inhibitor L-NAME abolished the protective action of cilnidipine. Pretreatment with cilnidipine also attenuated H9c2 cell death during H/R. Additional cilnidipine treatment during H/R did not significantly enhance its protective action. There was no significant difference in the protective effect of cilnidipine under normal and high Ca2+ conditions. Action potential duration (APD) of HL-1 cells was shortened by cilnidipine, with this shortening augmented after H/R. L-NAME attenuated the APD shortening caused by cilnidipine. These findings indicate that cilnidipine enhances NO production, shortens APD in part by L-type Ca2+ channel block, and thereby prevents HL-1 cell deaths during H/R.

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References

  1. Nabel EG, Braunwald E. A tale of coronary artery disease and myocardial infarction. N Engl J Med. 2012;366:54–63.

    CAS  Google Scholar 

  2. Spath NB, Mills NL, Cruden NL. Novel cardioprotective and regenerative therapies in acute myocardial infarction: a review of recent and ongoing clinical trials. Future Cardiol. 2016;12:655–72.

    CAS  Google Scholar 

  3. Hausenloy DJ, Botker HE, Engstrom T, Erlinge D, Heusch G, Ibanez B, et al. Targeting reperfusion injury in patients with ST-segment elevation myocardial infarction: trials and tribulations. Eur Heart J. 2017;38:935–41.

    CAS  Google Scholar 

  4. Prieto-Moure B, Lloris-Carsí JM, Barrios-Pitarque C, Toledo-Pereyra LH, Lajara-Romance JM, Berda-Antolí M, et al. Pharmacology of Ischemia Reperfusion. Translational research considerations. J Investig Surg. 2016;29:234–49.

    Google Scholar 

  5. Murry CE, Jennings RB, Reimer KA. Preconditioning with ischemia: a delay of lethal cell injury in ischemic myocardium. Circulation. 1986;74:1124–36.

    CAS  Google Scholar 

  6. Kunst G, Klein AA. Peri-operative anaesthetic myocardial preconditioning and protection - cellular mechanisms and clinical relevance in cardiac anaesthesia. Anaesthesia. 2015;70:467–82.

    CAS  Google Scholar 

  7. Schulz R, Kelm M, Heusch G. Nitric oxide in myocardial ischemia/reperfusion injury. Cardiovasc Res. 2004;61:402–13.

    CAS  Google Scholar 

  8. Cohen MV, Yang XM, Downey JM. Nitric oxide is a preconditioning mimetic and cardioprotectant and is the basis of many available infarct-sparing strategies. Cardiovasc Res. 2006;70:231–9.

    CAS  Google Scholar 

  9. Bice JS, Jones BR, Chamberlain GR, Baxter GF. Nitric oxide treatments as adjuncts to reperfusion in acute myocardial infarction: a systematic review of experimental and clinical studies. Basic Res Cardiol. 2016;111:23.

    Google Scholar 

  10. Arroyo-Martínez EA, Meaney A, Gutiérrez-Salmeán G, Rivera-Capello JM, GonzálezCoronado V, Alcocer-Chauvet A, et al. Is Local Nitric Oxide Availability Responsible for Myocardial Salvage after Remote Preconditioning? Arq Bras Cardiol. 2016;107:154–62.

    Google Scholar 

  11. Jones SP, Girod WG, Palazzo AJ, Granger DN, Grisham MB, Jourd’Heuil D, et al. Myocardial ischemia-reperfusion injury is exacerbated in absence of endothelial cell nitric oxide synthase. Am J Physiol. 1999;276:H1567–H1573.

    CAS  Google Scholar 

  12. Elrod JW, Greer JJ, Bryan NS, Langston W, Szot JF, Gebregzlabher H, et al. Cardiomyocyte-specific overexpression of NO synthase-3 protects against myocardial ischemia-reperfusion injury. Arterioscler Thromb Vasc Biol. 2006;26:1517–23.

    CAS  Google Scholar 

  13. Tominaga M, Ohya Y, Tsukashima A, Kobayashi K, Takata Y, Koga T, et al. Ambulatory blood pressure monitoring in patients with essential hypertension treated with a new calcium antagonist, cilnidipine. Cardiovasc Drugs Ther. 1997;11:43–48.

    CAS  Google Scholar 

  14. Kobayashi N, Mori Y, Mita S, Nakano S, Kobayashi T, Tsubokou Y, et al. Effects of cilnidipine on nitric oxide and endothelin-1 expression and extracellular signal-regulated kinase in hypertensive rats. Eur J Pharm. 2001;422:149–57.

    CAS  Google Scholar 

  15. Bahrudin U, Morisaki H, Morisaki T, Ninomiya H, Higaki K, Nanba E, et al. Ubiquitin-proteasome system impairment caused by a missense cardiac myosin-binding protein C mutation and associated with cardiac dysfunction in hypertrophic cardiomyopathy. J Mol Biol. 2008;384:896–907.

    CAS  Google Scholar 

  16. Tanno S, Yamamoto K, Kurata Y, Adachi M, Inoue Y, Otani N, et al. Protective effects of topiroxostat on an ischemia-reperfusion model of rat hearts. Circ J. 2018;82:1101–11.

    CAS  Google Scholar 

  17. Bahrudin U, Morikawa K, Takeuchi A, Kurata Y, Miake J, Mizuta E, et al. Impairment of ubiquitin-proteasome system by E334K cMyBPC modifies channel proteins, leading to electrophysiological dysfunction. J Mol Biol. 2011;413:857–78.

    CAS  Google Scholar 

  18. Courtemanche M, Ramirez RJ, Nattel S, Am J. Physiol. Ionic mechanisms underlying human atrial action potential properties: insights from a mathematical model. Am J Physiol. 1998;275:H301–H321. https://doi.org/10.1152/ajpheart.1998.275.1.H301

    Article  CAS  Google Scholar 

  19. Brutsaert DL. Cardiac endothelial-myocardial signaling: Its role in cardiac growth, contractile performance, and rhythmicity. Physiol Rev. 2003;83:59–115.

    CAS  Google Scholar 

  20. Boveris A, Costa LE, Poderoso JJ, Carreras MC, Cadenas E. Regulation of mitochondrial respiration by oxygen and nitric oxide. Ann N Y Acad Sci. 2000;899:121–35.

    CAS  Google Scholar 

  21. Ding Y, Vaziri ND. Calcium channel blockade enhances nitric oxide synthase expression by cultured endothelial cells. Hypertension. 1998;32:718–23.

    CAS  Google Scholar 

  22. Ishii M. Pharmacokinetics study of FRC-8653 (Cilnidipine). Jpn Pharm Ther. 1993;21:43–52.

    Google Scholar 

  23. Jiang J, Cyr D, Babbitt RW, Sessa WC, Patterson C. Chaperone-dependent regulation of endothelial nitric-oxide synthase intracellular trafficking by the cochaperone/ubiquitin ligase CHIP. J Biol Chem. 2003;278:49332–41.

    CAS  Google Scholar 

  24. Nedvetsky PI, Sessa WC, Schmidt HH. There’s NO binding like NOS binding: protein-protein interactions in NO/cGMP signaling. Proc Natl Acad Sci USA. 2002;99:16510–2.

    CAS  Google Scholar 

  25. Smolenski PA, Muller H, Kronich P, Kugler P, Walter U, Schnitzer JE, et al. Calcium-dependent membrane association sensitizes soluble guanylyl cyclase to nitric oxide. Nat Cell Biol. 2002;4:307–11.

    Google Scholar 

  26. Lee YM, Cheng PY, Chen SY, Chung MT, Sheu JR. Wogonin suppresses arrhythmias, inflammatory responses, and apoptosis induced by myocardial ischemia/reperfusion in rats. J Cardiovasc Pharm. 2011;58:133–42.

    CAS  Google Scholar 

  27. Li J, Zhang H, Zhang C. Role of inflammation in the regulation of coronary blood flow in ischemia and reperfusion: Mechanisms and therapeutic implications. J Mol Cell Cardiol. 2012;52:865–72.

    CAS  Google Scholar 

  28. Gottlieb RA. Cell death pathways in acute ischemia/reperfusion injury. J Cardiovasc Pharm Ther. 2011;16:233–8.

    CAS  Google Scholar 

  29. Koenitzer JR, Freeman BA. Redox signaling in inflammation: Interactions of endogenous electrophiles and mitochondria in cardiovascular disease. Ann N Y Acad Sci. 2010;1203:45–52.

    CAS  Google Scholar 

  30. Szewczyk A, Jarmuszkiewicz W, Koziel A, Sobieraj I, Nobik W, Lukasiak A, et al. Mitochondrial mechanisms of endothelial dysfunction. Pharm Rep. 2015;67:704–10.

    CAS  Google Scholar 

  31. Zhang P, Lu Y, Yu D, Zhang D, Hu W. Trap1 provides protection against myocardial ischemia-reperfusion injury by ameliorating mitochondrial dysfunction. Cell Physiol Biochem. 2015;36:2072–82.

    CAS  Google Scholar 

  32. Pike MM, Luo CS, Clark MD, Kirk KA, Kitakaze M, Madden MC, et al. Nmr measurements of na+ and cellular energy in ischemic rat heart: Role of na(+)-h+ exchange. Am J Physiol. 1993;265:2017–26.

    Google Scholar 

  33. Smith MA, Schnellmann RG. Calpains, mitochondria, and apoptosis. Cardiovasc Res. 2012;96:32–37.

    CAS  Google Scholar 

  34. Zhang JY, Wu F, Gu XM, Jin ZX, Kong LH, Zhang Y, et al. The blockade of transmembrane cl(-) flux mitigates i/r-induced heart injury via the inhibition of calpain activity. Cell Physiol Biochem. 2015;35:2121–34.

    CAS  Google Scholar 

  35. Zorov DB, Juhaszova M, Sollott SJ. Mitochondrial ros-induced ros release: an update and review. Biochim Biophys Acta. 2006;1757:509–17.

    CAS  Google Scholar 

  36. Endemann DH, Schiffrin EL. Endothelial dysfunction. J Am Soc Nephrol. 2004;15:1983–92.

    CAS  Google Scholar 

  37. Cheng O, Ostrowski RP, Wu B, Liu W, Chen C, Zhang JH. Cyclooxygenase-2 mediates hyperbaric oxygen preconditioning in the rat model of transient global cerebral ischemia. Stroke. 2011;42:484–90.

    CAS  Google Scholar 

  38. Thandroyen FT, McCarthy J, Burton KP, Opie LH. Ryanodine and caffeine prevent ventricular arrhythmias during acute myocardial ischemia and reperfusion in rat heart. Circ Res. 1988;62:306–14.

    CAS  Google Scholar 

  39. Zhao CY, Greenstein JL, Winslow RL. Mechanisms of the cyclic nucleotide cross-talk signaling network in cardiac L-type calcium channel regulation. J Mol Cell Cardiol. 2017;106:29–44.

    CAS  Google Scholar 

  40. Takahara A, Nakamura Y, Wagatsuma H, Aritomi S, Nakayama A, Satoh Y, et al. Long-term blockade of L/N-type Ca(2+) channels by cilnidipine ameliorates repolarization abnormality of the canine hypertrophied heart. Br J Pharm. 2009;158:1366–74.

    CAS  Google Scholar 

  41. Hayashi T, Yamaguchi T, Sakakibara Y, Taguchi K, Maeda M, Kuzuya M, et al. eNOS-dependent antisenscence effect of a calcium channel blocker in human endothelial cells. PLoS ONE. 2014;9:e88391.

    Google Scholar 

  42. Matsubara M, Yao K, Hasegawa K. Benidipine, a dihydropyridine-calcium channel blocker, inhibits lysophosphatidylcholine-induced endothelial injury via stimulation of nitric oxide release. Pharm Res. 2006;53:35–43.

    CAS  Google Scholar 

  43. Fan L, Yang Q, Xiao XQ, Grove KL, Huang Y, Chen ZW, et al. Dual actions of cilnidipine in human internal thoracic artery: inhibition of calcium channels and enhancement of endothelial nitric oxide synthase. J Thorac Cardiovasc Surg. 2011;141:1063–9.

    CAS  Google Scholar 

  44. Batova S, DeWever J, Godfraind T, Balligand JL, Dessy C, Feron O. The calcium channel blocker amlodipine promotes the unclamping of eNOS from caveolin in endothelial cells. Cardiovasc Res. 2006;71:478–85.

    CAS  Google Scholar 

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Acknowledgements

EA Pharma. Co. (Tokyo, Japan) and Fuji yakuhin Co. (Saitama, Japan) kindly provided cilnidipine and topiroxostat, respectively. Daiichi Sankyo Co. kindly provided azelnidipine.

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Authors and Affiliations

  1. Department of Anesthesiology and Critical Care Medicine, Tottori University Faculty of Medicine, 86 Nishi-cho, Yonago, 683-8503, Japan

    Hiroyuki Minato, Ryo Endo, Akihiro Otsuki & Yoshimi Inagaki

  2. Department of Genetic Medicine and Regenerative Therapeutics, Institute of Regenerative Medicine and Biofunction, Tottori University Graduate School of Medical Science, 86 Nishi-cho, Yonago, 683-8503, Japan

    Ichiro Hisatome, Tomomi Notsu, Motokazu Tsuneto & Yasuaki Shirayoshi

  3. Department of Physiology II, Kanazawa Medical University, Ishikawa, 920-0268, Japan

    Yasutaka Kurata

  4. Department of Biological Regulation, Tottori University, Yonago, 683-8503, Japan

    Naoe Nakasone & Haruaki Ninomiya

  5. Department of Community-Based Family Medicine, Tottori University Faculty of Medicine, Yonago, 683-8503, Japan

    Toshihiro Hamada

  6. Department of Molecular Medicine and Therapeutics, Tottori University Faculty of Medicine, Yonago, 683-8503, Japan

    Takuya Tomomori & Akihiro Okamura

  7. Department of Pharmacology, Tottori University Faculty of Medicine, Yonago, 683-8503, Japan

    Junichiro Miake

  8. Division of Pathological Biochemistry, Tottori University Faculty of Medicine, Yonago, 683-8503, Japan

    Futoshi Okada

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  1. Hiroyuki Minato
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Correspondence to Yasutaka Kurata.

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Conflict of interest

IH reported receiving lecturer’s fees from Mochida Pharmaceutical Company, Sanwa Kagaku Kenkyusho Co. Ltd, PFeizer Co. Ltd. and Fuji Yakuhin Co. Ltd., and research grants from EA Pharma. Co. Teijin Pharma, Fuji Yakuhin Co. Ltd. and Sanwa Kagaku Kenkyusho Co. Ltd.

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Minato, H., Hisatome, I., Kurata, Y. et al. Pretreatment with cilnidipine attenuates hypoxia/reoxygenation injury in HL-1 cardiomyocytes through enhanced NO production and action potential shortening. Hypertens Res 43, 380–388 (2020). https://doi.org/10.1038/s41440-019-0391-7

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