Effect of Captopril on Ca^(2+) Homeostasis and Cell Viability in Human Hepatoma Cells

I-Shu Chen ； Chiang-Ting Chou ； Wei-Zhe Liang ； Yuan-Yuarn Liu ； Chun-Chi Kuo ； Jue-Long Wang ； Lyh-Jyh Hao ； Chung-Ren Jan

《The Chinese Journal of Physiology》 61卷4期 (2018/08) Pp. 221-229

https://doi.org/10.4077/CJP.2018.BAH594

摘要

Captopril, an angiotensin-converting enzyme (ACE) inhibitor, induced different Ca^(2+) signaling responses in various cell models. However, the effect of captopril on Ca^(2+) homeostasis and cell viability in hepatoma cells is unknown. This study examined whether captopril altered Ca^(2+) homeostasis and viability in HepG2 human hepatoma cells. Intracellular Ca^(2+) concentrations in suspended cells were monitored by using the fluorescent Ca^(2+)-sensitive dye fura-2. Cell viability was examined by using 4-[3-[4-lodophenyl]-2-4(4-nitrophenyl)-2H-5-tetrazolio-1,3-benzene disulfonate] water soluble tetrazolium-1 (WST-1). Captopril at concentrations of 500-3000 µM induced cytosolic free concentrations of Ca^(2+) ([Ca^(2+)]_i) rises in a concentration-dependent manner. Ca^(2+) removal reduced the signal by approximately 15%. Mn^(2+) has been shown to enter cells through similar mechanisms as Ca^(2+) but quenches fura-2 fluorescence at all excitation wavelengths. Captopril (3000 μM)-induced Mn^(2+) influx indirectly suggested that captopril evoked Ca^(2+) entry. Captopril-induced Ca^(2+) entry was inhibited by 15% by a protein kinase C (PKC) activator (phorbol 12-myristate 13 acetate, PMA) and an inhibitor (GF109203X) and three inhibitors of store-operated Ca^(2+) channels: nifedipine, econazole and SKF96365. In Ca^(2+)-free medium, treatment with the endoplasmic reticulum Ca^(2+) pump inhibitor 2,5-di-tert-butylhydroquinone (BHQ) abolished captopril-evoked [Ca^(2+)]_i rises. Conversely, treatment with captopril abolished BHQ-evoked [Ca^(2+)]_i rises. Inhibition of phospholipase C (PLC) with U73122 inhibited 70% of captopril-induced [Ca^(2+)]_i rises. Captopril at concentrations between 150- 550 μM killed cells in a concentration-dependent fashion. Chelation of cytosolic Ca^(2+) with 1,2-bis(2- aminophenoxy)ethane-N,N,N',N'-tetraacetic acid/acetoxy methyl (BAPTA/AM) did not reverse captopril's cytotoxicity. Together, in HepG2 human hepatoma cells, captopril induced [Ca^(2+)]_i rises and caused cell death that was not triggered by preceding [Ca^(2+)]_i rises.

關鍵字

Ca^(2+) ； captopril ； endoplasmic reticulum ； human hepatoma cells ； viability

參考文獻

Andersson, P., Bratt, J., Heimbürger, M., Cederholm, T. and Palmblad, J. Inhibition of neutrophil-dependent cytotoxicity for human endothelial cells by ACE inhibitors. Scand. J. Immunol. 80: 339-345, 2014.

Attoub, S., Gaben, A.M., Al-Salam, S., Al Sultan, M.A., John, A., Nicholls, M.G., Mester, J. and Petroianu, G. Captopril as a potential inhibitor of lung tumor growth and metastasis. Ann. N. Y. Acad. Sci. 1138: 65-72, 2008.

Berridge, M.J. Calcium microdomains: organization and function. Cell Calcium 40: 405-412, 2006.

Bootman, M.D., Lipp, P. and Berridge, M.J. The organization and functions of local Ca2+ signals. J. Cell Sci. 114: 2213-2222, 2001.

Cao, X., Yang, F., Shi, T., Yuan, M., Xin, Z., Xie, R., Li, S., Li, H. and Yang, J.K. Angiotensin-converting enzyme 2/angiotensin-(1-7)/Mas axis activates Akt signaling to ameliorate hepatic steatosis. Sci. Rep. 6: 21592, 2016.