Ketoconazole-evoked [Ca2+]i rises and non-Ca2+-triggered cell death in rabbit corneal epithelial cells (SIRC).
The effect of ketoconazole on cytosolic free Ca2+ concentrations ([Ca2+]i) and proliferation has not been explored in corneal cells. This study examined whether ketoconazole alters Ca2+ levels and causes cell death in SIRC rabbit corneal epithelial cells. [Ca2+]i and cell viability were measured by using the fluorescent dyes fura-2 and WST-1, respectively. Ketoconazole at concentrations of 5 microM and above increased [Ca2+]i in a concentration-dependent manner. The Ca2+ signal was reduced partly by removing extracellular Ca2+. The ketoconazole-induced Ca2+ influx was insensitive to L-type Ca2+ channel blockers and protein kinase C modulators. In Ca2+-free medium, after pretreatment with 50 microM ketoconazole, thapsigargin-(1 microM)-induced [Ca2+]i rises were abolished; conversely, thapsigargin pretreatment nearly abolished ketoconazole-induced [Ca2+]i rises. Inhibition of phospholipase C with 2 microM U73122 did not change ketoconazole-induced [Ca2+]i rises. At concentrations between 5 and 100 microM, ketoconazole killed cells in a concentration-dependent manner. The cytotoxic effect of 50 microM ketoconazole was not reversed by prechelating cytosolic Ca2+ with BAPTA. In summary, in corneal cells, ketoconazole-induced [Ca2+]i rises by causing Ca2+ release from the endoplasmic reticulum and Ca2+ influx from unknown pathways. Furthermore, the cytotoxicity induced by ketoconazole was not caused via a preceding [Ca2+]i rise.
Lin
Muh-Chiou
MC
Department of Ophthalmology, Kaohsiung Veterans General Hospital, Kaohsiung, Taiwan.
Huang
Chorng-Chih
CC
Huang
Chun-Jen
CJ
Cheng
He-Hsiung
HH
Chou
Chiang-Ting
CT
Wang
Jue-Long
JL
Chen
I-Shu
IS
Liu
Shiuh-Inn
SI
Lu
Yih-Chau
YC
Chang
Hong-Tai
HT
Huang
Jong-Khing
JK
Chen
Jin-Shyr
JS
Jan
Chung-Ren
CR
D000818:Animals; D000935:Antifungal Agents / Q000494:pharmacology; D002118:Calcium / Q000737:chemistry / Q000378:metabolism*; D016923:Cell Death; D049109:Cell Proliferation; D002470:Cell Survival; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000378:metabolism; D007654:Ketoconazole / Q000494:pharmacology*; D008956:Models, Chemical; D011493:Protein Kinase C / Q000378:metabolism; D011817:Rabbits; D013997:Time Factors
D000935:Antifungal Agents; D011493:Protein Kinase C; D007654:Ketoconazole; D002118:Calcium