Diethylstilbestrol-induced estrogen receptor-dependent [Ca2+]i rises and apoptosis in Chinese hamster ovary (CHO) cells.
The effect of the synthetic estrogen diethylstilbestrol (DES) on cytosolic free Ca2+ concentrations ([Ca2+]i) and cell viability was explored in Chinese hamster ovary (CHO-K1). [Ca2+]i and cell viability were measured by using the fluorescent dyes fura-2 and WST-1, respectively. DES at concentrations>or=1 proportional, variant increased [Ca2+]i in a concentration-dependent manner. The Ca2+ signal was reduced partly by removing extracellular Ca2+. In Ca2+-free medium, after pretreatment with 50 proportional, variant DES, 1 proportional, variant thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor)-induced [Ca2+]i rises were abolished. Conversely, thapsigargin pretreatment abolished DES-induced [Ca2+]i rises. Inhibition of phospholipase C with U73122 did not alter DES-induced [Ca2+]i rises. At a concentration of 5 proportional, variant, DES increased cell viability. At concentrations of 100-200 microM, DES decreased viability in a concentration-dependent manner. The effect of 5 and 100 microM DES on viability was partly reversed by prechelating cytosolic Ca2+ with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N' -tetraacetic acid (BAPTA). DES-induced cell death was induced via apoptosis as demonstrated by propidium iodide staining. DES (100 microM)-induced [Ca2+]i rises were largely inhibited by pretreatment with the estrogen receptor antagonist ICI-182,780 (100 microM). ICI-182,780 did not affect 5 microM DES-induced increase in viability but partly reversed 100 microM DES-induced cell death. Collectively, in CHO-K1 cells, DES induced [Ca2+]i rises by stimulating estrogen receptors leading to Ca2+ release from the endoplasmic reticulum in a phospholipase C-independent manner, and Ca2+ influx. DES-caused cytotoxicity was mediated by an estrogen receptor- and Ca2+-dependent pathway.
Roan
Cherng-Jau
CJ
Department of Gynecology/Obstetrics, Ping Tung Christian Hospital, Pingtung, Taiwan.
Huang
Chorng-Chih
CC
Cheng
He-Hsiung
HH
Chien
Jau-Min
JM
Chou
Chiang-Ting
CT
Lin
Ko-Long
KL
Liu
Shiuh-Inn
SI
Lu
Yih-Chau
YC
Chang
Hong-Tai
HT
Huang
Jong-Khing
JK
Jan
Chung-Ren
CR
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D006224:Cricetinae; D003412:Cricetulus; D004054:Diethylstilbestrol / Q000494:pharmacology*; D004958:Estradiol / Q000031:analogs & derivatives / Q000494:pharmacology; D004963:Estrenes / Q000494:pharmacology; D000077267:Fulvestrant; D016257:Fura-2 / Q000378:metabolism; D042541:Intracellular Space / Q000187:drug effects / Q000378:metabolism; D011760:Pyrrolidinones / Q000494:pharmacology; D011960:Receptors, Estrogen / Q000378:metabolism*; D010738:Type C Phospholipases / Q000378:metabolism
D004963:Estrenes; D011760:Pyrrolidinones; D011960:Receptors, Estrogen; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; D000077267:Fulvestrant; D004958:Estradiol; D004054:Diethylstilbestrol; D010738:Type C Phospholipases; D002118:Calcium; D016257:Fura-2