Estrogen-dependent expression of the cystic fibrosis transmembrane regulator gene in a novel uterine epithelial cell line.
We have demonstrated previously the modulation of CFTR expression by estrogen in vivo in the rat uterine epithelium. The purpose of this study was to establish a suitable in vitro system to investigate the regulation of CFTR by steroid hormones. Primary cultures of rat uterine epithelial cells, which showed high levels of CFTR expression in vitro, were infected with an adeno/SV40 virus. One clone, UIT 1.16, which retained the morphology of the primary epithelial cells yet proliferated beyond the life span of the primary culture, was isolated and characterized. Successful immortalization of UIT 1.16 cells was verified by the presence of a band corresponding to the SV40 large T-antigen in western blots, as well as by their ability to proliferate continuously. Transmission electron microscopy studies revealed that these cells maintained the characteristics of a polarized epithelium with well-established membrane domains and specialized intercellular junctions. A high transepithelial electrical resistance was also observed when cells were assayed in modified Ussing chambers. When the basolateral cellular membrane of cells grown in vitrogen-coated filters was permeabilized with nystatin, a forskolin-stimulated Cl- permeability was observed in the apical membrane, similar to that present in other CFTR-expressing epithelial cells. UIT 1.16 cells showed high levels of CFTR expression on northern blots. The expression of CFTR was dependent on the presence of estrogen in the culture medium, since almost undetectable levels of CFTR mRNA were observed when the cells were cultured in medium containing serum depleted of steroid hormones. However, addition of estrogen to this medium prevented the disappearance of CFTR mRNA, confirming estrogen-regulated expression of CFTR in the UIT 1.16 cell line. The newly developed UIT 1.16 cell line provides a valuable model to analyze the regulation of CFTR expression by steroid hormones. Moreover, the cell line could also be used to investigate the role of CFTR in the uterus during the normal female cycle as well as for the study of other uterine epithelial functions and the agents that regulate them.
Rochwerger
L
L
Research Institute, Hospital for Sick Children, Toronto, Ontario, Canada.
Dho
S
S
Parker
L
L
Foskett
J K
JK
Buchwald
M
M
D000256:Adenoviridae / Q000502:physiology; D000818:Animals; D002461:Cell Line, Transformed; D016764:Cell Polarity; D002472:Cell Transformation, Viral; D002712:Chlorides / Q000378:metabolism; D005576:Colforsin / Q000494:pharmacology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D004717:Endometrium / Q000166:cytology*; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism; D004958:Estradiol / Q000494:pharmacology*; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects*; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D017208:Rats, Wistar; D013539:Simian virus 40 / Q000502:physiology
D002712:Chlorides; D008565:Membrane Proteins; D012333:RNA, Messenger; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D005576:Colforsin; D004958:Estradiol