title: Anti-apoptotic activity of porcine cFLIP in ovarian granulosa cell lines.
abstract: In mammalian ovaries, more than 99% of follicles undergo atresia during growth and development. Recently, we found that the expression of cellular FLICE-like inhibitory protein long form (cFLIP(L)) decreased during follicular atresia in granulosa cells of porcine ovaries. In humans and other species, both the short (cFLIP(S)) and long (cFLIP(L)) forms of cFLIP are considered to function as cell survival factors that inhibit death ligand receptor-mediated apoptosis. Since the anti-apoptotic activity of porcine cFLIP (pcFLIP) in granulosa cells had not been determined, we examined the effect of pcFLIP on survival using granulosa-derived cell lines. A human cervix adenocarcinoma cell line, HeLa, human ovarian granulosa tumor cell line, KGN, and porcine granulosa-derived cell line, JC-410, were used. By Western blotting, internal cFLIP(L) was detected in all cell lines, but only trace levels of cFLIP(S) were found in HeLa and KGN cells. To examine the anti-apoptotic activity, pcFLIP(S) or pcFLIP(L) was overexpressed in HeLa and KGN cells. Transfected cells in which pcFLIP(S) or pcFLIP(L) was overexpressed, survived the induction of Fas-mediated apoptosis, while almost all of the cells transfected with empty vector died. Then, we suppressed the expression of porcine cFLIP(S) and/or cFLIP(L) in JC-410 cells using small interfering RNA (siRNA). When both cFLIP(S) and cFLIP(L), or only cFLIP(L) was suppressed, cell viability declined significantly. From the results, we conclude that porcine cFLIP(S) and cFLIP(L) exhibit anti-apoptotic activity in granulosa-derived cells. It was strongly suggested that pcFLIP acts as a survival-promoting factor in granulosa cells and determines whether porcine ovarian follicles survive or undergo atresia.
authors.affiliation: Research Unit for Animal Life Sciences, Animal Resource Science Center, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Ibaraki-Kasama, Japan.
authors.forename: Fuko
authors.initials: F
authors.lastname: Matsuda-Minehata
authors.forename: Yasufumi
authors.initials: Y
authors.lastname: Goto
authors.forename: Naoko
authors.initials: N
authors.lastname: Inoue
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Sakamaki
authors.forename: P Jorge
authors.initials: PJ
authors.lastname: Chedrese
authors.forename: Noboru
authors.initials: N
authors.lastname: Manabe
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000235:genetics; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein / Q000235:genetics / Q000502:physiology*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000235:genetics; D005260:Female; D005496:Follicular Atresia; D006107:Granulosa Cells / Q000502:physiology*; D006801:Humans; D010053:Ovary / Q000166:cytology; D034421:Sus scrofa / Q000502:physiology*; D014162:Transfection
chemical_list: D053446:CASP8 and FADD-Like Apoptosis Regulating Protein