title: Gonadotropin-releasing hormone-agonist induces apoptosis of human granulosa-luteal cells via caspase-8, -9 and -3, and poly-(ADP-ribose)-polymerase cleavage.
abstract: Gonadotropin-releasing hormone-agonist (GnRH-Ag) used in controlled ovarian hyperstimulation (COH) for in vitro fertilization and embryo transfer (IVF-ET) has been known to directly affect apoptosis of human ovarian cells, but its mechanism is not clearly understood. Therefore, the purpose of the present study was to investigate whether caspase-8, -9, and -3 activation and poly-(ADP-ribose)-polymerase (PARP) cleavage are involved in the mechanism by which GnRH-Ag induces apoptosis in human granulosa-luteal cells. The prospective study was conducted in the research institute and clinical fertility center of university hospital. Human granulosa-luteal cells collected from IVF-ET patients were cultured and treated with 10-6 M GnRH-Ag or saline as a control. To access apoptosis in the cells, terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-digoxigenin nick end-labeling (TUNEL) assay and DNA fragmentation analysis were preformed 24 h after treatment. Activity of caspase-8, -9, and -3 in the cells was examined using a fluorogenic substrate. Caspase-8, -9, and -3 activation and PARP cleavage were analyzed by Western blot. DNA fragmentation in the cells increased at concentrations over 10-6M GnRH-Ag. In TUNEL assays, the rate of apoptotic cell formation in GnRH-Ag treatment increased significantly compared with that of saline treatment (p < 0.05). The activity of caspase-8, -9 and, -3 investigated using a fluorogenic substrate increased only in the apoptotic cells. In Western blot analysis, cells treated with GnRH-Ag revealed an increase in active forms of caspase-8, -9, and -3 compared with saline treatment. In addition, cleavage of PARP also increased in cells treated with GnRH-Ag. These results suggest that activation of caspase-8, -9, and -3 and cleavage of PARP might be involved in apoptosis of human granulosa-luteal cells induced by GnRH-Ag.
authors.affiliation: Department of Obstetrics and Gynecology, Eulji Medical Center of Eulji University, Seoul, Korea.
authors.forename: Eun Joo
authors.initials: EJ
authors.lastname: Park
authors.forename: Jung Whan
authors.initials: JW
authors.lastname: Shin
authors.forename: Yong Soo
authors.initials: YS
authors.lastname: Seo
authors.forename: Dae Wun
authors.initials: DW
authors.lastname: Kim
authors.forename: Seo Yoo
authors.initials: SY
authors.lastname: Hong
authors.forename: Won Il
authors.initials: WI
authors.lastname: Park
authors.forename: Byung Moon
authors.initials: BM
authors.lastname: Kang
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism*; D053181:Caspase 8 / Q000378:metabolism*; D053453:Caspase 9 / Q000378:metabolism*; D002478:Cells, Cultured; D053938:DNA Fragmentation / Q000187:drug effects; D005260:Female; D005307:Fertilization in Vitro; D007987:Gonadotropin-Releasing Hormone / Q000819:agonists*; D006801:Humans; D020287:In Situ Nick-End Labeling; D008184:Luteal Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism
chemical_list: D007987:Gonadotropin-Releasing Hormone; D011065:Poly(ADP-ribose) Polymerases; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9