Gonadotropin-releasing hormone antagonist cetrorelix down-regulates proliferating cell nuclear antigen and epidermal growth factor expression and up-regulates apoptosis in association with enhanced poly(adenosine 5'-diphosphate-ribose) polymerase expression in cultured human leiomyoma cells.
The objective of this study was to elucidate the effects of GnRH antagonist Cetrorelix on proliferation and apoptosis in human leiomyoma cells cultured in vitro. Isolated leiomyoma cells were subcultured in phenol red-free DMEM supplemented with 10% fetal bovine serum for 120 h and then stepped down to serum-free conditions in the presence or absence of graded concentrations of Cetrorelix (10(-5) to 10(-8) mol/liter) for 6 d. Cultured leiomyoma cells were used for semiquantitative RT-PCR, immunocytochemistry, Western blot analysis, and terminal deoxynucleotidyl transferase-mediated deoxyuridine 5-triphosphate nick-end labeling assay. RT-PCR analysis revealed the presence of mRNAs encoding for GnRH receptor and epidermal growth factor (EGF) in cultured leiomyoma cells. The number of viable cultured leiomyoma cells was significantly (P < 0.01) decreased by treatment with Cetrorelix compared with untreated control cultures. Immunocytochemical examination demonstrated that treatment with Cetrorelix attenuated the expression of proliferating cell nuclear antigen (PCNA) and EGF in cultured leiomyoma cells. Western blot analysis revealed that treatment with 10(-5) mol/liter Cetrorelix significantly (P < 0.01) decreased PCNA expression. In addition, treatment with 10(-5) mol/liter Cetrorelix remarkably increased the terminal deoxynucleotidyl transferase-mediated deoxyuridine 5-triphosphate nick-end labeling-positive rate and poly(ADP-ribose) polymerase expression at 24 h of treatment compared with untreated control cultures (P < 0.01). Furthermore, treatment with 10(-5) mol/liter Cetrorelix decreased immunoreactive EGF protein and EGF mRNA expression in cultured leiomyoma cells at 4 d of treatment. GnRH antagonist Cetrorelix may directly inhibit leiomyoma cell growth by down-regulating proliferation in association with a decrease in EGF mRNA expression and by up-regulating apoptosis in those cells.
Chen
Wei
W
Department of Obstetrics and Gynecology, Kobe University Graduate School of Medicine, Chuo-Ku, Kobe 650-0017, Japan.
Yoshida
Shigeki
S
Ohara
Noriyuki
N
Matsuo
Hiroya
H
Morizane
Mayumi
M
Maruo
Takeshi
T
D000328:Adult; D017209:Apoptosis / Q000187:drug effects*; D001483:Base Sequence; D002470:Cell Survival / Q000187:drug effects; D017931:DNA Primers; D004815:Epidermal Growth Factor / Q000235:genetics*; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D007987:Gonadotropin-Releasing Hormone / Q000031:analogs & derivatives* / Q000037:antagonists & inhibitors* / Q000494:pharmacology*; D006801:Humans; D007889:Leiomyoma / Q000235:genetics* / Q000473:pathology / Q000601:surgery; D011065:Poly(ADP-ribose) Polymerases / Q000235:genetics*; D017697:Premenopause; D018809:Proliferating Cell Nuclear Antigen / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured; D014594:Uterine Neoplasms / Q000235:genetics* / Q000473:pathology / Q000601:surgery
D017931:DNA Primers; D018809:Proliferating Cell Nuclear Antigen; D012333:RNA, Messenger; D007987:Gonadotropin-Releasing Hormone; D004815:Epidermal Growth Factor; D011065:Poly(ADP-ribose) Polymerases; C062876:cetrorelix