Cloning of equine prostaglandin dehydrogenase and its gonadotropin-dependent regulation in theca and mural granulosa cells of equine preovulatory follicles during the ovulatory process.
The mammalian ovulatory process is accompanied by a gonadotropin-dependent increase in follicular levels of prostaglandin E2 (PGE2) and PGF2alpha, which are metabolized by 15-hydroxy prostaglandin dehydrogenase (PGDH). Little is known about ovarian PGDH regulation in non-primate species. The objectives of this study were to characterize the structure of equine PGDH and its regulation in follicles during human chorionic gonadotropin (hCG)-induced ovulation. The full-length equine PGDH was obtained by RT-PCR, 5'- and 3'-rapid amplification of cDNA ends (RACE). Its open reading frame encodes a 266-amino acid protein that is 72-95% homologous to other species. Semi-quantitative RT-PCR/Southern blot were used to study PGDH regulation in follicles isolated 0-39 h post-hCG. Results showed that PGDH mRNA expression was low in follicles obtained at 0 h, increased at 12 and 24 h (P < 0.05), and decreased at 36-h post-hCG. This induction of expression was biphasic, with elevated abundance of transcripts at 12 and 33 h post-hCG (P < 0.05) in mural granulosa and theca cells. Immunohistochemistry and immunoblotting confirmed regulated expression of PGHD protein in both cell types of preovulatory follicles after hCG. High levels of PGDH mRNA were observed in corpus luteum and other non-ovarian tissues tested, except kidney, muscle, brain, and heart. Thus, this study is the first to report the gonadotropin-dependent regulation of PGDH during ovulation in a non-primate species. PGDH induction was biphasic in theca and mural granulosa cells differing from primates in which this induction was monophasic and limited to granulosa cells, suggesting species-specific differences in follicular control of PGDH expression during ovulation.
Sayasith
Khampoune
K
Centre de recherche en reproduction animale and Département de biomédecine vétérinaire, Faculté de médecine vétérinaire, Université de Montréal, 3200 Sicotte, Saint-Hyacinthe, Québec, Canada J2S 7C6. k.sayasith@umontreal.ca
Bouchard
Nadine
N
Doré
Monique
M
Sirois
Jean
J
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern / Q000379:methods; D015153:Blotting, Western / Q000379:methods; D001923:Brain Chemistry; D002478:Cells, Cultured; D006063:Chorionic Gonadotropin / Q000494:pharmacology; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000032:analysis; D004789:Enzyme Activation; D005260:Female; D006107:Granulosa Cells / Q000737:chemistry / Q000378:metabolism*; D006736:Horses / Q000502:physiology*; D006911:Hydroxyprostaglandin Dehydrogenases / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D007150:Immunohistochemistry; D007668:Kidney / Q000737:chemistry; D008969:Molecular Sequence Data; D009132:Muscles / Q000737:chemistry; D009206:Myocardium / Q000737:chemistry; D010060:Ovulation / Q000187:drug effects / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D013799:Theca Cells / Q000378:metabolism*
D006063:Chorionic Gonadotropin; D018076:DNA, Complementary; D006911:Hydroxyprostaglandin Dehydrogenases