title: A specific inhibitor of CDK1, RO-3306, reversibly arrests meiosis during in vitro maturation of porcine oocytes.
abstract: CDK1 plays pivotal role in meiotic progression of oocytes from G2 to metaphase II (MII) stage. In this study, we investigated the possibility of utilizing a selective inhibitor of CDK1, RO-3306, as a novel agent for the synchronization of oocyte maturation. Two groups of cumulus-oocyte complexes (COCs) were treated with 10 μM RO-3306. The first group was treated for 44 h, whereas the second group was transferred to drug-free medium after a 20 h treatment. MII-stage oocytes from each group were confirmed by cytoplasmic maturation and embryonic development assays. Treatment of immature porcine oocytes with RO-3306 for 20 h arrested them at the germinal vesicle (GV) stage. The GV-arrest effect of RO-3306 was reversible: when RO-3306-arrested COCs were subsequently cultured for 24h in the absence of RO-3306, 76.19 ± 2.68% of these oocytes reached the MII stage after 44 h of in vitro maturation, a rate similar to that of non-treated control oocytes (79.08 ± 3.23%). Furthermore, RO-3306-treated oocytes transferred to drug-free media did not differ significantly from controls (P>0.05) with respect to cleavage and blastocyst formation upon parthenogenetic activation. To explore the underlying molecular mechanisms, we examined the expression patterns of four representative maternal transcripts, CDK1, Cyclin B1, GDF9, and BMP15, by real-time polymerase chain reaction (PCR) and poly(A)-test PCR (PAT assay). RO-3306 treatment increased expression of CDK1 but had no effect on the expression of the other genes. These data suggest that RO-3306 efficiently blocks and synchronizes the meiotic progression of porcine oocytes at the GV stage without affecting their meiotic and cytoplasmic maturation.
authors.affiliation: Department of Animal Sciences, Chungbuk National University, Cheongju, Chungbuk 361-763, South Korea.
authors.forename: Woo-In
authors.initials: WI
authors.lastname: Jang
authors.affiliation: Department of Animal Sciences, Chungbuk National University, Cheongju, Chungbuk 361-763, South Korea.
authors.forename: Zi-Li
authors.initials: ZL
authors.lastname: Lin
authors.affiliation: Department of Animal Sciences, Chungbuk National University, Cheongju, Chungbuk 361-763, South Korea.
authors.forename: Sung Hyun
authors.initials: SH
authors.lastname: Lee
authors.affiliation: Department of Animal Sciences, Chungbuk National University, Cheongju, Chungbuk 361-763, South Korea.
authors.forename: Suk
authors.initials: S
authors.lastname: Namgoong
authors.affiliation: Department of Animal Sciences, Chungbuk National University, Cheongju, Chungbuk 361-763, South Korea. Electronic address: nhkim@chungbuk.ac.kr.
authors.forename: Nam-Hyung
authors.initials: NH
authors.lastname: Kim
mesh_terms: D000818:Animals; D016203:CDC2 Protein Kinase / Q000037:antagonists & inhibitors; D002478:Cells, Cultured; D005260:Female; D059471:In Vitro Oocyte Maturation Techniques / Q000662:veterinary; D008540:Meiosis / Q000187:drug effects*; D009865:Oocytes / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D009866:Oogenesis / Q000187:drug effects; D026723:Polyadenylation / Q000187:drug effects; D011804:Quinolines / Q000494:pharmacology*; D013552:Swine; D013844:Thiazoles / Q000494:pharmacology*
chemical_list: D011804:Quinolines; C512984:RO 3306; D013844:Thiazoles; D016203:CDC2 Protein Kinase
keywords: CDK1; In vitro maturation; Maturation/M-phase promoting factor; Porcine oocyte; RO-3306