The kinase inhibitor indirubin-3'-oxime prevents germinal vesicle breakdown and reduces parthenogenetic development of pig oocytes.
Oocytes undergo spontaneous germinal vesicle breakdown (GVBD) after being released from the follicular environment; this potentially prevents manipulation of the oocyte at the germinal vesicle (GV) stage. The objectives of this study were to investigate the effects of indirubin, a potent cdc2 kinase inhibitor, on GVBD and microtubular structure of porcine oocytes. Cumulus-oocyte-complexes (COCs) were collected from abattoir-derived ovaries and were randomly allocated to different concentrations of indirubin treatments (0, 10, 25, 50, and 100 microM in Experiment 1 and 0, 50, 75, and 100 microM in Experiment 2) during 44 h of IVM. The influences on the GVBD, microtubules, and maturation rates were evaluated using epifluorescence microscopy. The percentages of oocytes remaining at the GV stage were 0, 16, 26, 69, and 85% for oocytes treated with 0, 10, 25, 50, and 100 microM of indirubin, respectively, which differed among treatment groups (P<0.05). However, there were no significant differences between the oocytes treated with 75 and 100 microM (79 and 81%). The cytoplasmic microtubules were fragmented in oocytes maintained at the GV stage and the chromatin became condensed or aggregated. When COCs were incubated with indirubin (50-75 microM) for 22 h and then transferred to maturation medium for 44 h (Experiments 3-5), the percentages of oocytes reaching the metaphase II stage were generally higher than when the COCs were cultured in the presence of the drug for 44 h (62-65% versus 44-46%). However, the parthenogenetic development of the oocytes in Experiment 6 was reduced significantly in drug-treated oocytes. In summary, treatment with 50-75 microM of indirubin effectively prevented GVBD in porcine oocytes, but the developmental competence of the oocytes was compromised.
Hsieh
Chang-Hsing
CH
Taichung Military General Hospital, National Chung Hsing University, Taichung, Taiwan, ROC.
Tang
Pin-Chi
PC
Chang
Wen-Hung
WH
Weng
Ya-Chi
YC
Sha
Shih-Wen
SW
Tseng
Jung-Kai
JK
Chang
Lan-Hwa
LH
Ju
Jyh-Cherng
JC
D000818:Animals; D016203:CDC2 Protein Kinase / Q000037:antagonists & inhibitors; D002478:Cells, Cultured; D002843:Chromatin / Q000187:drug effects; D003599:Cytoskeleton / Q000187:drug effects; D005260:Female; D007150:Immunohistochemistry; D007211:Indoles / Q000494:pharmacology*; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000648:ultrastructure; D009865:Oocytes / Q000187:drug effects* / Q000502:physiology / Q000648:ultrastructure; D010091:Oximes / Q000494:pharmacology*; D010312:Parthenogenesis / Q000187:drug effects*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D013552:Swine
D002843:Chromatin; D007211:Indoles; D010091:Oximes; D047428:Protein Kinase Inhibitors; C474021:indirubin-3'-monoxime; D016203:CDC2 Protein Kinase