개 난자의 체외성숙율을 높이기 위하여 다양한 방법들이 시도되고 있지만 여전히 그 효율성은 낮다. 본 연구는 개 난자의 체외성숙 시, 성선 자극 호르몬인 황체형성호르몬(LH)과 난포자극호르몬(FSH), 상피세포성장인자(Epidermal growth factor, EGF) 그리고 시스테인(cysteine)을 각각 첨가하여 72시간 동안 체외성숙시킨 후 핵성숙율(GV: germinal vesicle, GVBD: germinal vesicle break down, MI: metaphase I, MII: metaphase II, UK: unknown stage)을 확인하였다. LH와 FSH를 첨가하였을 때 첨가하지 않은 군과 GV, MI 및 MII율에는 유의적인 차이는 없었다. 하지만 GVBD율은 첨가군이 유의적으로(p<0.05) 높았다. 성선 자극 호르몬을 첨가한 배지에 10ng/ml의 EGF를 첨가하였을 때 MII율이 첨가하지 않은 군보다 유의적으로(p<0.05) 높았다(4.54% vs. 7.06%). cysteine을 첨가하였을 경우, 핵성숙율에 유의적인 차이는 보이지 않았지만 전반적으로 핵성숙율이 향상된 것을 확인할 수 있었다. 따라서 개 난자의 체외성숙 시, $10 {mu}g/ml$의 LH와 FSH, 10ng/ml의 EGF 그리고 0.57mM의 cysteine을 첨가하는 것이 핵성숙율을 향상시키는 것으로 사료된다.
Despite many efforts to improving canine in vitro maturation(IVM), the efficiency is still low compared to that of other mammalian species. The present study investigated the effects of gonadotropin, epidermal growth factor and cysteine supplementation on in vitro maturation of canine oocytes. Cumulus-oocyte complexes (COCs) were matured in basic medium (TCM-199 containing 10% FBS, 0.11mg/ml sodium pyruvate supplemented with or without $10{mu}g/ml$ FSH, 10ng/ml EGF and 0.57mM cysteine) for 72 hr at $38.5^{circ}C$ in humidified atmosphere of 5% $CO_2$ in air. After culture, oocytes were stained with Hoechst 33342 $(10{mu}g/ml)$ for 30 min at $4^{circ}C$, and assessed their nuclear status (GV: germinal vesicle, GVBD: germinal vesicle break down, MI: metaphasse I, MII: metaphase II, UK: unknown stage). No differences were observed in GV, MI and MII rate except GVBD rate between with and without gonadotropins addition respectively (6.7%, vs. 17.2%). Supplementation of 10ng/ml of EGF in hormones added IVM medium resulted in significantly (p<0.05) higher MII rate (4.54% vs.7.06%). Although there are no significantly difference, total of MI and MII rates were increased by adding cysteine. In conclusion, the present study indicates that supplementation of $10{mu}g/ml$ LH and FSH, 10ng/ml EGF and 0.57mM cysteine in canine IVM medium show a positive influence on the progression of maturation to MII at 72 hr.
