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Glutamine 함유 배양액에서 glucose와 pyruvate의 조성이 착상전 생쥐 배아의 체외발달에 미치는 영향
김주환 ( Kim Ju Hwan ),서영석 ( Seo Yeong Seog ),최송기 ( Choe Song Gi ),이오성 ( Lee O Seong ),송해범 ( Song Hae Beom ),강길전 ( Kang Gil Jeon ),이기환 ( Lee Gi Hwan ) 대한산부인과학회 2004 Obstetrics & Gynecology Science Vol.47 No.1
Objective : The purpose of this study was to examine the effects on in vitro development of early preimplantation mouse embryos in DMEM medium with glutamine which was controlled by different composition of glucose and pyruvate. Methods : Four hundred and
생쥐 pre-antral follicle의 체외 배양체계의 확립: 호르몬 무첨가 배양액에서 협막-기질세포에 의한 난포의 체외성장 효과
이기환 ( Lee Gi Hwan ),김주환 ( Kim Ju Hwan ),서영석 ( Seo Yeong Seog ),송해범 ( Song Hae Beom ),강길전 ( Kang Gil Jeon ) 대한산부인과학회 2004 Obstetrics & Gynecology Science Vol.47 No.3
Objective : This present study was conducted to examine the effects on in vitro growth of pre-antral mouse follicles by thecal-stromal cells attached to around the follicular membrane in culture medium without hormones. Methods : Pre-antral follicles (100-130 ㎛)used in our studies were isolated mechanically by fine 30 G needles attached to 1 ㎖ insulin syringe from mice ovaries of 20-25 days old female ICR strain. Isolated pre-antral follicles were divided into three groups by attached status of thecal-stromal cell layers to follicular membrane. Follicular Initial diameters of group Ⅰ was 112.5 (mean)±6.2 (SD) ㎛ (n=31), group Ⅱ was 112.8±7.8 ㎛ (n=23), and group Ⅲ was 115.1±6.5 ㎛ (n=27). Divided pre-antral follicles were washed in fresh Ham`s F-10 medium and cultured in 20 ㎕ droplets of DMEM with glutamine, glucose and pyruvate under mineral oil on the 60 ㎜ culture dish. All experimental media were supplemented with 10% FBS without hormones. Media were exchanged wholly by fresh media every 2 days in culture. Diameters of intrafollicular oocytes and cultured pre-antral follicles were measured using an precalibrated cross ocular micrometer at ×200 every day during 6 days in vitro culture. Results were considered statistically significant when p value was less than 0.05 using Student`s t-test. Results : The mean and standard deviation (SD) of initial diameters of pre-antral follicles was 113.5±2.2 ㎛ in three groups. Follicular growth rate of group Ⅲ was significantly (p<0.05) higher in whole culture periods than group Ⅰ and group Ⅱ. Rate of follicular growth between group Ⅰ and group Ⅱ were not significant difference in culture for 1 to 3 days. However, group Ⅱ was significantly higher than group Ⅰ in culture for 4 days. Diameters of grown follicles for 6 days was 155.2±18.7 ㎛, 196.9±24.1 ㎛ and 284.2±47.6 ㎛ in group Ⅰ, group Ⅱ and group Ⅲ, respectively. Growth rate of intrafollicular oocytes between three groups were not difference and revealed to continuous growth patterns in whole culture periods. Follicular diameter after culture for 7 days were not measured because of disruption of follicular structure or outgrowth of granulosa cells. Conclusion : Pre-antral follicles were grown very well in DMEM medium without hormones. Thecal-stromal cells attached to the surface of follicular membrane has acted effectively in vitro growth of follicles.