|Budget Amount *help
¥1,900,000 (Direct Cost : ¥1,900,000)
Fiscal Year 1994 : ¥800,000 (Direct Cost : ¥800,000)
Fiscal Year 1993 : ¥1,100,000 (Direct Cost : ¥1,100,000)
Granulosa cells of follicles in avian species, such as chicken and Japanese quail, . proliferate during follicular growth, and differentiate to secrete progesterone in response to luteinizing hormone (LH), when the follicle becomes the largest and reaches to the time of ovulation. In order to study the mechanism and signal transduction pathway of growth and differentiation, quail granulosa cells were cultured with various gonadotrophins or growth factors, such as epidermal growth factor (EGF).
The culture medium was a mixture of McCoy's 5a and Ham's F12 (1 : 1) supplemented with glutamine, HEPES,Penicilline and Streptomycin. The concentration of fetal calf serum was 10% during the initial 6 hr of the culture, and then it was reduced to 1%. In order to evaluate the LH response of the cells after the culture, the cells were incubated with or without LH for 3 hr in Krebs-Ringer solution supplemented with bovine serum albumin, and progesterone production was measured in the medium during this 3 hr of incubation by the specific radio immunoassay.
When the granulosa cells were cultured with follicle stimulating hormone (FSH) for 72 hr, the response to LH was increased while the cell numbers were not significantly increased. When the granulosa cells were cultured with EGF for 72 hr, the cell numbers increased dramatically, but the LH response of the cells, which was evaluated by the production of progesterone, was diminished. These results suggested that the growth and differentiation of the granulosa cells during follicular maturation was controlled by both FSH and EGF.