Cyclin B1 is known to reflect the M-phase promoting factor (MPF), a universal regulator of G2/M-phase transition, activity during the process of oocytes maturation. To investigate whether culture condition affects the maturation rate and the expression of cyclin B1 protein, bovine immature oocytes are stimulated and cultured according to the following protocols: Experiment 1: denuded oocytes (denude) only, COC only, denuded oocytes + granulosa cells (denude + GCs) and COC + GCs; Experiment 2: no-activation (control), 7% ethanol for 5 min and 10 l/ml ionomycin for 5 min at immediately before maturation. The maturation rates of denude and no-activation group were significantly lower in both experiments (P<0.05), respectively. Co-culture or stimulation method in bovine immature oocytes culture increases the cyclin B1 expression significantly in both experiments (P<0.05). Based on these results, culture condition affects the maturation rate and the expression of cyclin B1 protein during the first meiotic maturation in bovine immature oocytes.

Pregnancy-associated plasma protein-A (PAPP-A) is a 200 kDa metalloprotease identified as an IGFBP-4 protease and likely an important regulator of IGF bioavailability. PAPPA mRNA is detected in bovine granulosa and theca cells and the PAPP-A protein is also found in follicular fluid. Proteolytic activity supposed to be due to the PAPP-A in bovine follicular fluid is induced by follicle stimulating hormone (FSH) treatment and PAPP-A-like activity appears concomitantly with increased E2 during follicular development. However, the effects of FSH, E2 and progesterone on the expression of PAPP-A in bovine corpus luteum of pregnancy have been evaluated in a limited number of studies. This study was performed to expression of PAPP-A and progesterone during early pregnancy in bovine corpus luteum. To determine the function of PAPP-A gene during early pregnancy, we collected the bovine pregnancy corpus luteum samples on 30, 60 and 90 day of pregnancy. The mRNA expression of PAPP-A, progesterone-receptor, VEGF and IGFBP4 gene was conducted by real-time PCR. And proteins expression of PAPP-A and progesterone antibody was detected by Immunohistochemistry and ELISA. The VEGF and PAPP-A mRNA expression was progressively increased on day 90 in the pregnancy corpus luteum. The mRNA expression of progesterone-receptor, IGFBP4 in the corpus luteum progressively was increased from 30 to 60 day, but decreased on 90 day of pregnancy. The proteins expressions of progesterone and PAPP-A were similar pattern in mRNA expression. Our results indicate that the IGFBP4 protease role of PAPP-A increases in response to pregnancy 90 day corpus luteum and suggest that progesterone is an connected for the expression of PAPP-A genes in luteal cells. Therefore, we suggest that PAPP-A stimulated with progesterone play a crucial role for IGF-I system in bovine early pregnancy. And could be used to predict the condition of normal pregnancy.