Effects of midkine during in vitro maturation of bovine oocytes on subsequent developmental competence

Midkine (MK) is known to be a member of a new family of heparin-binding gro wth/differentiation factors, together with pleiotrophin, and to be quite ri ch in bovine follicular fluid. To examine whether treatment with MK during in vitro maturation (IVM) of bovine granulosa-enclosed oocytes affects thei r nuclear maturation and postfertilization development to the blastocyst st age, bovine granulosa-enclosed oocytes obtained from slaughterhouse-derived ovaries were cultured for 24 h in IVM medium without (control) or with var ious concentrations (1-500 ng/ml) of MK, followed by in vitro fertilization (IVF) and culturing. Although the MK treatment during IVM did not affect t he rate of nuclear maturation or the postfertilization cleavage of oocytes, MK at greater than or equal to 10 ng/ml significantly (P < 0.05) increased the blastocyst yields per tested and per cleaved oocyte compared with the case of the control. Next, the effects of various glycosaminoglycans (hepar in, heparan sulfate, chondroitin sulfate A and C, and hyaluronic acid) prei ncubated with MK at 50 ng/ml were examined. The enhancing activity of MK wa s completely suppressed by heparin at 600 ng/ml but not by the other compou nds. The effects of MK during IVM were also tested on oocytes freed from gr anulosa cells (GCs), When the denuded oocytes were cultured in IVM medium, no blastocyst formation after IVF was observed, regardless of MK supplement ation. However, coculture of the denuded oocytes with isolated GC pellets e nhanced the cleavage rates and the blastocyst yield, and these effects were more pronounced with MK supplementation. These results indicate that the p resence of MK during IVM of bovine granulosa-enclosed oocytes can enhance t heir developmental competence to the blastocyst stage after IVF and suggest that the enhancing effects might be mainly mediated by GCs.