In Vitro Maturation (IVM) And Its Molecular Biology Preliminary Study On Immature Oocytes

The present study was conducted to apply the in vitro maturation (IVM) systerm for clinic treatments on infertile women. Human immature oocytes were matured in IVM medium supplied with human mature follicular fluid. After matured in vitro, the oocytes were fertilized in vitro by IVF or intracytoplasmic sperm injection (ICSI), the fertilization efficiency was compared between two ways. The differences of the fertilization competence of human oocytes matured in vitro or in vivo by ICSI using ejaculated and testicular spermatozoa were also compared. Finally, we studied the mechanism of molecular biology on maturation of oocytes preliminarily using proteomic analysis of mouse ovaries. Some specific proteins were selected from proteome to analysis their function by immunohistochemistry. We hope that we can evaluate the clinic safety of this IVM systerm basing on the preliminary molecular biology study on oocytes maturation.Part 1. Study on the fertilization competence of human oocytes matured in vitroDuring the past a few years, the technique of IVM of human oocytes has become an attractive option for the treatment of female infertility. The pregnancies and newborn infants have been obtained after maturation in vitro of oocytes recovered following ovariectomies, from polycystic ovarian syndrome (PCOS) patients, from women undergoing ICSI in natural cycles, and from poor responders or patients with retarded follicular development during a conventional stimulation IVF cycle. In IVM treatment, ICSI serves as a regular insemination method to tackle zona changes of in vitro matured oocytes resulted from long in vitro culture. It has been reported that the main factor affected the fertilizationresults of IVM oocytes is IVM medium instead of the manner of fertilization. So we believe that IVF may be a suitable way to be used for fertilization with IVM oocytes following improvement on IVM medium. It should be stressed that the difference between IVM oocytes and in vivo matured oocytes was mature environment, and the difference would affect the results of fertilization and embryonic development. Additionally, it has been reported that a twin pregnancy had been achieved from IVM-matured oocytes from unstimulated cycles fertilized by ICSI using spermatozoa from epididymis by percutaneous sperm aspiration (PESA). However, there is no report on whether pregnancy and live birth can be established from IVM oocytes fertilized with testicular aspirated spermatozoa. Moreover, little is known for differences of the fertilization competence of human oocytes matured in vitro and in vivo when fertilized with spermatozoa from ejaculates and testicle following ICSI.Therefore the objectives of the present study were 1) to compare the fertilization ability of human IVM oocytes by different ways including IVF and ICSI following improvement on IVM medium, 2) to investigate the differences of the fertilization and embryonics developmental competence of human oocytes matured in vitro and in vivo when fertilized with spermatozoa from ejaculates and testicle following ICSI.The main results were as follows: 1. Human immature oocytes were matured in IVM medium with 10% human mature follicular fluid (HMFF). After matured in vitro, oocytes were fertilized in vitro by IVF (IVF group) or ICSI (ICSI group). There was no significant difference on the rates of fertilization, the cleavage rates and the percentages of embryos suited for embryo transfer (ET) between groups (P>0.05), although the rate of abnormal fertilization and the percentages of embryos suited for ET in IVF group were higher than those in ICSI group, and no good quality embryos developed in IVF group on ET day. 2. Thepercentages of oocytes achieving normal fertilization in the groups that human oocytes matured in vitro fertilized with spermatozoa in ejaculates (Group 1: 72.9%) and oocytes matured in vivo fertilized with spermatozoa in ejaculates (Group 3: 67.0%) was significantly higher than the groups that human oocytes matured in vitro fertilized with spermatozoa from testicle (Group 2: 54.5...