The Effect Of Culture Immature Oocytes In Vitro By Adding Follicular Fluid Or Hmg To Medium

The majority of in vitro fertilization-embryo(IVF) transfer programs conventionally adopt controlled ovarian hyperstimulation technology(COH). This usually can obtain10to14mature oocytes. However about15-20%of oocytes can not reach mature stage, part of which are often discarded. Studies had shown that denuded oocytes and eggs with granulosa cells could be cultured to reach mature stage, and the results were similar to fertilization rate, cleavage rate and available embryo rate. These confirms that the immature denuded oocytes can be matured in vitro. Taking full advantage of this part of the immature oocytes after ICSI to culture them to mature can increase the number of embryos in ICSI cycles and the pregnancy rate in these patients can also been improved. Although the IVM technology expanded rapidly in the past10years, IVM technology is still at the exploratory stage. It is still low in the maturation rate, fertilization rate, cleavage rate, planting rate and other issues. The key to solve these problems lies in the IVM culture medium composition. Now international companies have developed some systems of human immature oocytes in vitro maturation culture medium, but the medium still could not escape the defects of the low success rate. Our study aims to research these issues.ObjectivesWe aims to explore the effect on immature oocyte after COH in IVF process by adding the different components to commercial medium of immature oocytes in vitro maturation and then establish an effective human oocyte in vitro mature system.MethodsIn this study, we collected265immature oocytes in216ICS1cycles. These patients accepted ICSI in the Third Affiliated Hospital of Zhengzhou University from March to November in2011.And ICSI indications were all male factor.Preparing IVM medium:Group A(basis medium of IVM):commercial medium of IVM; Group B (hormone group):add0.075IU/ml of FSH and0.075IU/ml of LH to the basis medium of IVM; Group C(mature follicular fluid):mixture of50%basis medium of IVM and50%mature follicular fluid.In ICSI process,the oocytes were stripped of granule cells and observed. The immature oocytes were placed to the prior preparation of IVM fluid and observed in each group at24h and48h. If they matured,they had been oprated ICSI and then observed fertilization, the embryos were scored. Lastly the final statistical were analysed.This study had been approved by the committee of ethics of the hospital and obtained the patients agreement.Spss16.0statistical package was applied. Counting data used the χ2test, Fisher’s exact test. The test level was a=0.05.Results1General situationIn216ICSI patients, average age were35.9±5.4years old. Obtained immature oocytes were265:155in MI stage and110GV stage.By IVM,126oocytes matured at24h with a mature rate of47.5%and165oocytes matured at48h wih a mature rate of62.3%.2The effect on immature oocytes in vitro maturation by three different medium:At24h,there was no significant difference in maturation rate in the three groups (P>0.05). At48h, maturation rate in group C (74.7%) was significantly higher than in group A (53.4%) and group B (57.3%), the difference was statistically significant (P<0.05). There was no significant difference in the maturation rate between group A and Group B (P>0.05). At MI stage oocytes,48h maturation rate (81.7%) in group C was significantly higher than group B (62.2%)(P<0.05).At GV stage oocytes,48h maturation rate (62.9%) in group C was significantly higher than group A (36.8%)(P<0.05).3The result of the different period immature oocytes in vitro maturation:At MI stage oocytes,24h、48h maturation rate (59.3%,77.4%) were significantly higher than GV stage oocytes (30.9%,50.0%)(P<0.05)4The result of fertilization rate, cleavage rate and available embryo rate after IVM in three groupsIn group C cleavage rate, available embryo rate (91.3%,66.7%) was significantly higher than group B (71.0%,40.9%) and group A(67.6%,36%)(P <0.05). There were no significant difference between group B and group A(P>0.05). Fertilization rate had no significant difference in the three groups.Conclusions1Adding follicular fluid line in vitro maturation could improve the maturation rate and developmental potential of immature oocytes.2The effect of culture adding hormones was better than the commercialization IVM medium culture.3The maturation rate of MI stage oocytes was higher than that of GV stage oocytes in vitro culture.