Effects Of Puberty And Gonadotropins On Mouse Oocyte Developmental Capacity In Vitro

In vitro matured (IVM) oocytes of Prepubertal mammals can obtain a large mount of oocyte for IVF and animal in vitro embryo production and cloning.But the quality of the IVM Prepubertal mammals oocytes is generally lower than that of Adult, the embryo developmental ability of oocytes matured and fertilized in Prepubertal oocytes is very low. Several studies have found some difference in Prepubertal and Adult oocyte,such as follicular diameter,oocyte diameter,mitochondrial distribution,microtubule configurations and mRNA and protein synthesis.These factors result in incomplete cytoplasmic maturation,fertilized rates and blastocysts rates decreased. Therefore, it is very important for us to find the difference between prepubertal and adult oocyte and improve oocyte developmental capacity by improveing IVM system of oocyte.In this study, we use mouse oocyte as experimental model to find the difference between prepubertal and adult oocyte further and improve oocyte developmental capacity (1)follicular and oocyte diameter is a important parameters in oocyte development,large follicular and oocyte will get a better developmental capacity, synchrony between follicle and oocyte growth is reflect the quantities of follicular cells.if these question is exist in prepubertal and adult oocytes.(2)Polyspermic is an universal problem in prepubertal oocyte fertilized in vitro, Polyspermic result in early cleavage and developmental failure.So,we study the factor in Polyspermic to decrease Polyspermic and increase embryo developmental capacity.(3)Gonadotropins can increase the mount of big antral follicular,but the role in oocyte developmental capacity is controversial.we use mouse oocyte model to study if Gonadotropins can increase the developmental capacity in prepubertal oocyte.(4) Intracellular GSH levels are very important for the IVF process and the first stages of embryo development,Precious study concluded that cysteamine and cystine when added to IVM culture mediums of oocytes can improve GSH synthesis and embryo development.If it can improve prepubertal oocyte developmental capacity.(5) GSH is synthesized via the action of they-glutamylcysteine synthetase (yGCS) and GSH synthetase, yGCS is the rate-limiting enzyme in GSH synthesis and is composed of a catalytic heavy subunit and a regulatory light subunit, which are encoded by different genes,we will study if these gene has difference in prepubertal and adult oocytes.(6)If gonadotropins has effect on GSH synthesis and GSH synthetase mRNA expressionThe results were summarized as follows:(1)SrCl2 optimum activation concentration of IVM oocytes is 20mM,It can get better activated rates and blastocysts rates(2)After IVF and parthentic,the blastocysts rates is lower in prepubertal oocyte than adult oocyte.DO oocytes is also low,there is no difference between 4 weeks and 6-8 weeks oocyte.PMSG can improve developmental capacity in prepubertal and adult oocyte,there is no difference of the blastocysts rates in prepubertal and adult oocyte(3)there is no difference of the Penetrated spermatozoa rates and pronuclear diameter in prepubertal and adult oocyte,polyspermic rates and polyspermy but no pronuclear rates in prepubertal oocyte is higher than adult.PMSG increased nomal Fertilization rates,male pronuclear former rates and pronuclear diameters.(4) there is no difference of cortical granule distribution complete rates in 3 and 4 weeks mouse oocytes,4 and 6-8weeks mouse oocyte has also no difference,6-8weeks mouse oocyte cortical granule distribution complete rates higher than 3 weeks. PMSG increased cortical granule distribution complete rates in 3 and 6-8weeks mouse oocyte.(5)GSH in oocyte of prepubertal is lower than adult oocyte,PMSG increased GSH in prepubertal and adult mouse oocyte.(6)oocytes were cultured in TCM-199 supplemented with 100μuM cysteamine and 200μM cystine can improve developmental capacity in prepubertal and adult mouse oocyte.200μM cysteamine and 400μM cystine can not improve further.PMSG treated groups also improved when added 200μM cysteamine and 400μM cystine.(7) catalytic subunit and regulatory subunit of GSH synthesis mRNA expression is higher in adult mouse oocyte,there is no difference of GSS in prepubertal and adult mouse oocyte.PMSG can increase the three gene mRNA expression.(8)follicular oocytes diameter and growth synchrony is not the factor of prepubertal oocyte development capacity poor.