BDNF Val66Met Polymorphism Is Associated With Female Infertility

BackgroudBrain-derived neurotrophic factor (BDNF) is an important member of neurotrophin superfamily, which has been proved to be the most abundant and widely expressed neurotrophin in the mammalian nervous system with the functions of neuronal survival, migration, phenotypic differentiation, axonal and dendritic growth, and synapse formation such as long-term potentiation, learning, and memory. Recently, a BDNF gene196G/A polymorphism was identified, which results in an amino acid substitution (Val/Met) in the position66at BDNF prodomain and leads to altered intracellular trafficking and decreased regulated secretion of BDNF in neurons.Besides, in many other organs, BDNF and its receptor TrkB also existed, like hypothalamus, pituitary, ovary and uterus. Many studies demonstrate that BDNF play important role in oocytes maturation and fertilized egg development. Moreover, plasma and follicular fluid BDNF levels exhibit dynamic changes during the menstrual cycle in normally cycling fertile women or in the process of controlled ovarian stimulation for IVF. These findings suggested that BDNF may also play a role in female reproduction.In this experiment, we will take endometriosis for instance, to explore the effect of BDNFMetgene polymorphism on infertility. At the same time, we use BDNFMettransgenic mice to investigate the mechanism of BDNF Val66Met gene polymorphism associated with infertility.ObjectiveThough BDNFMetgene polymorphism has been reported to interact with some neurologic disease, little is known about the role of BDNFMetgene polymorphism in reproductive system. Therefore, in the present study.two questions are raised. First, does BDNFMetgene polymorphism play a role in human and mice reproductive system? Second, what is the mechanism?Methods1. Association between BDNF Va166Met polymorphism and human infertility1.1Human genotyping1.2Infertile woman IVF-ET test1.3Detection BDNF protein levels in plasma and follicular fluid2. Association between BDNF Val66Met polymorphism and mice infertility2.1Mice mating test2.2Mice estrous cycle test at10-week-old and28-week-old2.3Paraffin embedding the organs of ovary and uterus2.4HE staining2.5ELISA, BDNF protein level test2.6SuperovulationResults1. The role of BDNF Val66Met polymorphism in endometriosis-related infertility1.1BDNF Val66Met polymorphism increase the incidence of endometriosis-related infertility.In control population, the frequencies of BDNF Val66Met genotypes and allele showed no statistically different between the fertile and infertile subgroups(P>0.05). However, compared with those in control infertile women, higher BDNFMet/Met genotype(P=0.043) and Met allele frequencies(P=0.015) were shown in endometriosis-related infertile patients.1.2BDNF Val66Met polymorphism decrease the IVF outcomes of infertile patients. We further analyzed the effect of BDNF Val66Met polymorphism on IVF outcomes of198endometriosis-related infertile patients, whose control were102tubal obstructed infertile patients. No significant differences were found in some indexes, such as age, BMI, basal FSH levels etc.(P>0.05). However, though administrated with higher doses of gonadotropins (P<0.01), patients with BDNFMet/Met genotype had achieved fewer mature oocytes (P<0.01), lower fertilization rate (P<0.01) and lower good quality rate of embryos (P<0.05) than those in BDNFVal/Val carriers. A similar result were also found in tubal obstructed infertile patients between BDNFMet/Met and BDNFVal/Val carriers.1.3The plasma/follicular fluid BDNF levels in endometriosis-related and tubal obstructed infertile patientsWe measured the plasma and follicular fluid BDNF levels on the day of oocyte retrieval in102control infertile women and198endometriosis-related infertile women with IVF treatment. No significant differences were found in plasma BDNF levels among different BDNF genotype in tubal obstructed and endometriosis-related infertile groups. However, follicular-fluid BDNF levels in different BDNF Val66Met genotypes showed significant difference in both groups.2. The effect of BDNF Val66Met polymorphism on mice reproduction2.1Val66Met polymorphism induces subfertility in female miceFertile males were bred with10-week-old female littermates for1week. BDNFMet/Met mice exhibited reduced fertility by showing a decrease in the rate of pregnancy(P<0.05), although the litter size is almost equal compared with BDNFvai/vai mice2.2The effects of Val66Met polymorphism on the estrous cycleThere was no significant difference in the length of each stage. Mean estrous cycle duration (ECD) of10-week-old female mice was roughly5days and did not differ between the two genotypes. However,28-week-old female BDNFMet/Met mice have a irregulate estrous cycle.2.3Morphological changes in adult mice ovaries, not in immature mice, and there is no changes in uterus There was no significant difference in the number of follicles when comparing the two genotypes at3weeks of age. At12weeks of age, the ovaries of BDNFMet/Met mice showed fewer antral follicles and corpora lutea, and more atretic follicles compared to BDNFVal/Val mice(P<0.05), showing that there was no difference in follicle reservation, but in the course of follicle development.In addition, uterine function was also examined. The uteri of BDNFMet/Met mice developed equally compared to BDNFVal/Val mice(P>0.05), no matter the mice age.2.4Comparison BDNF protein levels in HPO axis between different genotype miceCompared with control wild-type mice, BDNFMet/Met mice shows a lower level of BDNF protein in hypothalamus, pituitary, and ovary. However, in uterus, no significant difference has been found.2.5Ovarian function was impaired in BDNFMet/Met miceThe protocol consisted of a single i.p. injection with7.5units of pregnant mare serum gonadotropin for48h, followed by10units of human chorionic gonadotropin for an additional16h. BDNFMet/Met produced significantly fewer oocytes (P<0.05) induced by exogenous gonadotropins, compared to wild type mice, which is indicated that the reaction properties of BDNFMet/Met mice to exogenous gonadotropins is reduced.Conclusions1. BDNF Val66Met polymorphism is associated with endometriosis-related infertility in Chinese Han population. Moreover, this SNP can lead to poor IVF outcomes both in endometriosis-related and tubal obstructed infertile women, which may due to the decreased BDNF levels in follicular fluid.2. BDNFMet/Met mice shows a lower fertility, which might associate with follicle maldevelopment or the decreased BDNF levels in hypothalamus, pituitary, and ovary. Ovarian function was impaired in BDNFMet/Met mice.