Knockdown of regulatory associated protein of TOR (raptor) in hypothalamus-stimulated folliculogenesis and induced ovarian cysts.

CONTEXT: Mammalian target of rapamycin complex 1 (mTORC1) is an essential sensor that regulates fundamental biological processes like cell growth, proliferation and energy metabolism. The treatment of disease by sirolimus, a mTORC1 inhibitor, causes adverse effects, such as female fertility disorders. AIMS: The objective of the study was to decipher the reproductive consequences of a downregulation of mTORC1 in the hypothalamus. METHODS: The reduced expression of mTORC1 was induced after intracerebroventricular injection of lentivirus expressing a short hairpin RNA (shRNA) against regulatory associated protein of TOR (raptor) in adult female mice (ShRaptor mice). KEY RESULTS: The ShRaptor mice were fertile and exhibited a 15% increase in the litter size compared with control mice. The histological analysis showed an increase in antral, preovulatory follicles and ovarian cysts. In the hypothalamus, the GnRH mRNA and FSH levels in ShRaptor mice were significantly elevated. CONCLUSIONS: These results support the hypothesis that mTORC1 in the central nervous system participates in the regulation of female fertility and ovarian function by influencing the GnRH neuronal activity. IMPLICATIONS: These results suggest that a lower mTORC1 activity directly the central nervous system leads to a deregulation in the oestrous cycle and an induction of ovarian cyst development.

Perinatal Exposure to Methoxychlor Affects Reproductive Function and Sexual Behavior in Mice.

Numerous chemicals derived from human activity are now disseminated in the environment where their exert estrogenic endocrine disrupting effects, and therefore represent major health concerns. The present study explored whether Methoxychlor (MXC), an insecticide with xenoestrogens activities, given during the perinatal period (from gestational day 11 to postnatal day 8) and at an environmentally dose [20 µg/kg (body weight)/day], would affect reproductive physiology and sexual behavior of the offspring in mice. While MXC exposure did not induce any differences in the weight gain of animals from birth to 4 months of age, a clear difference (although in opposite direction according to the sexes) was observed on the anogenital distance between intact and exposed animals. A similar effect was also observed on preputial separation and vaginal opening, which reflects, respectively, in males and females, puberty occurrence. The advanced puberty observed in females was associated with an enhanced expression of kisspeptin cells in the anteroventral periventricular region of the medial preoptic area. Exposure to MXC did not induce in adult females changes in the estrous cycle or in the weight of the female reproductive tract. By contrast, males showed reduced weight of the epididymis and seminiferous vesicles associated with reduced testosterone levels and seminiferous tubule diameter. We also showed that both males and females showed deficits in mate preference tests. As a whole, our results show that MXC impacts reproductive outcomes.

Specific deletion of AMP-activated protein kinase (α1AMPK) in mouse Sertoli cells modifies germ cell quality.

The AMP-activated protein kinase (AMPK) is an important regulator of cellular energy homeostasis which plays a role in fertility. Complete disruption of the AMPK catalytic subunit α1 gene (α1AMPK KO) in male mice results in a decrease in litter size which is associated with the production of altered sperm morphology and motility. Because of the importance of Sertoli cells in the formation of germ cells, we have chosen to selectively disrupt α1AMPK only in the Sertoli cells in mice (Sc-α1AMPK-KO mice). Specific deletion of the α1AMPK gene in Sertoli cells resulted in a 25% reduction in male fertility associated with abnormal spermatozoa with a thin head. No clear alterations in testis morphology or modification in the number of Sertoli cells in vivo were observed, but a dysregulation in energy metabolism in Sertoli cells occurred. We have reported an increase in lactate production, in lipid droplets, and a reduction in ATP production in Sc-α1AMPK-KO Sertoli cells. These perturbations were associated with lower expression of mitochondrial markers (cytochrome c and PGC1-α). In addition another metabolic sensor, the deacetylase SIRT1, had a reduction in expression which is correlated with a decline in deacetylase activity. Finally, expression and localization of junctions forming the blood-testis barrier between Sertoli cells themselves and with germ cells were deregulated in Sc-α1AMPK-KO. In conclusion, these results suggest that dysregulation of the energy sensing machinery exclusively through disruption of α1AMPK in Sertoli cells translates to a reduction in the quality of germ cells and fertility.

Effects of mono-(2-ethylhexyl) phthalate (MEHP) on chicken germ cells cultured in vitro.

In recent decades, many toxicological tests based on in vivo or in vitro models, mainly from mammalian (rat-mouse) and fish species, were used to assess the risks raised by contact or ingestion of molecules of pharmaceutical, agricultural, or natural origin. But no, or few, in vitro tests using other non-mammalian models such as bird have been explored despite their advantages: the embryonic gonads of birds have a high plasticity of development sensitive to estrogen, and sperm production is nearly two times faster than in rodents. Hence, we have established an in vitro culture of germ cells and somatic cells from chicken post-natal testis, and we have evaluated the sensitivity against the endocrine disruptor compound mono-(2-ethylhexyl) phthalate (MEHP) in comparison to previous studies using rodent and human models. After 96 h of exposure in presence of 10 µM MEHP, chicken seminiferous tubules cultures present a structural alteration, a reduction in cell proliferation and in germ cells population. Apoptosis of germ and somatic cells increases in presence of 1 µM MEHP. Furthermore, MEHP does not affect inhibin B and lactate production by Sertoli cells. These results are in accordance with previous studies using rat, mice, or human culture of testicular cells and in similar range of exposures or even better sensitivity for some "end-points" (biological parameters). In conclusion, the establishment of this postnatal testicular cells culture could be considered as an alternative method to in vivo experiments frequently used for evaluating the impact on the terrestrial wildlife species. This method could be also complementary to mammal model due to the limiting number of animals used and its elevated sensitivity.