Effects of frutalin on early follicle morphology, ultrastructure and gene expression in cultured goat ovarian cortical tissue.

Frutalin is a galactose-binding lectin that has an irreversible cytotoxic effect on HeLa cervical cancer cells, by inducing apoptosis and inhibiting cell proliferation. It was previously shown that after in vitro incubation, frutalin is internalized into HeLa cells nucleus, which indicates that frutalin apoptosis-inducing activity might be linked with its nuclear localization. Considering that drugs commonly used for cancer treatment have a deleterious effect on germ cells, the aim of this study was to evaluate the effect of frutalin on the activation, survival, ultrastructure and gene expression in follicles cultured within ovarian tissue. Goat ovarian fragments were cultured for 6 days in α-MEM⁺ alone or supplemented with frutalin (1, 10, 50, 100 or 200 µg/ml). Non-culturad and cultured tissues were processed for histological and ultrastructural analysis and they were also stored to evaluate the expression of anti- and pro-apoptotic genes by quantitative polymerase chain reaction (qPCR). The results showed that the frutalin, at all concentrations tested, reduced follicular survival when compared with control medium. Higher concentrations of frutalin (50, 100 or 200 µg/ml) also reduced follicular survival when compared with those tissues cultured with 1 or 10 µg/ml of frutalin. The ultrastructural analysis showed that atretic cultured follicles had retracted oocytes and a large number of vacuoles spread throughout the cytoplasm. In addition, signs of damage of mitochondrial membranes and cristae were observed. Moreover, although a dose-response effect on gene expression has not been observed, when compared with tissues culture in control medium, the presence of frutalin increased in mRNA expression pro-apoptotic genes. In conclusion, frutalin reduces follicular survival at all concentrations tested, its effects being more pronounced when high concentrations of this lectin (50, 100 and 200 µg/ml) are used. Gene expression profile and ultrastrutural features of cultured follicles suggest that follicular death in goat ovarian tissue cultured in presence of frutalin occurs via necrosis.

In vitro differentiation of primordial germ cells and oocyte-like cells from stem cells.

Infertility is the result of failure due to an organic disorder of the reproductive organs, especially their gametes. Recently, much progress has been made on generating germ cells, including oocytes, from various types of stem cells. This review focuses on advances in female germ cell differentiation from different kinds of stem cells, with emphasis on embryonic stem cells, adult stem cells, and induced pluripotent stem cells. The advantages and disadvantages of the derivation of female germ cells from several types of stem cells are also highlighted, as well as the ability of stem cells to generate mature and functional female gametes. This review shows that stem cell therapies have opened new frontiers in medicine, especially in the reproductive area, with the possibility of regenerating fertility.

Effects of jacalin and follicle-stimulating hormone on in vitro goat primordial follicle activation, survival and gene expression.

This study aims to investigate the effects of jacalin and follicle-stimulating hormone (FSH) on activation and survival of goat primordial follicles, as well as on gene expression in cultured ovarian tissue. Ovarian fragments were cultured for 6 days in minimum essential medium (MEM) supplemented with jacalin (10, 25, 50 or 100 µg/ml - Experiment 1) or in MEM supplemented with jacalin (50 µg/ml), FSH (50 ng/ml) or both (Experiment 2). Non-cultured and cultured tissues were processed for histological and ultrastructural analysis. Cultured tissues from Experiment 2 were also stored to evaluate the expression of BMP-15, KL (Kit ligand), c-kit, GDF-9 and proliferating cell nuclear antigen (PCNA) by real-time polymerase chain reaction (PCR). The results of Experiment 1 showed that, compared with tissue that was cultured in control medium, the presence of 50 µg/ml of jacalin increased both the percentages of developing follicles and viability. In Experiment 2, after 6 days, higher percentages of normal follicles were observed in tissue cultured in presence of FSH, jacalin or both, but no synergistic interaction between FSH and jacalin was observed. These substances had no significant effect on the levels of mRNA for BMP-15 and KL, but FSH increased significantly the levels of mRNA for PCNA and c-kit. On the other hand, jacalin reduced the levels of mRNA for GDF-9. In conclusion, jacalin and FSH are able to improve primordial follicle activation and survival after 6 days of culture. Furthermore, presence of FSH increases the expression of mRNA for PCNA and c-kit, but jacalin resulted in lower GDF-9 mRNA expression.