Effect of reduced water intake on ovarian reserve, leptin immunoexpression and impact of leptin on the in vitro culture of sheep secondary follicles.

This study evaluated the effect of reduced water intake on survival, apoptosis and immunoexpression of leptin in sheep preantral follicles, activation of primordial follicles, serum levels of leptin, estradiol (E2) and progesterone (P4), and in vitro maturation (IVM) of oocytes antral follicles, as well evaluated the effects of leptin on in vitro culture of secondary follicles isolated these animals. Ewes (n = 32) were divided into four groups: water ad libitum (Control - 100%), 80%; 60% and 40% of ad libitum intake. Blood was collected to determine, leptin, E2 and P4, before and after experiment. After the slaughter, ovarian cortex was used to histological and immunohistochemistry analysis and oocytes IVM. Moreover, isolated secondary follicles were cultured in vitro for 12 days in control medium (α-MEM+) or α-MEM+ with 10 or 25 ng/mL leptin. The reduction of water intake caused a linear decreasing effect on the percentages of normal preantral follicles, especially of primordial (P < 0.05), increased the apoptosis (P < 0.05) and decreased leptin expression in preantral follicles. The treatment with 60% of water intake showed greater total growth rate of isolated secondary follicles cultured with 25 ng/L leptin (P < 0.05), compared to those cultured in α-MEM+ . In conclusion, reduced water intake impaired the number of normal sheep preantral follicles, especially of primordial follicles, increased apoptosis and decreased leptin expression in preantral follicles. Moreover, secondary follicles from of ewes that receive 60% water intake increased follicular growth after in vitro culture with 25 ng/mL leptin.

Impact of antioxidant supplementation during in vitro culture of ovarian preantral follicles: A review.

The in vitro culture systems of ovarian preantral follicles have been developed for studying follicular and oocyte growth, for future use of immature oocytes as sources of fertilizable oocytes and for screening ovarian toxic substances. One of the key limitations of the in vitro culture of preantral follicles is the oxidative stress by accumulation of reactive oxygen species (ROS), which can impair follicular development and oocyte quality. Several factors are associated with oxidative stress in vitro, which implies the need for a rigorous control of the conditions as well as addition of antioxidant agents to the culture medium. Antioxidant supplementation can minimize or eliminate the damage caused by ROS, supporting follicular survival and development and producing mature oocytes competent for fertilization. This review focuses on the use of antioxidants and their role in preventing follicular damage caused by oxidative stress in the in vitro culture of preantral follicles.

Protective effect of gallic acid on doxorubicin-induced ovarian toxicity in mouse.

The aims of the present study were to evaluate the protective effects of gallic acid against doxorubicin-induced ovarian toxicity in mice, and to verify the possible involvement of PI3K and mTOR signaling pathway members (PTEN, Akt, FOXO3a and rpS6) in the gallic acid protective actions. Mice were pretreated with NaCl (0.15 M, p.o.) (control and doxorubicin groups) or gallic acid (50, 100 or 200 mg/kg body weight, p.o.) once daily, for 5 days, and on the third day of treatment, after 1 h of treatment administration, the mice received saline solution (i.p.) (control group) or doxorubicin (10 mg/kg of body weight, i.p.). Next, the ovaries were harvested for histological (follicular morphology and activation), fluorescence (GSH and mitochondrial activity), and immunohistochemical (PCNA, cleaved caspase-3, TNF-α, p-PTEN, Akt, p-Akt, p-rpS6 and p-FOXO3a) analyses. The results showed that cotreatment with 50 mg/kg gallic acid plus doxorubicin preserved the percentage of normal follicles and cell proliferation, reduced the percentage of cleaved caspase-3 follicles, prevented inflammation, and increased GSH concentrations and mitochondrial activity compared to doxorubicin treatment alone. Furthermore, cotreatment 50 mg/kg gallic acid plus doxorrubicin increased expression of Akt, p-Akt, p-rpS6 and p-FOXO3a compared to the doxorubicin alone. In conclusion, 50 mg/kg gallic acid protects the mouse ovary against doxorubicin-induced damage by improving GSH concentrations and mitochondrial activity and cellular proliferation, inhibiting inflammation and apoptosis, and regulating PI3K and mTOR signaling pathway.