The role of oxidative stress in chemotherapy-induced gonadotoxicity in a rat model, and the protective effects of Nigella Sativa oil on oxidative stress, the anti-Müllerian hormone level, and apoptosis.

OBJECTIVE: This study aimed to determine the role of oxidative stress (OS) in carboplatin-induced gonadotoxicity and whether Nigella Sativa oil (NSO), an herbal antioxidant, has a protective effect on ovarian apoptosis, OS, and the anti-Müllerian hormone (AMH) level in a rat model. MATERIALS AND METHODS: The study included 24 adult female rats that were divided into 4 treatment groups. Group A saline + saline (sham group); group B: NSO + saline; group C: saline + carboplatin; group D: NSO + carboplatin. Saline, NSO, and carboplatin were administered intraperitoneally 24 and/or 48 h before sacrification as 4 mL/kg, 4 mL/kg, and 80 mg/kg, respectively. Apoptosis, OS parameters, and AMH were measured. RESULTS: Oxidant levels and apoptosis were higher, whereas AMH and the antioxidants were lower in group C than in group A. Apoptosis, OS parameters, and AMH levels were negatively affected by chemotherapy (CTx) in group C whilst improvement in those parameters was observed in group D following NSO pretreatment. The levels of apoptosis and malondialdehyde (MDA), an OS parameter, in group D were lower than in group C as they declined from 34.3% to 8.65% (p = 0.002) and from 199.4 nmol/g tissue to 136.4 nmol/g tissue (p = 0.002), respectively. However, the slight increase in AMH level from 2.7 ng/mL to 3.5 ng/mL due to the NSO effect was not significant between groups C and D. CONCLUSIONS: The present findings show that carboplatin has adverse effects on AMH, ovarian tissue apoptosis, and OS parameters. NSO pretreatment might protect ovarian tissue and decrease CTx-induced ovarian injury by decreasing OS and apoptosis, but the protective effect of NSO on AMH is limited.

A Docosahexaenoic Acid Derivative (N-Benzyl Docosahexaenamide) as a Potential Therapeutic Candidate for Treatment of Ovarian Injury in the Mouse Model.

Commonly used clinical chemotherapy drugs, such as cyclophosphamide (CTX), may cause injury to the ovaries. Hormone therapies can reduce the ovarian injury risk; however, they do not achieve the desired effect and have obvious side effects. Therefore, it is necessary to find a potential therapeutic candidate for ovarian injury after chemotherapy. N-Benzyl docosahexaenamide (NB-DHA) is a docosahexaenoic acid derivative. It was recently identified as the specific macamide with a high degree of unsaturation in maca (Lepidium meyenii). In this study, the purified NB-DHA was administered intragastrically to the mice with CTX-induced ovarian injury at three dose levels. Blood and tissue samples were collected to assess the regulation of NB-DHA on ovarian function. The results indicated that NB-DHA was effective in improving the disorder of estrous cycle, and the CTX+NB-H group can be recovered to normal levels. NB-DHA also significantly increased the number of primordial follicles, especially in the CTX+NB-M and CTX+NB-H groups. Follicle-stimulating hormone and luteinizing hormone levels in all treatment groups and estradiol levels in the CTX+NB-H group returned to normal. mRNA expression of ovarian development-related genes was positive regulated. The proportion of granulosa cell apoptosis decreased significantly, especially in the CTX+NB-H group. The expression of anti-Müllerian hormone and follicle-stimulating hormone receptor significantly increased in ovarian tissues after NB-DHA treatment. NB-DHA may be a promising agent for treating ovarian injury.

Effects of Bushen Yiqi Huoxue Decoction in Treatment of Patients with Diminished Ovarian Reserve: A Randomized Controlled Trial.

OBJECTIVE: To explore the therapeutic effect of Bushen Yiqi Huoxue Decoction BYHD) in patients with diminished ovarian reserve (DOR). METHODS: A total of 180 patients with DOR diagnosed from December 2013 to December 2014 were equally assigned into progynova and duphaston (E+D) group, Zuogui Pill group and BYHD group with 60 cases in each by computerized randomization. Patients received E+D, Zuogui Pill or BYHD for 12 months, respectively. Follicle stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E2), anti-Müllerian hormone (AMH), antral follicle count (AFC), ovarian volume, endometrial thickness, and the resistance indices (RIs) of ovarian arteries and uterine arteries were observed before and after treatment. RESULTS: Nine women (4 from the E+D group, 3 from the Zuogui Pill group, and 2 from the BYHD group) withdrew from the study. After 6 months, Zuogui Pill and BYHD significantly decreased FSH and LH and increased endometrial thickness and AMH (all P<0.01). BYHD also resulted in E2 elevation (P<0.05), ovary enlargement (P<0.05), AFC increase (P<0.01), and RI of ovarian arteries decrease (P<0.05). After 12 months, further improvements were observed in the Zuogui Pill and BYHD groups (all P<0.01), but BYHD showed better outcomes, with lower FSH, larger ovaries and a thicker endometrium compared with the Zuogui Pill group (all P<0.01). However, E+D only significantly increased endometrial thickness (P<0.01) and no significant improvements were observed in the RI of uterine arteries in the three groups. CONCLUSIONS: BYHD had a favorable therapeutic effect in patients with DOR by rebalancing hormone levels, promoting ovulation, and repairing the thin endometrium. The combination of tonifying Shen (Kidney), benefiting qi and activating blood circulation may be a promising therapeutic strategy for DOR.

Effects of Bushen Yiqi Huoxue Decoction in Treatment of Patients with Diminished Ovarian Reserve: A Randomized Controlled Trial / 中国结合医学杂志

OBJECTIVE@#To explore the therapeutic effect of Bushen Yiqi Huoxue Decoction BYHD) in patients with diminished ovarian reserve (DOR).@*METHODS@#A total of 180 patients with DOR diagnosed from December 2013 to December 2014 were equally assigned into progynova and duphaston (E+D) group, Zuogui Pill group and BYHD group with 60 cases in each by computerized randomization. Patients received E+D, Zuogui Pill or BYHD for 12 months, respectively. Follicle stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E2), anti-Müllerian hormone (AMH), antral follicle count (AFC), ovarian volume, endometrial thickness, and the resistance indices (RIs) of ovarian arteries and uterine arteries were observed before and after treatment.@*RESULTS@#Nine women (4 from the E+D group, 3 from the Zuogui Pill group, and 2 from the BYHD group) withdrew from the study. After 6 months, Zuogui Pill and BYHD significantly decreased FSH and LH and increased endometrial thickness and AMH (all P<0.01). BYHD also resulted in E2 elevation (P<0.05), ovary enlargement (P<0.05), AFC increase (P<0.01), and RI of ovarian arteries decrease (P<0.05). After 12 months, further improvements were observed in the Zuogui Pill and BYHD groups (all P<0.01), but BYHD showed better outcomes, with lower FSH, larger ovaries and a thicker endometrium compared with the Zuogui Pill group (all P<0.01). However, E+D only significantly increased endometrial thickness (P<0.01) and no significant improvements were observed in the RI of uterine arteries in the three groups.@*CONCLUSIONS@#BYHD had a favorable therapeutic effect in patients with DOR by rebalancing hormone levels, promoting ovulation, and repairing the thin endometrium. The combination of tonifying Shen (Kidney), benefiting qi and activating blood circulation may be a promising therapeutic strategy for DOR.

Effect of anti-Müllerian hormone in hypothalamic Kiss-1- and GnRH-producing cell models.

Purpose: Anti-Müllerian hormone (AMH) is one of the local factors involved in follicle development. In addition, AMH and its receptor are broadly expressed throughout the body. In this study, we examined how AMH modifies gene expression of Kiss-1 and GnRH.Materials and methods: mHypoA-50 and mHypoA-55 cells were originated from the hypothalamic anteroventral periventricular nucleus (AVPV) and arcuate nucleus (ARC), respectively, and these cells are known as Kiss-1 (which encodes kisspeptin) expressing cell models. These cells also express gonadotropin-releasing hormone (GnRH) genes. Our experiments were performed useing these cell models.Results: Both mHypoA-50 and mHypoA-55 hypothalamic cells expressed AMH and AMH receptor type 2 (AMHR2). Exogenous AMH failed to alter the expression levels of the Kiss-1 gene in both cell models but significantly increased GnRH gene expression by 1.73 ± 0.2-fold at 100 pM in mHypoA-50 AVPV cells and by 1.74 ± 0.17-fold at 1 nM in mHypoA-55 ARC cells. AMH also augmented GnRH protein expression in both cell models. Similar to the phenomenon observed in the hypothalamic cell lines, 100 pM AMH significantly increased GnRH, but not Kiss-1, mRNA expression in primary cultures of fetal rat brain cells. Kisspeptin-10 (KP10) increased Kiss-1 gene expression in mHypoA-55 ARC cells but this was blocked by AMH. AMH did not alter the expression of the kisspeptin receptor (Kiss1R) or that of neurokinin B or dynorphin A in mHypoA-55 ARC cells.Conclusions: It was demonstrated that AMH participates in hypothalamic-pituitary-gonadal axis control by stimulating GnRH expression. In addition, AMH might be a potent repressor of Kiss-1 gene expression induced by KP10.

Localization, expression and role of Orexin A and its receptor in testes of neonatal mice.

Orexin A (OXA), a hypothalamic neuropeptide, and its receptor (OX1R) are primarily expressed in lateral hypothalamus and are involved in the control of various biological functions. Expressions of OXA and OX1R have also been reported in peripheral organs like gastrointestinal and genital tracts. In the present study, expressions of OXA and OX1R have been observed in the testis of Parkes strain neonatal mice by semi-quantitative RT-PCR and western blot analyses. Immunohistochemical study also revealed their presence on spermatogonia, Sertoli cells and in the interstitium of the testis. In order to understand the role of OXA and OX1R in testicular development, an in vitro study was also performed. For this, binding of OXA to OX1R was blocked using OX1R specific antagonist, SB-334867. Eighteen mice were sacrificed and their testes were cultured in complete media containing vehicle and two doses (0.1 and 4.0µg/ml media) of SB-334867 for 72h in CO2 incubator at 37°C. At the end of culture period, testes were used for western blot and RT-PCR analyses to study the expression of various markers of gonadal development, such as steroidogenic factor 1 (SF-1), Wilms' tumor 1 (Wt1), Mullerian inhibiting substance (MIS) and stem cell factor (SCF). Further, expressions of OXA, OX1R and glucose transporter 3 (GLUT 3) were also studied. A marked increase in the expression of SF-1 and a decrease in the expression of Wt1 at both transcript and protein levels were noted, while there was a decrease in the expression of SCF and MIS at transcript level at both doses of the antagonist; this suggests that blockage of OXA binding to OX1R by SB-334867 affects testicular development. The decrease in expressions of OXA, OX1R and GLUT 3 in the test is in response to both doses of the antagonist points to their down-regulation causing inefficient uptake of glucose by the testicular cells, thereby affecting gonadal development. In conclusion, our results suggest that the binding of OXA to OX1R is important for the development of the testis.