Ginseng alleviates folliculogenesis disorders via induction of cell proliferation and downregulation of apoptotic markers in nicotine-treated mice.

BACKGROUND: Ginseng is a powerful phytoestrogen with high antioxidant properties. OBJECTIVE: This study aimed to evaluate the effect of Panax Ginseng (PG) on folliculogenesis, proliferation, and apoptosis in the ovary impaired by nicotine. METHODS: Forty adult mice were divided into five groups. Control, sham, and nicotine groups, and co-treated groups of nicotine and ginseng in doses of 0.5 and 1 g/kg. Folliculogenesis was assessed via histopathology and serum evaluation of estradiol, progesterone, follicle-stimulating hormone (FSH), and luteinizing hormone (LH) by ELISA. Lipid peroxidation and antioxidant enzyme activities both in homogenate tissue and serum were assayed by colorimetric analysis. Apoptotic markers of cytochrome c (Cyt c), Bax, and Bcl-2 were evaluated by RT-PCR. Proliferative index was studied by the Ki-67 immunostaining procedure. RESULTS: In comparison to the control or sham groups, nicotine significantly reduced the levels of FSH, LH, and estradiol hormones. An insignificant reduction was observed in the progesterone hormone. Nicotine reduced all healthy follicle numbers, except primordial (P = 0.001). Malondialdehyde (MDA) was increased in tissue and serum in the nicotine group (P = 0.01). Serum catalase (CAT) and enzymatic activity of superoxide dismutase (SOD) both were reduced in tissue and the serum, in the nicotine group. Nicotine induced a reduction in the proliferative indexes of granulosa and theca cells in pre-antral and antral follicles (P = 0.001). However, its effect on the proliferative index of stroma cells was not significant. Apoptotic markers were elevated in the nicotine group (P = 0.001). Co-treatment with ginseng elevated all sex hormones, increased healthy follicles, and reduced tissue or serum lipid peroxidation, compared with the nicotine group (p < 0.05). Co-Treatment with ginseng also reduced the expression of apoptotic markers and increased the proliferative indexes in granulosa and theca cells in pre-antral and antral follicles and also in stroma cells, in comparison to the nicotine group (P = 0.001). All above-mentioned alterations following treatment with ginseng were remarkable, especially in the dose of 1 g/kg. CONCLUSION: This study showed ginseng protects folliculogenesis via alteration of hypothalamic- pituitary-gonadal (HPG) axis, induction of proliferation in ovarian somatic cells, reduction of lipid peroxidation, and downregulation of apoptotic markers in the mouse ovary, treated with nicotine.

Long non-coding RNA Xist regulates oocyte loss via suppressing miR-23b-3p/miR-29a-3p maturation and upregulating STX17 in perinatal mouse ovaries.

The fecundity of female mammals is resolved by the limited size of the primordial follicle (PF) pool formed perinatally. The establishment of PF pool is accompanied by a significant programmed oocyte death. Long non-coding RNAs (lncRNA) are central modulators in regulating cell apoptosis or autophagy in multiple diseases, however, the significance of lncRNAs governing perinatal oocyte loss remains unknown. Here we find that Yin-Yang 1 (YY1) directly binds to the lncRNA X-inactive-specific transcript (Xist) promoter and facilitates Xist expression in the perinatal mouse ovaries. Xist is highly expressed in fetal ovaries and sharply downregulated along with the establishment of PF pool after birth. Gain or loss of function analysis reveals that Xist accelerates oocyte autophagy, mainly through binding to pre-miR-23b or pre-miR-29a in the nucleus and preventing the export of pre-miR-23b/pre-miR-29a to the cytoplasm, thus resulting in decreased mature of miR-23b-3p/miR-29a-3p expression and upregulation miR-23b-3p/miR-29a-3p co-target, STX17, which is essential for timely control of the degree of oocyte death in prenatal mouse ovaries. Overall, these findings identify Xist as a key non-protein factor that can control the biogenesis of miR-23b-3p/miR-29a-3p, and this YY1-Xist-miR-23b-3p/miR-29a-3p-STX17 regulatory axis is responsible for perinatal oocyte loss through autophagy.

Evidences for Red Pigment Concentrating Hormone (RPCH) and Beta-Pigment Dispersing Hormone (ß-PDH) Inducing Oocyte Meiotic Maturation in the Chinese Mitten Crab, Eriocheir sinensis.

Red pigment concentrating hormone (RPCH) and pigment dispersing hormone (PDH) are crustacean neuropeptides involved in broad physiological processes including body color changes, circadian rhythm, and ovarian growth. In this study, the full-length cDNA of RPCH and PDH were identified from the brain of the Chinese mitten crab Eriocheir sinensis. The deduced RPCH and PDH mature peptides shared identical sequence to the adipokinetic hormone/RPCH peptides family and the ß-PDH isoforms and were designated as Es-RPCH and Es-ß-PDH, respectively. Es-RPCH and Es-ß-PDH transcripts were distributed in the brain and eyestalks. The positive signals of Es-RPCH and Es-ß-PDH were localized in the neuronal clusters 6, 8, 9, 10, and 17 of the brain as revealed by in situ hybridization. The expression level of Es-RPCH and Es-ß-PDH mRNA in nervous tissues were all significantly increased at vitellogenic stage, and then decreased at the final meiotic maturation stage. The administrated with synthesized Es-RPCH peptide results in germinal vesicles shift toward the plasma membrane in vitellogenic oocyte, and significant decrease of the gonad-somatic index (GSI) and mean oocyte diameter as well as the expression of vitellogenin mRNA at 30 days post injection in vivo. Similar results were also found when injection of the Es-ß-PDH peptide. In vitro culture demonstrated that Es-RPCH and Es-ß-PDH induced germinal vesicle breakdown of the late vitellogenic oocytes. Comparative ovarian transcriptome analysis indicated that some reproduction/meiosis-related genes such as cdc2 kinase, cyclin B, 5-HT-R and retinoid-X receptor were significantly upregulated in response to Es-RPCH and Es-ß-PDH treatments. Taken together, these results provided the evidence for the inductive effect of Es-RPCH and Es-ß-PDH on the oocyte meiotic maturation in E. sinensis.

Understanding Transcriptomic and Serological Differences between Forced Molting and Natural Molting in Laying Hens.

Molting is natural adaptation to climate change in all birds, including chickens. Forced molting (FM) can rejuvenate and reactivate the reproductive potential of aged hens, but the effect of natural molting (NM) on older chickens is not clear. To explore why FM has a dramatically different effect on chickens compared with NM, the transcriptome analyses of the hypothalamus and ovary in forced molted and natural molted hens at two periods with feathers fallen and regrown were performed. Additionally, each experimental chicken was tested for serological indices. The results of serological indices showed that growth hormone, thyroid stimulating hormone, and thyroxine levels were significantly higher (p < 0.05) in forced molted hens than in natural molted hens, and calcitonin concentrations were lower in the forced molted than in the natural molted hens. Furthermore, the transcriptomic analysis revealed a large number of genes related to disease resistance and anti-aging in the two different FM and NM periods. These regulatory genes and serological indices promote reproductive function during FM. This study systematically revealed the transcriptomic and serological differences between FM and NM, which could broaden our understanding of aging, rejuvenation, egg production, and welfare issues related to FM in chickens.

Identification, functional characterization, and estrogen regulation on gonadotropin-releasing hormone in the spotted scat, Scatophagus argus.

Gonadotropin-releasing hormone (GnRH) is a key neuropeptide of the reproductive system. However, little is known about the role of GnRH in the spotted scat (Scatophagus argus). Here, three GnRH subtypes (cGnRH-II, sGnRH, and sbGnRH) were identified in the spotted scat. cGnRH-II and sGnRH were only expressed in the brains and gonads of both male and female fish, exhibiting a tissue-specific expression pattern, while sbGnRH was expressed at different transcription levels in all examined tissues. During ovarian maturation, hypothalamus-associated sbGnRH was upregulated, while the expression of sGnRH was variable and cGnRH-II first increased and then decreased. In vivo experiments showed that sbGnRH significantly promoted the expression of fsh and lh genes in a dose-dependent manner and exhibited a desensitization effect on lh expression at high concentrations. For sGnRH and cGnRH-II, only high concentrations could induce fsh and lh expression. Furthermore, treatment with highly concentrated sbGnRH peptide also induced fsh and lh expression, whereas the sGnRH and cGnRH-II peptides only induced fsh expression in vitro. 17ß-Estradiol (E2) significantly inhibited the expression of sbGnRH mRNA in a dose-dependent manner and did not impact sGnRH and cGnRH-II mRNA levels in vivo or in vitro. The inhibitory effect of E2 on sbGnRH expression was attenuated by the estrogen receptor (ER) broad-spectrum antagonist (fulvestrant) and the ERα-specific antagonist (methyl-piperidinopyrazole), respectively, implying that the feedback regulation on sbGnRH is mediated via ERα. This study provides a theoretical basis for the reproductive endocrinology of the spotted scat by studying GnRH.

Expression profiles of types 2 and 3 iodothyronine deiodinase genes in relation to vitellogenesis in a tropical damselfish, Chrysiptera cyanea.

Thyroid hormone (TH) is involved in regulating the reproduction of vertebrates. Its physiological action in the target tissues is due to the conversion of TH by iodothyronine deiodinases. In this study, we aimed to clone and characterize type 2 (sdDio2) and type 3 (sdDio3) of the sapphire devil Chrysiptera cyanea, a tropical damselfish that undergoes active reproduction under long-day conditions, and to study the involvement of THs in the ovarian development of this species. When the cDNAs of sdDio2 and sdDio3 were partially cloned, they had deduced amino acid sequences of lengths 271 and 267, respectively, both of which were characterized by one selenocysteine residue. Real-time quantitative PCR (qPCR) revealed that both genes are highly expressed in the whole brain, and sdDio2 and sdDio3 are highly transcribed in the liver and ovary, respectively. In situ hybridization analyses showed positive signals of sdDio2 and sdDio3 transcripts in the hypothalamic area of the brain. Little change in mRNA abundance of sdDio2 and sdDio3 in the brain was observed during the vitellogenic phases. It is assumed that simultaneous activation and inactivation of THs occur in this area because oral administration of triiodothyronine (T3), but not of thyroxine (T4), upregulated mRNA abundance of both genes in the brain. The transcript levels of sdDio2 in the liver and sdDio3 in the ovary increased as vitellogenesis progressed, suggesting that, through the metabolism of THs, sdDio2 and sdDio3 play a role in vitellogenin synthesis in the liver and yolk accumulation/E2 synthesis in the ovary. Taken together, these results suggest that iodothyronine deiodinases act as a driver for vitellogenesis in tropical damselfish by conversion of THs in certain peripheral tissues.

Sodium nitrite negatively affects reproductive ability and offspring survival in female mice.

Sodium nitrite (NaNO2) is an industrial chemical that is frequently used as a food additive to prevent botulism and enhance glossiness, such as curing meat. In addition, in some regions, water source NaNO2 concentrations exceed standard regulatory levels. Whether the excessive intake of NaNO2 has toxic effects on female fertility and fetal development remain unknown. In this study, we administered ICR mice control saline, low-dose NaNO2 (60 mg/kg/day), or high-dose NaNO2 (120 mg/kg/day) by intragastric gavage for 21 days. We then assessed oocyte morphology, spindle-chromosome dynamics, mitochondrial distribution, ATP content, apoptotic cell numbers, DNA damage levels, histone modifications, reactive oxygen species (ROS) levels, and offspring survival. Results showed that NaNO2 treatment decreased oocyte number, impaired polar body extrusion, and increased zona pellucida thickness in oocytes. Furthermore, NaNO2 disrupted MII spindle integrity, caused abnormal mitochondrial distribution, decreased ATP content, and increased levels of ROS and H3K4me2. Moreover, the number of oocytes in early stages of apoptosis and with levels of DNA damage increased in NaNO2-treated mice along with decreased offspring numbers and survival rates. We demonstrated the negative effects of NaNO2 on female reproductive abilities in mice.

Kuntai capsule attenuates premature ovarian failure through the PI3K/AKT/mTOR pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Kuntai capsule (KTC), a type of herb formulas, was first described in the book of Shang Han Za Bing Lun in the third century. KTC has been widely used for the clinical treatment of menopausal syndrome. Considering that premature ovarian failure is also known as premature menopause, this study was designed to investigate the effects and mechanisms of KTC on a mouse model of premature ovarian failure. MATERIALS AND METHODS: Forty-five female C57BL/6 mice were chosen for this study. Fifteen of the mice were separated into the Control group. The remaining thirty were used to establish the premature ovarian failure model by injecting intraperitoneally with 75 mg/kg cyclophosphamide and then by randomly dividing the mice into two groups. One group was considered the Model group, the other group treated with the Kuntai capsule intragastrically every day for one week called the KTC group. After treatment, mice were sacrificed for sampling. The ovaries morphology of mice was observed by hematoxylin and eosin (HE) staining, and all follicles were counted under microscope. Western blotting was used to detect the PI3K/AKT/mTOR pathway activation. Serum follicle-stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E2) and anti-mullerian hormone (AMH)levels were measured by enzyme-linked immunosorbent assay (ELISA). The fertility was observed by giving treated mice 8 weeks for breeding. RESULTS: We found that primordial follicle counts were increased in the KTC group compared to the Model group. The phosphorylation of PI3K, AKT, mTOR, 4E-BP1 and S6K in the KTC group significantly reduced compared to Model group. Serum FSH and LH levels in the KTC group were decreased compared to the Model group, while, serum E2 and AMH levels in the KTC group were increased compared with the Model group. The litter size in the KTC group was improved compared to Model group. CONCLUSIONS: The KTC showed protective potentials of ovarian reserve and fertility to attenuate premature ovarian failure, which was relatively associated with activation of the PI3K/AKT/mTOR signaling pathway.

Comparative Transcriptome Analysis Reveals Differentially Expressed Genes and Signaling Pathways Between Male and Female Red-Tail Catfish (Mystus wyckioides).

Sexual dimorphism is widespread in fish species. The red-tail catfish (Mystus wyckioides) is a commercially important catfish in the lower reaches of the Lancang River and the Mekong basin, and it shows a growth advantage in males. Here, RNA-seq was for the first time used to explore the gene expression difference between the sexes in the hypothalamus and pituitary of red-tail catfish, respectively. In the hypothalamus, 5732 and 271 unigenes have significantly higher and lower expressions, respectively, in males compared with females. KEGG analysis showed that 212 DEGs were annotated to 216 signaling pathways, and enrichment analysis suggested different levels of cAMP and glutamatergic synapse signaling between male and female hypothalami and some of the DEGs appear involved in gonad development and growth. In the pituitary, we found only 19 differentially expressed unigenes, which were annotated to 32 signaling pathways, most of which play important roles in gonad development.

Maternal ß-hydroxy-ß-methylbutyrate (HMB) supplementation during pregnancy affects early folliculogenesis in the ovary of newborn piglets.

Beta-hydroxy-beta-methylbutyrate (HMB) is a leucine metabolite with protein anabolic effects. This study was designed to determine whether prenatal HMB treatment has an effect on oogenesis and folliculogenesis in the ovary of newborn piglets. HMB decreased the number of egg nests and primordial follicles and increased the pool of developing follicles compared to the control group. Although the percentage of TUNEL-positive oocytes within the egg nests was higher in HMB-treated group no increase in the Bax/Bcl-2 ratio and active caspase-3 expression was observed. Moreover, the granulosa cell proliferation index and StAR protein expression were higher in HMB-treated group. In contrast to the control group, the expression of E-cadherins was reduced after the HMB treatment. In addition, a significant increase in the serum level of gonadotropins and steroid hormones was detected in HMB-treated piglets. In conclusion, prenatal HMB treatment dysregulates hormonal homeostasis which impairs early folliculogenesis in piglets.

Effect of photoperiod on ovarian morphology, reproductive hormone secretion, and hormone receptor mRNA expression in layer ducks during the pullet phase.

We evaluated the effect of photoperiod on ovarian morphology, reproductive hormone secretion, and hormone receptor mRNA expression in layer ducks during the pullet phase. A total of 480 71-d-old Jinding layer ducks were randomly divided into 5 groups that received 6L (hours of light):18D (hours of darkness), 8L:16D, 10L:14D, 12L:12D, or 14L:10D, respectively. Each group had 6 replicates with 16 birds each. The photoperiod feeding trial lasted 80 d until 150 d of age. The age at first egg (AFE), the total number, and weight of eggs increased linearly with increasing photoperiods (P < 0.05); lower values of AFE occurred with photoperiods ≥8 h, whereas a higher total number and weight of eggs occurred with photoperiods ≥10 h, compared with 6L:18D (P > 0.05). Oviduct weight, ovary percentage, and initial and bare stroma (weight and percentage) increased quadratically with increasing photoperiods (P < 0.05), and 10.24, 10.01, and 10.10 h were the optimal photoperiods for oviduct weight, bare stroma (follicles ≥2 mm in diameter removed) weight, and bare stroma percentage, respectively, as calculated from reliable regression equations (R2 ≥ 0.5791). Compared with 6L:18D, 10L:14D had a higher total large white follicle weight, small yellow follicle number, and weight (P < 0.05). In addition, higher serum levels of follicle-stimulating hormone, luteinizing hormone, and progesterone were observed with ≥10-h photoperiods (P < 0.05), as were levels of hormone receptor mRNA expression in ovarian follicles (P < 0.05), with the highest values for both measures at 10L:14D. In the hypothalamus, mRNA expression of gonadotropin-releasing hormone increased in ≥8-h photoperiods, with the highest value at 10L:14D. In contrast, gonadotropin-inhibitory hormone increased in photoperiods ≥12 h (P < 0.05). In conclusion, an appropriate photoperiod led to early sexual maturity and improved the development of reproductive organs and ovarian follicles through effects on reproductive hormones and their receptors; 10 to 10.24 h is an adequate photoperiod for layer ducks during the pullet phase.

A unified scale for female reproductive stages in the Norway lobster (Nephrops norvegicus): Evidence from macroscopic and microscopic characterization.

Knowledge of the reproductive cycle in exploited species is important for a sustainable management of fisheries. Standardized scales to assess maturity stages are a fundamental tool to understand the demographic composition of exploited populations. Staging scales for female Norway lobster, Nephrops norvegicus, have been subject to a series of changes, and multiple inconsistent scales are in use in different fisheries regions. A unified, evidence-based scale has not previously been established. We reviewed previous staging scales for the female ovary maturation and propose a revised scale based on the correlation between macroscopic and microscopic ovary characteristics. To provide better-informed tools for future stock assessment, female stages were characterized through external observation on ovary color and size, and the progress of vitellogenesis. This study clarifies several biological phases that were conflated in previous scales. First, we demonstrate how to distinguish between immature ovaries in juvenile females versus the earliest ovary maturation stage in adults. Second, the new scale differentiates between "mottled" ovaries seen in two separate biological stages: the spent ovaries that undergo partial resorption in berried females, versus ovaries of females which failed to spawn and undergo full resorption. To ensure consistent application, colors are assessed relative to international standards (RAL/Pantone). This new, practical staging scheme clarifies the correlation between microscopic characteristics and macroscopically observable details in ovary maturation. Adoption of this unified staging scale will improve maturity analyses, help to identify stocks with potentially reduced reproductive capacity, and facilitate broad-scale comparisons.

Effects of dietary crude oil exposure on molecular and physiological parameters related to lipid homeostasis in polar cod (Boreogadus saida).

Polar cod is an abundant Arctic key species, inhabiting an ecosystem that is subjected to rapid climate change and increased petroleum related activities. Few studies have investigated biological effects of crude oil on lipid metabolism in this species, despite lipids being a crucial compound for Arctic species to adapt to the high seasonality in food abundance in their habitat. This study examines the effects of dietary crude oil exposure on transcription levels of genes related to lipid metabolism (peroxisome proliferator-activated receptors [ppar-α, ppar-γ], retinoic X receptor [rxr-ß], palmitoyl-CoA oxidase [aox1], cytochrome P4507A1 [cyp7α1]), reproduction (vitellogenin [vtg-ß], gonad aromatase [cyp19a1]) and biotransformation (cytochrome P4501A1 [cyp1a1], aryl hydrocarbon receptor [ahr2]). Exposure effects were also examined through plasma chemistry parameters. Additional fish were exposed to a PPAR-α agonist (WY-14,643) to investigate the role of PPAR-α in their lipid metabolism. The dose-dependent up-regulation of cyp1a1 reflected the activation of genes related to PAH biotransformation upon crude oil exposure. The crude oil exposure did not significantly alter the mRNA expression of genes involved in lipid homeostasis except for cyp7α1 transcription levels. Plasma levels of cholesterol and alanine transaminase showed significant alterations in fish exposed to crude oil at the end of the experiment. WY exposure induced a down-regulation of ppar-α, an effect contrary to studies performed on other fish species. In conclusion, this study showed clear effects of dietary crude oil exposure at environmentally relevant concentrations on xenobiotic biotransformation but revealed only weak alterations in the lipid metabolism of polar cod.

Effectiveness of ZiYin Xiehuo granules and Zishen Qinggan granules on partial precocious puberty in girls: a multicenter, randomized, single-blind, controlled trial.

OBJECTIVE: To evaluate the effect of ZiYin Xiehuo granules (ZYXH) and Zishen Qinggan granules (ZSQG) on partial precocious puberty (PPP). METHODS: This was a multicenter, randomized, single-blind, positive-controlled trial. A total of 143 patients were assigned to either the ZYXH group or the ZSQG group using a random number table. The ZYXH group received ZYXH three times daily for 6 months and the ZSQG group received ZSQG three times daily for 6 months. Mammary nucleus diameter; the results of uterus, ovarian, and maximum follicle measures; and Chinese medicine symptom pattern scores were compared at baseline and after 3 months and 6 months of treatment. RESULTS: After 3 months' treatment, there were no significant differences between the two groups in mammary nucleus index changes (left 3.4 ± 3.1 vs 3.5 ± 3.1, P = 0.790; right 3.0 ± 2.9 vs 3.6 ± 3.0, P = 0.719). The uterine volume in the ZYXH group was smaller than that in the ZSQG group (2.1 ± 1.6 vs 2.6 ± 2.2, P = 0.006). There were no significant between-group differences in ovarian volume and maximum follicular diameter on either side (ovarian volume: left 1.2 ± 0.7 vs 1.3 ± 0.6, P = 0.8; right 1.2 ± 0.7 vs 1.4 ± 1.1, P = 0.984; maximum follicular diameter: left 3.9 ± 1.7 vs 3.5 ± 2.2, P = 0.158; right 3.5 ± 1.7 vs 3.9 ± 2.1, P = 0.314). CONCLUSION: ZYXH granules and ZSQG granules both affected the size of the mammary nucleus in girls with PPP, and improved Chinese medicine symptom patterns. ZYXH granules showed slight advantages over ZSQG granules in terms of the decrease in the size of the uterus, ovaries, and ovarian follicles.

Seasonal Reproductive Biology of Drosophila suzukii (Diptera: Drosophilidae) in Temperate Climates.

Drosophila suzukii (Matsumura; Diptera: Drosophilidae) is a key pest of sweet cherry and small fruits worldwide. The present studies were designed to describe the reproductive physiology in both sexes, through dissections of their reproductive organs. We extensively dissected female D. suzukii throughout the season from 2013 to 2016 and classified the reproductive status flies based on five recognizable ovarian maturation stages: 1) no ovaries; 2) unripe ovaries 3) ripening eggs in ovarioles; 4) mature eggs in ovarioles; and 5) mature eggs in the abdomen. Development was examined as a function of calendar days as well as degree-days (DD). Results obtained from winter collections revealed that females collected from November to March contained a lower percentage of mature eggs than females collected from April to September. These data suggest that environmental conditions during the dormant period induce reproductive diapause. Oogenesis likely increased with an increase in mean monthly temperatures and DD. The first overwintered females with mature eggs were dissected as early as 21 February 2014 in Trento (7 DD). Additionally, we found that a low proportion of males (less than 50%) had sperm in their testes between January and March, yet during the same period females already have sperm stored in their spermathecal. Ivy berries was an alternative but unfavorable non-crop host during the late dormant period, as evidenced by emergence of smaller adults when compared to individuals emerging from cherry fruits. This study showed that D. suzukii females have great potential for oviposition early in the season, posing a risk to early season maturing crop hosts.

Preconception exposure to dietary levels of genistein affects female reproductive outcomes.

Genistein is a phytoestrogen found in soy and soy-based products. Previously, we found that genistein adversely affected estradiol levels and follicle growth in vitro. Proper hormone production and follicle growth are key regulators of normal fertility. Therefore, we hypothesized that genistein adversely affects female fertility and pregnancy outcomes. To test this hypothesis, we dosed sexually mature female CD-1 mice (35days) with 0, 300, 500, or 1000ppm genistein for 30, 60, 150, and 240days. At the end of the dosing periods, we measured mating rate, pregnancy rate, fertility rate, gestation time, parturition time, pup mortality, litter size, average pup weight, and estradiol and progesterone levels. We found that chronic, preconception exposure to genistein affects gestation time, parturition time, litter size, pup weight, and pup mortality. Additionally, genistein exposure for 240days appears to have a protective effect on fertility rate, but does not affect hormone levels in vivo.

Sodium fluoride adversely affects ovarian development and reproduction in Drosophila melanogaster.

The study demonstrates the effects of chronic sub-lethal exposure of sodium fluoride (NaF) on reproductive structure and function of female Drosophila melanogaster. As a part of treatment, flies were maintained in food supplemented with sub-lethal concentrations of NaF (10-100 µg/mL). Fecundity, ovarian morphology, presence and profusion of viable cells from ovary and fat body were taken into consideration for evaluating changes in reproductive homeostasis. Wing length (a factor demonstrating body size and reproductive fitness) was also monitored after NaF exposure. Significant reduction in fecundity, alteration in ovarian morphology along with an increase in apoptosis was observed in treated females. Simultaneous decline in viable cell number and larval weight validates the result of MTT assay. Furthermore, altered ovarian Glucose-6-phosphate dehydrogenase and catalase activities together with increased rate of lipid peroxidation after 20 and 40 µg/mL NaF exposure confirmed the changes in reproduction related metabolism. Enhanced lipid peroxidation known for ROS generation might have induced genotoxicity which is confirmed through Comet assay. The enzyme activities were not dose dependent, rather manifested a bimodal response, which suggests a well-knit interaction among the players inducing stress and the ones that help establish physiological homeostasis.

The Application of Dehydroepiandrosterone on Improving Mitochondrial Function and Reducing Apoptosis of Cumulus Cells in Poor Ovarian Responders.

Poor ovarian responders (PORs) pose a great challenge for in vitro fertilization (IVF). Previous studies have suggested that dehydroepiandrosterone (DHEA) may improve IVF outcomes in PORs. The current study attempted to investigate the clinical benefits of DHEA in PORs and the possible mechanisms of DHEA on cumulus cells (CCs). This was a prospective study performed at one tertiary center from January 2015 to March 2016. A total of 131 women who underwent IVF treatment participated, including 59 normal ovarian responders (NORs) and 72 PORs. PORs were assigned to receive DHEA supplementation or not before the IVF cycle. For all patients, CCs were obtained after oocyte retrieval. In the CCs, mRNA expression of apoptosis-related genes and mitochondrial transcription factor A (TFAM) gene, terminal deoxynucleotidyl transferase dUTP nick end labeling assay, mitochondrial dehydrogenase activity and mitochondrial mass were measured. The results indicated that PORs with DHEA supplementation produces a great number of top-quality embryos at day 3 and increased the number of transferred embryos and fertilization rate compared with those without DHEA supplementation. Additionally, supplementation with DHEA in PORs decreased DNA damage and apoptosis in CCs while enhancing the mitochondrial mass, mitochondrial dehydrogenase activity and TFAM expression in CCs. In conclusion, our results showed that the benefits of DHEA supplementation on IVF outcomes in PORs were significant, and the effects may be partially mediated by improving mitochondrial function and reducing apoptosis in CCs.

Effect of injectable trace mineral complex supplementation on development of ovarian structures and serum copper and zinc concentrations in over-conditioned Holstein cows.

This study evaluated the effect of injecting trace minerals on reproductive performance in over-conditioned Holstein cows before synchronized estrus. Multiparous non-lactating, over-conditioned repeat breeder cows (n=20) were assigned randomly to one of two treatments: 1) control (n=10), and 2) supplementation with an injectable trace mineral complex 25days before expected synchronized estrus (n=10). Follicular waves were synchronized by intravaginal insertion of a CIDR for eight days and an intramuscular (i.m.) injection of a GnRH analogue. Estrus was induced at CIDR removal by an i.m. injection of PGF2α. Blood samples were collected before and after synchronized estrus. The response variables were follicle population (FP), diameter of the preovulatory follicle at CIDR removal (DFP0) and at estrus detection (DFP1), time of estrus after CIDR removal (TE), area of corpus luteum (ACL), pregnancy rate and copper and zinc serum concentrations. The statistical analysis of the variables was carried out with SAS. The FP, DFP0, DFP1, TE, ACL and serum concentrations of copper and zinc were not affected by the trace mineral injection (P>0.05). Even though pregnancy rate at 40 (77.78±13.46 vs 44.44±16.56%) and 60days after AI (66.67±15.71 vs 33.33±15.71%) was numerically higher for cows injected with trace minerals than for the control group, the differences were not significant (P>0.05). In conclusion, while follicular and corpus luteum development were not affected by trace mineral injection, it may be a feasible way to increase the pregnancy rate in over-conditioned cows.

Effects and mechanisms of waterborne copper exposure influencing ovary development and related hormones secretion in yellow catfish Pelteobagrus fulvidraco.

The present study was conducted to determine the effects and mechanism of waterborne copper (Cu) exposure influencing ovary development and related hormones secretion in yellow catfish Pelteobagrus fulvidraco. To this end, two experiments were conducted. In Exp. 1, the partial cDNA sequences of three steroidogenesis-related genes (androgen receptor (ar), steroidogenic factor 1 (sf-1) and steroidogenic acute regulatory protein (star)) were firstly characterized from P. fulvidraco. The predicted amino acid sequences for the P. fulvidraco ar, sf-1 and star contained the main structural features characteristic in other species. In Exp. 2, P. fulvidraco were exposed to three waterborne Cu concentrations (control, 30µg/l and 60µg/l, respectively) for 56days. Sampling occurred on day 28 and day 56, respectively. On day 28, the levels of serum sex-steroid hormones (FSH and LH) and the mRNA levels of steroidogenesis-related genes (3ß-hsd, cyp11a1, cyp17, cyp19a, sf-1 and star) were significantly increased in ovary of P. fulvidraco exposed to 30µg Cu/l. The immunohistochemical analysis showed the positive reaction of ER, VTG and aromatase in low dose exposure group. These indicated that in low dose and relative short-term exposure, Cu was beneficial. In contrast, 60µg Cu/l exposure significantly reduced the levels of serum FSH, LH, E2 and P, and the mRNA levels of ovarian 20ß-hsd, cyp19a and erα in P. fulvidraco. On day 56, waterborne Cu concentration exposure reduced the levels of serum gonadotropins and sex hormones, and down-regulated the mRNA levels of steroidogenesis-related genes, indicating long-term Cu exposure had toxic effect on the secretion of sex-steroid hormone in P. fulvidraco. For the first time, our study cloned cDNA sequences of ar, sf-1 and star in P. fulvidraco, and demonstrated the effects and mechanism of waterborne Cu exposure influencing hormones secretion and synthesis in dose- and time-dependent manner in P. fulvidraco, which will help to understand the Cu-induced reproductive toxicity at both protein and transcriptional levels in fish.