Protective effects of polydatin on cyclophosphamide-induced ovarian and uterine damage in rats via modulation of hormonal, oxidative, inflammatory, and histopathological alterations.

The effects of repeated controlled ovarian stimulation (COS) on the female reproductive system are still controversial. This study investigated the effects of repeated COS on the ovaries and uterus of mice and its possible mechanism. Female ICR (Institute of Cancer Research) mice were subjected to the COS using pregnant mare serum gonadotropin (PMSG) and human chorionic gonadotropin (hCG) for 1, 3, 5, and 7 cycles. Serum hormone levels, reactive oxidative stress (ROS), 8-hydroxy-2'-deoxyguanosine (8-OHdG), total antioxidant capacity (T-AOC), and superoxide dismutase (SOD) in the mouse ovary and uterus were analyzed by ELISA. The morphology of the ovary and endometrium, ovarian apoptosis, and expressions of the vascular endothelial growth factor (VEGF), leukemia inhibitory factor (LIF), PI3K, AKT, Bax, and Bcl-2 in the ovarian and uterine tissues were tested by hematoxylin-eosin (HE) staining, immunohistochemistry, and western blot. The results showed that repeated COS significantly decreased the hormone level (estradiol, progesterone and anti-Müllerian hormone), high-quality of the MII oocyte ratio, oocyte and embryo number, antioxidant capacity (T-AOC, SOD activity), and the protein level of Bcl-2, LIF, and VEGF, but increased the oxidative damage (ROS, 8-OHdG content), embryo fragment ratio, and expression of pro-apoptotic protein Bax. In addition, the expressions of p-PI3K and p-AKT also decreased with the increase of COS cycle. In conclusion, repeated COS causes ovarian and uterus damage possibly through the PI3K/AKT signaling pathway, and this finding may provide some experimental basis for guiding clinical treatment.

The combination of medroxyprogesterone acetate (MPA) and gonadotropin chorionic (eCG) has been widely used to synchronize oestrus cycle in sheep, but their effects on the gene expression in uterine tissue are yet to be elucidated. To evaluate the effect of MPA + eCG or prostaglandin analogue (PA) treatments on the rate of oestrus cycle synchronization, as well as further hormone production and gene expression profiles in uterine tissue, 14 Santa Inês ewes were randomly selected. The MPA + eCG group (n=7) received intravaginal insertion of MPA-impregnated sponges for 14 days and was administered 350 IU eCG on the day of sponge withdrawal. The PA group (n=7) was administered two doses of 100 μg of PA separated by 12 days. The ewes were assessed for the rate of oestrus cycle synchronization and the serum concentrations of progesterone (P4) and estradiol (E2). Additionally, the expression of estrogen receptor (ERα), progesterone receptor (P4R), and immunolocalization of interferon receptor (IFNAR1) in the uterine tissue samples collected 15th day post-mating were examined. The rate of oestrus cycle synchronization was 100% (n=7/7) and 57.14% (n=4/7) in the MPA + eCG and PA groups, respectively. Moreover, the MPA + eCG group exhibited higher serum concentration of P4 than the PA group (p < 0.05). However, the E2 serum concentration did not differ between the two groups (p > 0.05). The relative expression of P4R and ERα mRNA analyzed using real-time PCR and immunodetection of IFNAR1 were similar between the two groups tested (p > 0.05). Conclusively, MPA + eCG treatment improved the rate of oestrus cycle synchronization and endogenous P4 production; however, it did not affect the expression of sex steroid receptors and IFNAR1 in uterine ovine tissue.

Background:Embryo implantation relies on precise hormonal regulation, associated gene expression changes, and appropriate female reproductive tract tissue architecture. Female mice exposed neonatally to the phytoestrogen genistein (GEN) at doses similar to those in infants consuming soy-based infant formulas are infertile due in part to uterine implantation defects.Objectives:Our goal was to determine the mechanisms by which neonatal GEN exposure causes implantation defects.Methods:Female mice were exposed to GEN on postnatal days (PND)1–5 and uterine tissues collected on PND5, PND22–26, and during pregnancy. Analysis of tissue weights, morphology, and gene expression was performed using standard histology, confocal imaging with three-dimensional analysis, real-time reverse transcription polymerase chain reaction (real-time RT-PCR), and microarrays. The response of ovariectomized adults to (E2) and artificial decidualization were measured. Leukemia inhibitory factor (LIF) injections were given intraperitoneally and implantation sites visualized. Gene expression patterns were compared with curated data sets to identify upstream regulators.Results:GEN-exposed mice exhibited reduced uterine weight gain in response to E2 treatment or artificial decidualization compared with controls; however, expression of select hormone responsive genes remained similar between the two groups. Uteri from pregnant GEN-exposed mice were posteriorized and had reduced glandular epithelium. Implantation failure was not rescued by LIF administration. Microarray analysis of GEN-exposed uteri during early pregnancy revealed significant overlap with several conditional uterine knockout mouse models, including Foxa2, Wnt4, and Sox17. These models exhibit reduced endometrial glands, features of posteriorization and implantation failure. Expression of Foxa2, Wnt4, and Sox17, as well as genes important for neonatal uterine differentiation (Wnt7a, Hoxa10, and Msx2), were severely disrupted on PND5 in GEN-exposed mice.Discussion:Our findings suggest that neonatal GEN exposure in mice disrupts expression of genes important for uterine development, causing posteriorization and diminished gland function during pregnancy that contribute to implantation failure. These findings could have implications for women who consumed soy-based formulas as infants. https://doi.org/10.1289/EHP6336

ABSTRACTWe investigated the use of melatonin to improve fertility and reduce uterine damage caused by torsion of the uterus in pregnant rats. We used 35 pregnant rats at gestational age 18 days. The animals were randomized into five groups. Group 1 was anesthetized only. Group 2 was subjected to experimental uterine torsion of 360° and the torsion was corrected after 6 h. Group 3 was subjected to uterine torsion of 360°, the torsion was corrected after 6 h and melatonin was administered at the time of correction. Group 4 rats were subjected to 360º uterine torsion and melatonin was administered 6 h later at the time of correction. Group 5 was administered melatonin followed by uterine torsion of 360 degrees followed by correction of torsion 6 h later. Samples were obtained from the uterine horns on the day 1 postpartum. We used Bax, Bcl-2 and caspase 3 staining to measure apoptosis in the uterine tissues. The mRNA levels of Rho-associated, coiled-coil containing protein kinases 1 (ROCK1), homeobox D10 (Hox4 HoxD10), TLR4, NFκB1, caveolin 1 (Cav1) heat shock protein 90 alpha (cytosolic), class B member 1 (Hsp90ab1) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were determined using quantitative real-time polymerase chain reaction analysis (qRT-PCR). Bax, Bcl-2 and caspase 3 were detected using immunohistochemistry. No difference was observed among groups with respect to abortion, neonatal mortality or congenital abnormalities. Compared to the control group, the mRNA levels of Rock1, Hox4, TLR4, NFκB1, Cav1 and Hsp90 genes were decreased significantly in the study groups; the decrease was greater in groups 3 and 4, which were treated with melatonin. The greatest amount of Bax staining was found in group 1 and the least amount of Bcl-2 staining was found in groups 4 and 5; the greatest amount of caspase 3 staining was found in group 2. Our findings indicate that melatonin reduced uterine torsion related tissue damage and that its application during torsion was more effective than application following removal of torsion.

In a mouse model of menstruation stromal cells respond to breakdown of the tissue by changing their identity to become epithelial cells that are incorporated into the luminal epithelium which is rapidly 'healed' without scarring.

In a mouse model of menstruation stromal cells respond to breakdown of the tissue by changing their identity to become epithelial cells that are incorporated into the luminal epithelium which is rapidly 'healed' without scarring.

In utero diethylstilbestrol (DES) exposure has long term consequences including increased risk of vaginal and breast cancer. Epigenetic factors are suspected to contribute to some of these increased risks and animal studies show DES-mediated effects on DNA methylation in both uterine tissue and blood. However, there are currently no studies examining the effect of fetal DES exposure on DNA methylation in humans. Therefore, we investigated differences in DNA methylation between women exposed to DES in utero compared to unexposed women. All our study subjects were participants in the Sister Study -a prospective cohort focused on environmental and familial risk factors for breast cancer and other diseases. Within this study 1,156 women (2.3%) said that they were definitely or probably exposed to DES in utero. In a special sub study (including both women who reported positive exposure and women who reported no exposure), we confirmed exposure history with the woman's mother and from those with matching mother daughter reports we randomly selected 100 exposed and 100 unexposed women. We analyzed DNA methylation patterns in blood using the Illumina 450k array to assess methylation at 473,844 CpG sites. Primary analysis did not reveal any differentially methylated CpGs (dmCpGs) below the false discovery rate (FDR, q<0.05), nonetheless there were about 8000 dmCpGs with p-values <0.05. Stratification of women into subgroups -based on menopausal status, fertility related diagnoses or hormone intake- did not alter the results. Previous work using mouse models shows gene specific changes in gene expression and DNA methylation following perinatal exposure to DES. When we compared these expression changes in ten genes to changes in promoter methylation in our data we found an inverse correlation between gene expression in mouse and the direction of methylation in our samples. This was especially evident in the ERBB2 promoter region. We also examined the subset of genes known to directly interact with DES: in four of the seven direct DES targets (AR, TTR, AFP, and GLI3) we found a majority of the 5’ CpGs had lower methylation levels in exposed women, although these differences were not statistically significant. This is the first study investigating genome wide methylation changes in women subjected to an unusual and very potent prenatal exposure that is known to have severe consequences into adulthood. Study weaknesses include lack of information regarding dose and exact timing of exposure as well as use of whole blood instead of uterine target tissue, but we still observe changes at p<0.05. If DNA methylation changes are detectable in blood decades after exposure this strengthens the idea that DES exposure leads to epigenetic changes and should be possible to verify even further using samples from uterine or vaginal tissue. Citation Format: S Sophia Harlid, Zongli Xu, Vijayalakshmi Panduri, Aimee D'Aloisio, Lisa DeRoo, Dale Sandler, Jack A. Taylor. Fetal exposure to diethylstilbestrol and DNA methylation in adult women. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3643. doi:10.1158/1538-7445.AM2013-3643

To investigate the protective effect of Lycium barbarum polysaccharide (LBP) on reproductive system damage induced by exposure to high-altitude hypoxic environment in female rats, and to explore the mechanisms involved. After undergoing physiological synchronization, 30 female Wistar rats were randomly and evenly assigned to 3 groups, including a plain control (C) group, a high-altitude hypoxia (H) group, and a high-altitude hypoxia + LBP (H-LBP) group. The C group was placed in a region at an altitude of 1500 m above sea level (with an oxygen volume fraction of 18.55%), while the H group and the H-LBP group were placed in a region at an altitude of 4010 m above sea level (with an oxygen volume fraction of 12.70%). Rats in the H-LBP group were fed with LBP at 75 mg/kg via gastric gavage, while the C and H groups received normal saline once a day for 14 days in a row. Changes in estrous cycles were documented throughout the experiment. At the end of the experiment, the serum levels of reproductive hormones and the levels of oxidative stress in the ovarian and uterine tissues were measured. Morphological changes in the ovarian and uterine tissues were assessed using hematoxylin-eosin (HE) staining. A component-target-pathway network diagram was constructed using network pharmacology methods to analyze the key targets and pathways. Compared with the C group, rats in the H group had disrupted estrous cycles and significantly lower serum levels of reproductive hormones (all P<0.05). In addition, rats in the H group had increased oxidative stress damage and experienced pathological damage in the ovarian and uterine tissues. However, compared with those of the H group, the estrous cycle in the H-LBP group became normalized after the administration of LBP and the serum levels of estradiol (E2), progesterone (P), luteinizing hormone (LH), and anti- Müllerian hormone (AMH) were significantly increased in H-LBP group (all P<0.05). In the ovarian tissue, the malondialdehyde (MDA) content was significantly reduced, superoxide dismutase (SOD) activity was increased, and the content of reduced glutathione (GSH) was increased. In addition, in the uterine tissue, the MDA content was reduced and SOD activity was increased (all P<0.05), with LBP significantly improving the pathological damage to the reproductive organs of female rats caused by high-altitude hypoxic environment. Through network pharmacology analysis, we identified 76 potential targets for the protective effect of LBP against high-altitude hypoxia-induced reproductive injury, and the targets were mainly involved in the signaling pathways such as calcium channels, PI3K-Akt, MAPK, and HIF-1. LBP can ameliorate high-altitude hypoxia-induced reproductive damage in female rats. The mechanisms involved may be associated with the regulation of PI3K-Akt, MAPK, and HIF-1 pathways.

Modulatory effects of fenugreek seeds powder on the histopathology, oxidative status, and immune related gene expression in Nile tilapia (Oreochromis niloticus) infected with Aeromonas hydrophila

BackgroundNicotine administration can generate severe oxidative stress and lipid peroxidation. Royal jelly, with its antioxidant properties, acts as a scavenger of reactive oxygen species. This study describes the apitherapy effects of royal jelly on testicular damage following nicotine administration.MethodsForty‐eight male BALB/c mice were divided into 8 groups (n = 6): saline, 3 different doses of royal jelly (100, 150, and 200 mg/kg body weight (BW) per day), nicotine (1.5 mg/kg), and 3 different groups of Nic + Roy (1.5 mg/kg of Nic + 100, 150, and 200 mg/kg BW per day of royal jelly). Nicotine was administrated intraperitoneally, and royal jelly was prescribed orally for 10 consecutive days. Serum levels of hormones (testosterone, luteinizing hormone, and follicle‐stimulating hormone), total antioxidant capacity, nitric oxide (NO) status, malondialdehyde levels, sperm DNA fragmentation, sperm parameters, histopathological changes (H&E staining), immunohistochemistry against apoptotic proteins, and gene expression of Bcl‐2, p53, Caspase‐3, and Nrf2 (real‐time PCR) were assessed to evaluate the molecular and histological changes.ResultsHormone levels, sperm parameters, and status of antioxidants were decreased significantly (p < .05) following nicotine administration. Moreover, royal jelly treatment normalized hormonal and antioxidant characteristics, decreased apoptotic gene expression, increased Nfr2 gene expression, and restored histopathological alteration to the physiological status significantly (p < .05).ConclusionRoyal jelly upregulates the antioxidant status, inhibits the mitochondrial‐dependent apoptosis pathway, and increases the rate of proliferation. This therapeutic agent effectively protected the testis against nicotine‐associated damages by antioxidant and anti‐apoptotic effects.

Objective: To investigate the effect of dietary Spirulina platensis supplementation on growth performance, organ colonization, expression of inflammatory-related genes, and antioxidant status in quail challenged with Salmonella enteritidis.Design: Randomized controlled experimental study.Animals: hundred eighty-seven-day-old Japanese quail.Procedures: birds were randomly allocated into 6 equal groups.G1, G2, and G3: non challenged and fed abasal diet supplemented with Spirulina at 0, 1, and 2 g/kg diet, respectively.G4, G5, and G6: challenged and fed a basal diet supplemented with Spirulina at 0, 1, and 2 g/kg diet, respectively.At 21 days of age, all challenged groups were orally inoculated with 1ml of (1.00x10 7 Salmonella enteritidis/ml).The collected samples were serum for determining biochemical and antioxidant parameters, cecal tissue samples for determination of gene expression of inflammatory-related genes, and tissue samples from liver, heart, spleen, and caecum for bacterial reisolation. Results:The dietary supplementation of Spirulina significantly improved growth performance parameters and reduced organ colonization.The cecal pro-inflammatory gene expressions (IL-6, IL1 ß and TNFα) were significantly downregulated while the anti-inflammatory cytokine (IL-10) was significantly increased.In addition, gene expression of cecal serum amyloid (SAA) was significantly down-regulated.The antioxidant and serum biochemical parameters were improved.Conclusion and clinical relevance: dietary supplementation of Spirulina could be a helpful strategy for mitigating the harmful effects of Salmonella enteritidis in quail.

Simple SummaryThe progression of several inflammatory diseases, including endometritis, has been proven to be enhanced by the process of neutrophil extracellular traps (NETs) formation in pathogenesis. Therefore, averting the formation of NETs could be an effective method to combat endometritis. Cl-amidine has been reported to treat certain inflammatory diseases, but its therapeutic role in combating endometritis remains an enigma as it may likely attenuate NETs formation. Hence, we investigate the potential of NETs formation in the pathogenesis of endometritis using rats and the probable therapeutic indices of Cl-amidine against endometritis. It was revealed that LPS induced endometritis in rats, and the formation of NETs was detected in the rats’ inflamed uterine tissues. The treatment with Cl-amidine showed the regression of LPS-induced endometritis and protection against uterine tissue damage. Cl-amidine effectively reduced the expression of specific proteins that could enhance the formation of NETs, thereby attenuating the inflammatory response to LPS-induced endometritis in rats. Our research showed that Cl-amidine possesses potentially therapeutic properties against endometritis.Endometritis is a common disease that affects the production in dairy cows and leads to severe losses in the dairy industry. Neutrophil extracellular traps (NETs) formation promotes pathogenic invasions of the lumen of the tissue, leading to inflammatory diseases such as mastitis, pancreatitis, and septic infection. However, research that could show the relationship between NETs and endometritis is scarce. Cl-amidine has been shown to ameliorate the disease squealing and clinical manifestation in various disease models. In this study, we investigated the role of NETs in LPS-triggered endometritis in rats and evaluated the therapeutic efficiency of Cl-amidine. An LPS-induced endometritis model in rats was established and found that the formation of NETs can be detected in the rat’s uterine tissues in vivo. In addition, Cl-amidine treatment can inhibit NETs construction in LPS-induced endometritis in rats. Myeloperoxidase (MPO) activity assay indicated that Cl-amidine treatment remarkably alleviated the inflammatory cell infiltrations and attenuated the damage to the uterine tissue. The Western blot results indicated that Cl-amidine decreased the expression of citrullinated Histone H3 (Cit-H3) and high-mobility group box 1 protein (HMGB1) protein in LPS-induced rat endometritis. The ELISA test indicated that Cl-amidine treatment significantly inhibited the expression of the pro-inflammatory cytokines IL-1β, IL-6, and TNF-α. The NETs were determined by Quant-iTTMPicoGreen dsDNA kit®, which indicated that Cl-amidine significantly inhibited the NETs in rat serum. All results showed that Cl-amidine effectively reduced the expression of Cit-H3 and HMGB1 proteins by inhibiting the formation of NETs, thereby attenuating the inflammatory response to LPS-induced endometritis in rats. Hence, Cl-amidine could be a potential candidate for the treatment of endometritis.

Simple SummaryThe beneficial effect of polyunsaturated fatty acids (PUFA) in different body systems and their function in mammals is well known. The use of linseed oil (LO, rich in alpha-linolenic acid/ALA) as a feed additive has been investigated to improve the reproductive and productive performance of different livestock species. However, their effects on the antioxidative defense systems of female reproductive structures are scarcely documented. The results obtained in the present study demonstrate that dietary supplementation of by-pass LO enhances the antioxidative system in luteal and uterine tissues on maternal recognition of the pregnancy period (on Days 14 and 16 after mating) in ewes.This study analyzed the effects of dietary supplementation with by-pass linseed oil (LO; rich in α-linolenic acid) on maternal antioxidant systems at Days 14 and 16 of pregnancy in Sarda ewes. This trial used sixteen dry ewes. Eight ewes (CT group) were fed with a control diet without LO, and eight ewes (LO group) were fed with a diet supplemented with LO (10.8 g of α-linolenic acid/ewe/day). Both diets had similar crude protein and energy levels. The experiment included 10 days of an adaptation period and 31 days of a supplementation period. This supplementation period was divided into Period −2 (from Day −15 to −8), Period −1 (from Day −7 to −1; before synchronized mating period/Day 0), Period +1 (from Day +1 to + 7 after mating), and Period +2 (from Day +8 to +15 after mating). Estrous synchronization was induced in all the ewes using an intravaginal sponge (45 mg fluorgestone acetate) for 14 days and equine chorionic gonadotropin (350 UI/ewe) at the end of the treatment. On Days 14 (CT, N = 4; LO, N = 4) and 16 (CT, N = 4; LO, N = 4) after mating, the ewes were slaughtered. Samples of plasma, uterine, and luteal tissues were collected. Thiols, total antioxidant activity (TEAC), superoxide dismutase (SOD) activity, and malondialdehyde (MDA) content were measured. On Day 16, thiol and TEAC in luteal tissues were higher in the LO group when compared with the control one (p < 0.05). Moreover, TEAC was higher for the LO group in uterine tissues on Days 14 and 16 (p < 0.05). SOD activity was higher in the LO group in luteal and uterine tissues on Day 14 and Day 16, respectively (p < 0.001). On Day 16, uterine MDA content was lower for the LO group (p < 0.001). No differences were found between groups at the plasmatic level. However, the by-pass LO supplementation enhanced the analyzed antioxidant parameters in luteal and uterine tissues. In conclusion, these results demonstrate that by-pass LO supplementation exerted a positive effect on antioxidative defenses on maternal structures during the embryo-maternal recognition period in ewes. Thus, this could contribute to improving the maternal environment during the embryo-maternal recognition period in mammals.

Arsenic, an environmental and industrial pollutant causes female reproductive disturbances and female infertility. Several researchers found that the use of Camellia sinensis (CS) (green tea) is effective as an alternative therapeutic strategy in the management of several health ailments. This study explores the role of CS extract against arsenic-induced rat uterine tissue damage. Methanolic extract of CS (10 mg/kg BW) was tested concomitantly in arsenic-treated (10 mg/kg BW) rats for a duration of two-oestrous cycle length (8 days). CS effectively attenuated arsenic-induced antioxidantdepletion and necrosis in uterine tissue. Rats treated with sodium arsenite showed significantlyreduced activities of enzymatic antioxidants like superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) in uterine tissue as evidenced by the results of spectrophotometric and electrozymographic analysis. Co-administration of CS significantly reversed the above oxidative stress markers in uterine tissue along with the histopathological changes in ovarian and uterine tissue. Moreover, an increase in the level of transcription factor NF-κB in the uterine tissue in association with reduced serum levels of vitamin B12 and folic acid were mitigated in arsenic fed rats following CS co-administration.

Dietary supplementation with selenomethionine enhances antioxidant capacity and selenoprotein gene expression in layer breeder roosters