F1 Generation Rats Research Articles

Prenatal exposure to nitrate alters uterine morphology and gene expression in adult female F1 generation rats.

Nitrate is ubiquitously found in the environment and is one of the main components of nitrogen fertilizers. Previous studies have shown that nitrate disrupts the reproductive system in aquatic animals, but no study has evaluated the impact of nitrate exposure on the uterus in mammals. This study aimed to evaluate the impact of maternal exposure to nitrate during the prenatal period on uterine morphology and gene expression in adult female F1 rats. Pregnant Wistar rats were either treated with sodium nitrate 20 mg/L or 50 mg/L dissolved in drinking water from the first day of pregnancy until the birth of the offspring or were left untreated. On postnatal day 90, the uteri of female offspring rats were collected for histological and gene expression analyses. Morphometric analyses of the uterine photomicrographs were performed to determine the thickness of the layers of the uterine wall (endometrium, myometrium, and perimetrium) and the number of endometrial glands. The highest nitrate dose increased the myometrial thickness of the exposed female rats. Treatment with both nitrate doses reduced the number of endometrial glands compared with no treatment. Additionally, nitrate treatment significantly increased the expression of estrogen receptors and reduced the expression of progesterone receptors in the uterus. Our results strongly suggest that prenatal exposure to nitrate programs gene expression and alters the uterine morphology in female F1 rats, potentially increasing their susceptibility to developing uterine diseases during adulthood.

Extended one generation reproductive toxicity study and effect on gut flora of genetically modified rice rich in β-carotene in wistar rats

To evaluate the reproductive toxicity of gene modified rice generated by introducing phytoene synthase (Psy) and bacterial phytoene desaturase (CrtI) from maize and Erwinia uredovora, Wistar rats were allocated into 3 groups and fed with Psy and CrtI gene modified rice mixture diet (GM group), non-gene modified rice mixture diet (non-GM group), and AIN-93 diet (Blank control group) from parental generation (F0) to the offsprings (F1). GM rice, Heijinmi (HJM) and Non-GM rice, Heishuai (HS), were both formulated into diets at ratios of 73.5% and 75.5% according to the AIN93 diet for rodent animals, respectively. Relative to the non-GM group, no biologically relevant differences were observed in GM group rats concerning reproductive performance such as fertility rate, gestation rate, mean duration, hormone level, and reproductive organ pathology. The developmental parameters results were not significantly different from the non-GM group such as body weight, food consumption, developmental neurotoxicity, behavior, hematology, and serum chemistry. In terms of immunotoxicity, the IgG indicators of offspring from the GM group improved in contrast with the non-GM group. Additional gut flora analysis of F0 generation rats resulted as that the treatment elicited an increased gut microflora diversity of F0 rats. And no horizontal gene transfer of Psy and CrtI genes in rats fed a GM rice HJM diet. In conclusion, we found no adverse effects related to GM rice in the extended one-generation reproductive toxicity study, indicating that GM rice is a safe alternative for its counterpart rice regarding reproductive toxicity.

Long term excessive zinc supplementation in diet induced alteration in serum lipids, hormones and minerals profile in wistar rats and has carry over effect in their F1 generation rats

The use of the zinc in various areas such as animal husbandry, agricultural techniques and multivitamin supplements has increased in recent decades and will increase over time. In this study, the impact of Zn overload for long period in the body, and their carry-over effect on the next generation is determined. Three groups of rats were fed on basal diets having 20 mg Zn in one kg diet (group-1), control group, 50 mg Zn in one kg diet (Group-2) and 80 mg Zn in one kg diet (Group-3) for 180 days. The F1- generation rats obtained from each group of parent rats were fed only on rat pellet diet. The investigation revealed that body weight rise with increased concentration of Zn in parent and F1-generation rats. The blood lipid profile in serum showed an increase in triglycerides, cholesterol, very-low-density lipoprotein (VLDL) cholesterol whereas lowered level of high-density Lipoprotein (HDL) cholesterol levels was recorded in both generation of rats. A significant increase in serum Insulin, HbA1c, and C-reactive protein level was recorded in group-2 and 3 rats while no significant changes in the FI- generation of rats. In F1- rats, Zn showed increase in its level while decreased level of Mn and Cu in the kidney and liver of group-F1-2 and F1-3 was observed in comparison to their control counterparts. The results of the study suggest long term treatment of diet rich in Zn induced alteration in serum lipids, hormones and profile of minerals and has carry over effect to their F1- generations.

Effect of coumestrol on the biochemical constituents of ovary and uterus of pups of Female albino rats

In the current research, attempts were undertaken to evaluate the impact of coumestrol on some biochemical constituents of rats of F1 generation when these animals reached the ages of 2,4, and 6 months. The administration of coumestrol at different doses of 5,10 and 25 ug/kg during different days of pregnancy (Group-1 on the 10th and 13th day) and (Group-2 on the 15th and 18th day) caused a substantial rise in different parameters like the wet weight of uterus and ovary, protein contents, glycogen contents, the acid phosphatase, and alkaline phosphatase activity even at the age of 6 months.

Transcriptomic profiling identified altered expression of genes associated with testicular functions in adult F1 rats exposed to carbimazole during fetal period

The central goal of this study was to investigate the alterations in transcriptome of testis in F1 generation adult rats exposed to carbimazole prenatally. At post-natal day 100, the testis of rats delivered to carbimazole exposed (time-mated pregnant rats orally administered with carbimazole from gestation day 9 to 21) and control (untreated pregnant rats) groups were subjected to transcriptomic analysis using NGS platform. A total of 187 differentially expressed (up regulated: 49 genes; down regulated: 138) genes were identified in carbimazole exposed rats over controls and the major processes associated with these altered testicular transcripts were examined. Functional clustering analysis suggest that the involvement of identified DEGs were linked to intrinsic and extrinsic apoptotic pathways, mitochondrial solute carriers slc25a members, nuclear receptors/zinc family members, steroidogenic pathway and cholesterol synthesis, and growth factors and protein kinases and thus represent potential mediators of the developmental toxic effects of carbimazole in F1 generation rats. Based on the findings, it can be concluded that prenatal exposure to carbimazole prominently affects expression of multiple transcripts implicating key regulatory events associated with testicular functions, spermatogenesis and steroidogenesis in rats at their adulthood. These results support our earlier findings and hypothesis. This background information obtained at the testicular transcriptome during gestational hypothyroidism might be helpful for future studies and experiments to gain additional in-depth analysis and to develop strategies to protect F1 generation male reproductive health. SignificanceThe rationale for the paper described thyroid gland changes in the off springs. Antithyroid drugs are widely used to control thyroid disorders and used to control thyroid hormone levels during surgeries. Carbimazole is one of the antithyroid drugs and is a parent molecule of methimazole. Both the drugs can able to cross placenta. During fetal period, the development of thyroid gland is not completely formed and hence, the fetus entirely depends on the maternal thyroid hormones. Therefore, it is conceivable that the disturbances at the level of maternal thyroid hormones could interfere with the development of vital organs such as testis and glands including thyroid gland (Kala et al., 2012). To address this notion, the present study was designed and executed.

Maternal exposure to di(2-ethylhexyl) phthalate (DEHP) causes multigenerational adverse effects on the uterus of F1 and F2 offspring rats.

Phthalates are one of the ubiquitous chemicals found in day-to-day products like food packaging, children's toys, and other consumer commodities. There is rising concern that repeated exposure to phthalates during pregnancy and lactation could have long-term effects on maternal and fetal health. We hypothesize that exposure to DEHP during the developmental windows might affect the expression of molecules that regulate uterine function and that this effect would be passed on to further generations. Rat dams were treated with olive oil (vehicle) or DEHP (100 mg/kg b.wt./day) orally from gestational day 9 (GD 9) to the end of lactation (PND 21). F0 maternal DEHP exposure resulted in multigenerational (F1 and F2) reproductive toxicity, as evidenced by an extended estrous cycle, decreased mating, fertility, and fecundity indices. Serum progesterone and estradiol levels were decreased and their cognate receptors (PR and ERα) in the uterus were decreased in the DEHP-exposed offspring rats. Further analysis of the expression of estrogen and progesterone regulatory genes such as Hox a11, VEGF A, Ihh, LIFR, EP4, PTCH, NR2F2, BMP2, and Wnt4 were reduced in the uteri of adult F1 and F2 generation rats born from DEHP-exposed F0 dams. Decreased expression of these crucial proteins due to DEHP exposure may lead to defects in epithelial proliferation and secretion, uterine receptivity, and decidualization in the uteri of successive generations. This study showed that maternal DEHP exposure impairs the expression of molecules that regulate uterine function and this multigenerational effect is transmitted via maternal lineage.

Transforming growth factor-β receptor 1: An intervention target for genetic poor cartilage quality induced by prenatal dexamethasone exposure

IntroductionFetal-originated osteoarthritis is relative to poor cartilage quality and may exhibit transgenerational genetic effects. Previous findings revealed prenatal dexamethasone exposure (PDE) induced poor cartilage quality in offspring. ObjectivesThis study focused on further exploring molecular mechanism, heritability, and early intervention of fetal-originated osteoarthritis. MethodsPregnant rats (F0) were segregated into control and PDE groups depending upon whether dexamethasone was administered on gestational days (GDs) 9–20. Some female offspring were bred with healthy males during postnatal week (PW) 8 to attain the F2 and F3 generations. The F3-generation rats were administrated with glucosamine intragastrically at PW12 for 6 weeks. The knee cartilages of male and female rats at different time points were harvested to assay their morphologies and functions. Furthermore, primary chondrocytes from the F3-generation rats were isolated to confirm the mechanism and intervention target of glucosamine. ResultsCompared with the control, female and male rats in each generation of PDE group showed thinner cartilage thicknesses; shallower and uneven staining; fewer chondrocytes; higher Osteoarthritis Research Society International scores; and lower mRNA and protein expression of SP1, TGFβR1, Smad2, SOX9, ACAN and COL2A1. After F3-generation rats were treated with glucosamine, all of the above changes could be reversed. In primary chondrocytes isolated from the F3-generation rats of PDE group, glucosamine promoted SP1 expression and binding to TGFβR1 promoter to increase the expression of TGFβR1, p-Smad2, SOX9, ACAN and COL2A1, but these were prevented by SB431542 (a potent and selective inhibitor of TGFβR1). ConclusionsPDE induced chondrodysplasia in offspring and stably inherited in F3-generation rats, which was related to decreased expression of SP1/TGFβR1/Smad2/SOX9 pathway to reduce the cartilage matrix synthesis, without major sex-based variations. Glucosamine could alleviate the poor genetic cartilage quality in offspring induced by PDE by up-regulating SP1/TGFβR1 signaling, which was prevented by a TGFβR1 inhibitor. This study elucidated the molecular mechanism and therapeutic target (TGFβR1) of genetic chondrodysplasia caused by PDE, which provides a research basis for precisely treating fetal-originated osteoarthritis.

Prenatal exposure to di(n-butyl) phthalate delays the spermatogenic cycle in rats: Investigation using a BrdU-injection method

Di(n-butyl) phthalate (DBP) esters are plasticizers that are used to provide transparency and flexibility in household plastic products but can easily leach out to contaminate organisms and the environment. We investigated whether prenatal DBP exposure affects spermatogenesis in rats. Pregnant Sprague–Dawley rats were injected with DBP 10, 50, and 100 mg/kg, or vehicle, administered intragastrically, on gestation days 12–21. At 9 or 17 weeks, 5-bromodeoxyuridine (BrdU) 50 mg/kg was injected intraperitoneally, and one testis was removed 3 h later. The remaining testis was excised 12.95 days + 3 h after the BrdU injection. Immunohistochemical analysis of BrdU was performed with periodic acid-Schiff and hematoxylin counterstaining for a quantitative analysis of the delay in one cycle of spermatogenesis. The DBP 100 mg group showed that the ratio of the appearance of seminiferous tubules in stages VII and VIII were significantly decreased, but those of stages IX and X were significantly increased compared to the Vehicle group. The reference value for the duration of spermatogenesis per cycle was set at 310.8 h. The DBP 100 mg group showed a significant delay in the duration of one cycle of spermatogenesis (16.95 h at puberty and 19.01 h at adulthood) compared with the Vehicle group. This study determined that F1-generation rats with prenatal DBP 100 mg exposure revealed significant accumulation of spermatogenic cells at stages IX to X in the second and third cycles, and the significant delay in the duration of spermatogenesis was more prominent at adulthood than in puberty.

Transgenerational evidence of increases in dopamine D2 receptor sensitivity in rodents: Impact on sensorimotor gating, the behavioral response to nicotine and BDNF.

Neonatal quinpirole (NQ) treatment to rats increases dopamine D2 (DAD2) receptor sensitivity in adult animals. We investigated if increased DAD2 sensitivity would be passed to the next (F1) generation, and if these animals demonstrated sensorimotor gating deficits and enhanced behavioral responses to nicotine. Male and female rats were intraperitoneal (IP) administered quinpirole (1 mg/kg) or saline (NS) from postnatal day (P)1-21. Animals were either behaviorally tested (F0) or raised to P60 and mated, creating F1 offspring. Experiment 1 revealed that F1 generation animals that were the offspring of at least one NQ-treated founder increased yawning behavior, a DAD2-mediated behavioral event, in response to acute quinpirole (0.1 mg/kg). F1 generation rats also demonstrated increased striatal β arrestin-2 and decreased phospho-AKT signaling, consistent with increased G-protein independent DAD2 signaling, which was equal to F0 NQ-treated founders, although this was not observed in all groups. RNA-Seq analysis revealed significant gene expression changes in the F1 generation that were offspring of both NQ-treated founders compared to F0 NQ founders and controls, with enrichment in sensitivity to stress hormones and cell signaling pathways. In Experiment 2, all F1 generation offspring demonstrated sensorimotor gating deficits compared to controls, which were equivalent to F0 NQ-treated founders. In Experiment 3, all F1 generation animals demonstrated enhanced nicotine behavioral sensitization and nucleus accumbens (NAcc) brain-derived neurotrophic factor (BDNF) protein. Further, F1 generation rats demonstrated enhanced adolescent nicotine conditioned place preference equivalent to NQ-treated founders conditioned with nicotine. This represents the first demonstration of transgenerational effects of increased DAD2 sensitivity in a rodent model.

Epigenome-wide association study for atrazine induced transgenerational DNA methylation and histone retention sperm epigenetic biomarkers for disease

Atrazine is a common agricultural herbicide previously shown to promote epigenetic transgenerational inheritance of disease to subsequent generations. The current study was designed as an epigenome-wide association study (EWAS) to identify transgenerational sperm disease associated differential DNA methylation regions (DMRs) and differential histone retention regions (DHRs). Gestating female F0 generation rats were transiently exposed to atrazine during the period of embryonic gonadal sex determination, and then subsequent F1, F2, and F3 generations obtained in the absence of any continued exposure. The transgenerational F3 generation males were assessed for disease and sperm collected for epigenetic analysis. Pathology was observed in pubertal onset and for testis disease, prostate disease, kidney disease, lean pathology, and multiple disease. For these pathologies, sufficient numbers of individual males with only a single specific disease were identified. The sperm DNA and chromatin were isolated from adult one-year animals with the specific diseases and analyzed for DMRs with methylated DNA immunoprecipitation (MeDIP) sequencing and DHRs with histone chromatin immunoprecipitation (ChIP) sequencing. Transgenerational F3 generation males with or without disease were compared to identify the disease specific epimutation biomarkers. All pathologies were found to have disease specific DMRs and DHRs which were found to predominantly be distinct for each disease. No common DMRs or DHRs were found among all the pathologies. Epimutation gene associations were identified and found to correlate to previously known disease linked genes. This is one of the first observations of potential sperm disease biomarkers for histone retention sites. Although further studies with expanded animal numbers are required, the current study provides evidence the EWAS analysis is effective for the identification of potential pathology epimutation biomarkers for disease susceptibility.

Developmental origins of transgenerational sperm histone retention following ancestral exposures

Numerous environmental toxicants have been shown to induce the epigenetic transgenerational inheritance of disease and phenotypic variation. Alterations in the germline epigenome are necessary to transmit transgenerational phenotypes. In previous studies, the pesticide DDT (dichlorodiphenyltrichloroethane) and the agricultural fungicide vinclozolin were shown to promote the transgenerational inheritance of sperm differential DNA methylation regions, non-coding RNAs and histone retention, which are termed epimutations. These epimutations are able to mediate this epigenetic inheritance of disease and phenotypic variation. The current study was designed to investigate the developmental origins of the transgenerational differential histone retention sites (called DHRs) during gametogenesis of the sperm. Vinclozolin and DDT were independently used to promote the epigenetic transgenerational inheritance of these DHRs. Male control lineage, DDT lineage and vinclozolin lineage F3 generation rats were used to isolate round spermatids, caput epididymal spermatozoa, and caudal sperm. The DHRs distinguishing the control versus DDT lineage or vinclozolin lineage samples were determined at these three developmental stages. DHRs and a reproducible core of histone H3 retention sites were observed using an H3 chromatin immunoprecipitation-sequencing (ChIP-Seq) analysis in each of the germ cell populations. The chromosomal locations and genomic features of the DHRs were analyzed. A cascade of epigenetic histone retention site alterations was found to be initiated in the round spermatids and then further modified during epididymal sperm maturation. Observations show that in addition to alterations in sperm DNA methylation and ncRNA expression previously identified, the induction of differential histone retention sites (DHRs) in the later stages of spermatogenesis also occurs. This novel component of epigenetic programming during spermatogenesis can be environmentally altered and transmitted to subsequent generations through epigenetic transgenerational inheritance.

Transgenerational effects following chronic circadian disturbance given to mother rats in F1 generation rats

It has not been documented whether circadian disturbance of mother may influence fetal neurodevelopment. Therefore, this study aims to find the root cause of congenital neuronal defects, which is explosively increasing today, in one of the environmental factors; circadian disturbance. Female rats were randomly divided into 3 groups; L/D group with 12/12 hr light and dark cycle, LL group with 24 hr constant light, DD group with 24 hr constant dark. All subjected females were assigned to each light condition one week prior to pregnancy. Then, they were mated with males, and F1 offsprings were obtained at three weeks after the mating. The future generations were exposed exactly same light conditions. The animals in each group were randomly divided into control and melatonin‐treated subgroups. All animals were engaged in physiological, histological, and neurobehavioral analyses. LL and DD groups showed several abnormal features and decreased cecum/colon length (p<.05). Also, LL and DD groups showed lower activation of intracranial δ‐wave than L/D group showed. However, in LL and DD groups treated with melatonin improved intestinal integrity and activated intracranial δ‐wave. Axonal sprouting was evaluated by Golgi‐cox staining for identifying the abnormal neurodevelopment in all groups. The LL and DD groups showed reduction of axonal sprouting than L/D group. However, axonal branching in neuron increased in LL and DD groups treated with melatonin. In addition, neurobehavioral tests including open field test, novel objective recognition test, social interaction test, spontaneous alternation Y‐maze test, and elevated plus maze test were performed. LL and DD groups showed abnormal neurobehaviors (p<.05), whereas, melatonin treatment resulted in the normalized neurobehaviors (p<.05). In conclusion, F1 generation born from mothers exposed to chronic circadian disturbance showed abnormal neurodevelopment and neurobehaviors. Melatonin was considered as a therapeutic agent for congenital neuronal deficit.Support or Funding InformationFundings: National Research Foundation (NRF‐2017R1A2A2A01067169, NRF‐2019R1A6A3A01091422), Korea

Epigenome-wide association study (EWAS) for potential transgenerational disease epigenetic biomarkers in sperm following ancestral exposure to the pesticide methoxychlor.

Environmental exposures such as chemical toxicants can alter gene expression and disease susceptibility through epigenetic processes. Epigenetic changes can be passed to future generations through germ cells through epigenetic transgenerational inheritance of increased disease susceptibility. The current study used an epigenome-wide association study (EWAS) to investigate whether specific transgenerational epigenetic signatures of differential DNA methylation regions (DMRs) exist that are associated with particular disease states in the F3 generation great-grand offspring of F0 generation rats exposed during gestation to the agricultural pesticide methoxychlor. The transgenerational epigenetic profiles of sperm from F3 generation methoxychlor lineage rats that have only one disease state were compared to those that have no disease. Observations identify disease specific patterns of DMRs for these transgenerational rats that can potentially serve as epigenetic biomarkers for prostate disease, kidney disease, obesity, and the presence of multiple diseases. The chromosomal locations, genomic features, and gene associations of the DMRs are characterized. Disease specific DMR sets contained DMR-associated genes that have previously been shown to be associated with that specific disease. Future epigenetic biomarkers could potentially be developed and validated for humans as a disease susceptibility diagnostic tool to facilitate preventative medicine and management of disease.

Transgenerational effect of parental obesity and chronic parental bisphenol A exposure on hormonal profile and reproductive organs of preadolescent Wistar rats of F1 generation: A one-generation study

There is a global concern about adverse health effects of endocrine-disrupting chemicals (EDCs). Bisphenol A (BPA), an estrogenic and obesogenic compound, used in the plastic and medical industry has a dominant position among EDCs as far as human health and regulatory scenario are concerned. Due to its omnipresence across the biosphere, population of all age groups and health status is unavoidably exposed to BPA. Transgenerational exposure to BPA and its effects have also been recognized. However, there is no report on the transgenerational effect of BPA on metabolically disordered parents, such as obese ones. We studied effect of BPA exposure in F0 generation and its impact on F1 generation and factored parental obesity in transgenerational effect of concurrent exposure to low dose BPA (10 ppm × 180 days) in Wistar rats in a one-generation study protocol. The exposed F0 generation animals were crossed and F1 generation was analyzed 35 days after birth for indications of reproductive toxicity. We observed changes in hormone levels and disturbance in glucose and lipid homeostasis. Animals showed increased serum cholesterol and triglycerides along with higher birth weight and rapid weight gain. Histopathological evidence confirmed the presence of regressive and inflammatory changes in the ovary and testis. The test group showed metabolic disturbances in comparison to control group. Our study showed the additive effect of parental obesity in transgenerational reproductive toxicity of BPA. Female animals of F1 generation of BPA-treated obese parents showed more insulin resistance than males with similar exposure scenario. Our study highlights the confounding role of metabolic disorders such as obesity in the transgenerational toxicity of BPA, which otherwise itself is implicated in the aetiology of such metabolic disorders, directly or indirectly.

Epigenetic transgenerational inheritance of testis pathology and Sertoli cell epimutations: generational origins of male infertility.

Male reproductive health has been in decline for decades with dropping sperm counts and increasing infertility, which has created a significant societal and economic burden. Between the 1970s and now, a general decline of over 50% in sperm concentration has been observed in the population. Environmental toxicant-induced epigenetic transgenerational inheritance has been shown to affect testis pathology and sperm count. Sertoli cells have an essential role in spermatogenesis by providing physical and nutritional support for developing germ cells. The current study was designed to further investigate the transgenerational epigenetic changes in the rat Sertoli cell epigenome and transcriptome that are associated with the onset of testis disease. Gestating female F0 generation rats were transiently exposed during the period of fetal gonadal sex determination to the environmental toxicants, such as dichlorodiphenyltrichloroethane (DDT) or vinclozolin. The F1 generation offspring were bred (i.e. intercross within the lineage) to produce the F2 generation grand-offspring that were then bred to produce the transgenerational F3 generation (i.e. great-grand-offspring) with no sibling or cousin breeding used. The focus of the current study was to investigate the transgenerational testis disease etiology, so F3 generation rats were utilized. The DNA and RNA were obtained from purified Sertoli cells isolated from postnatal 20-day-old male testis of F3 generation rats. Transgenerational alterations in DNA methylation, noncoding RNA, and gene expression were observed in the Sertoli cells from vinclozolin and DDT lineages when compared to the control (vehicle exposed) lineage. Genes associated with abnormal Sertoli cell function and testis pathology were identified, and the transgenerational impacts of vinclozolin and DDT were determined. Alterations in critical gene pathways, such as the pyruvate metabolism pathway, were identified. Observations suggest that ancestral exposures to environmental toxicants promote the epigenetic transgenerational inheritance of Sertoli cell epigenetic and transcriptome alterations that associate with testis abnormalities. These epigenetic alterations appear to be critical factors in the developmental and generational origins of testis pathologies and male infertility.

Beta cypermethrin exposure and perinatal reproductive development of female f1 generation of albino rats

BackgroundPrevious studies have shown that cypermethrin has a teratogenic effect on rat feti born to exposed dam or buck with no information on its effect on their reproductive parameters at maturity. The present study was aimed at evaluating the reproductive effect of perinatal beta cypermethrin (β-cyp) exposure on female F1 generation rat.MethodsFifteen pregnant animals (day 0 = day of mating, average body-weight = 190 g) were randomly divided into 3 groups. Group A (control) received 0.5 ml olive oil, group B (15 mg/kg β-cyp), and group C (30 mg/kg β-cyp) by oral gavage from gestational day (GD) 1—post natal day (PND) 20. On PND 21, the pups were weaned and bred to 12 weeks of age (maturity). At maturity, 5 females were randomly taken from each group. The estrous cycle was determined for 14 days. Thereafter, the animals were anesthetized and blood was collected for hormonal assay (LH, FSH, estrogen, progesterone) using enzyme immunoassay.Resultsβ-cyp had no significant effect (p > 0.05) on the estrous cycle. There was no significant (p > 0.05) change in the serum concentration of the sex hormones except the LH concentration where there was a dose-dependent significant (p < 0.05) decrease in treated groups relative to the control.ConclusionIt is therefore concluded that perinatal beta cypermethrin exposure could have a deleterious effect on the reproductive development of female F1 generation rats, even at maturity.

Transgenerational sperm DNA methylation epimutation developmental origins following ancestral vinclozolin exposure

ABSTRACTA number of environmental factors from nutrition to toxicants have been shown to promote the epigenetic transgenerational inheritance of disease and phenotypic variation. This requires alterations in the germline (sperm or egg) epigenome. Previously, the agricultural fungicide vinclozolin was found to promote the transgenerational inheritance of sperm differential DNA methylation regions (DMRs) termed epimutations that help mediate this epigenetic inheritance. The current study was designed to investigate the developmental origins of the transgenerational DMRs during gametogenesis. Male control and vinclozolin lineage F3 generation rats were used as a source of embryonic day 13 (E13) primordial germ cells, embryonic day 16 (E16) prospermatogonia, postnatal day 10 (P10) spermatogonia, adult pachytene spermatocytes, round spermatids, caput epididymal spermatozoa, and caudal sperm. The DMRs between the control versus vinclozolin lineage samples were determined for each developmental stage. The top 100 statistically significant DMRs for each stage were compared. The developmental origins of the caudal epididymal sperm DMRs were assessed. The chromosomal locations and genomic features of the different stage DMRs were investigated. In addition, the DMR associated genes were identified. Previous studies have demonstrated alterations in the DMRs of primordial germ cells (PGCs). Interestingly, the majority of the DMRs identified in the current study for the caudal sperm originated during the spermatogenic process in the testis. A cascade of epigenetic alterations initiated in the PGCs appears to be required to alter the epigenetic programming during spermatogenesis to modify the sperm epigenome involved in the transgenerational epigenetic inheritance phenomenon.

Extenuation of in utero toxic effects of MeHg in the developing neurons by Fisetin via modulating the expression of synaptic transmission and plasticity regulators in hippocampus of the rat offspring

The neurotoxic environmental contaminant, methylmercury (MeHg), has shown to have detrimental effects on the developing brain when exposed during gestation. We have shown in our earlier studies that gestational administration of 3,3′,4′,7-Tetrahydroxyflavone or Fisetin reduces the toxic effects of MeHg in the developing rat brain. The current study has pivoted to study the mechanism behind the mitigating action of Fisetin against prenatal MeHg exposure induced neurotoxicity. Negligible data is available about the toxicity targets of MeHg in the developing brain. Studies have exhibited that MeHg exposure cause toxic effects on synaptic transmission and plasticity in the offspring brain. Hence, we aimed to study the effect of Fisetin on MeHg induced alterations in the expressions of regulatory genes and proteins involved in synaptic plasticity and transmission. Pregnant rats were grouped according to the type of oral administration as, (i) Control, (ii) MeHg (1.5 mg/kg b. w.), (iii) MeHg + Fisetin (30 mg/kg b. w.) and (iv) Fisetin (30 mg/kg b. w). Maternal administration of Fisetin prevented MeHg exposure induced downregulation of neurogranin (Nrgn), dendrin (Ddn), Syntaxin 1 A (Stx1a), Lin-7 homolog A (Lin7a), Complexin-2 (Cplx2) and Exocyst complex component 8 (Exoc8) genes in the offspring rat. Fisetin also prevented MeHg exposure induced downregulation of brain derived neurotrophic factor (BDNF), Glial-cell derived neurotrophic factor (GDNF) protein expressions and hampered reactive astrogliosis in the hippocampus of F1 generation rats. Hence, through this study, we conclude that Fisetin modulates the expression of regulatory genes and proteins involved in synaptic transmission and plasticity and extenuates MeHg neurotoxicity in the developing rat brain.

Developmental origins of transgenerational sperm DNA methylation epimutations following ancestral DDT exposure

Epigenetic alterations in the germline can be triggered by a number of different environmental factors from diet to toxicants. These environmentally induced germline changes can promote the epigenetic transgenerational inheritance of disease and phenotypic variation. In previous studies, the pesticide DDT was shown to promote the transgenerational inheritance of sperm differential DNA methylation regions (DMRs), also called epimutations, which can in part mediate this epigenetic inheritance. In the current study, the developmental origins of the transgenerational DMRs during gametogenesis have been investigated. Male control and DDT lineage F3 generation rats were used to isolate embryonic day 16 (E16) prospermatogonia, postnatal day 10 (P10) spermatogonia, adult pachytene spermatocytes, round spermatids, caput epididymal spermatozoa, and caudal sperm. The DMRs between the control versus DDT lineage samples were determined at each developmental stage. The top 100 statistically significant DMRs at each stage were compared and the developmental origins of the caudal epididymal sperm DMRs were assessed. The chromosomal locations and genomic features of the different stage DMRs were analyzed. Although previous studies have demonstrated alterations in the DMRs of primordial germ cells (PGCs), the majority of the DMRs identified in the caudal sperm originated during the spermatogonia stages in the testis. Interestingly, a cascade of epigenetic alterations initiated in the PGCs is required to alter the epigenetic programming during spermatogenesis to obtain the sperm epigenetics involved in the epigenetic transgenerational inheritance phenomenon.

Vinclozolin induced epigenetic transgenerational inheritance of pathologies and sperm epimutation biomarkers for specific diseases.

Exposure to vinclozolin has been shown to induce the epigenetic transgenerational inheritance of increased susceptibility to disease, and to induce transgenerational changes to the epigenome. In the current study, gestating F0 generation rats were exposed to vinclozolin, and the subsequent F1, F2 and transgenerational F3 generations were evaluated for diseases and pathologies. F1 and F2 generation rats exhibited few abnormalities. However, F3 generation rats showed transgenerational increases in testis, prostate, and kidney disease, changes in the age of puberty onset in males, and an increased obesity rate in females. Overall there was an increase in the rate of animals with disease, and in the incidence of animals with multiple diseases. The objective of the current study was to analyze the sperm epigenome of F3 generation rats with specific abnormalities and compare them to rats without those abnormalities, in an effort to find epigenetic biomarkers of transgenerational disease. Unique signatures of differential DNA methylation regions (DMRs) in sperm were found that associated with testis disease, prostate disease and kidney disease. Confounding factors identified were the presence of multiple diseases in the analysis and the limited number of animals without disease. These results further our understanding of the mechanisms governing epigenetic transgenerational inheritance, and may lead in the future to the use of epigenetic biomarkers that will help predict an individual’s susceptibility for specific diseases.