Shift and longtime light induces endometrioid adenocarcinoma via activation of PKC-α/Akt pathway in female golden hamster: Involvement of altered Aanat and Bmal1 rhythm.

Female night-workers get exposed to frequent light shifts, hence have altered circadian rhythm and are at high risk of endometrial cancer; the underlying mechanism however is still not clear. We, therefore examined the effect of long light exposure (16L:8D, LD1) and regular shift (8 h) in long nighttime (LD2) on endometrial changes of female golden hamsters. Morphometric analysis, scanning electron microscopy imaging, alcian blue staining, and cytological nuclear atypia of endometrial stromal cells confirmed the incidence of endometrial adenocarcinoma in LD2 exposed hamsters. But, less severe pathomorphological alterations were noted in uterus of LD1 exposed hamsters. Altered Aanat and Bmal1 mRNA, melatonin rhythm, downregulation of important marker gene of adenocarcinoma like Akt, 14-3-3, and PR protein expression and upregulation PKCα, pAkt-S473 and vascular epithelial growth factor (VEGF) were observed in LD2 exposed hamsters suggesting the endometrial adenocarcinoma. Further, our western blot analysis supported the immunohistochemical localization of PR, PKCα, and VEGF in uterine tissues along low progesterone. Overall, our data indicates that light shift and long light exposure potentially induced endometrioid adenocarcinoma via activation of PKC-α/Akt pathway in female hamsters. Therefore, duration of light is essential for female normal uterine function.

BPS-induced ovarian dysfunction: Protective actions of melatonin via modulation of SIRT-1/Nrf2/NFĸB and IR/PI3K/pAkt/GLUT-4 expressions in adult golden hamster.

Ever-increasing occurrence of plastic-manufacturing industries leads to environmental pollution that has been associated with declined human health and increased incidence of compromised reproductive health. Female subfertility/infertility is a complex phenomenon and environmental toxicants as well as lifestyle factors have a crucial role to play. Bisphenol S (BPS) was believed to be a "safer" replacement of bisphenol A (BPA) but recent data documented its neurotoxic, hepatotoxic, nephrotoxic, and reprotoxic attributes. Hence based on the scarcity of reports, we investigated molecular insights into BPS-induced ovarian dysfunction and protective actions of melatonin against it in adult golden hamsters, Mesocricetus auratus. Hamsters were administered with melatonin (3 mg/kg BW i.p. alternate days) and BPS (150 mg/kg BW orally every day) for 28 days. BPS treatment disrupted hypothalamo-pituitary-ovarian (HPO) axis as evident by reduced gonadotropins such as luteinizing hormone (LH) and follicle-stimulating hormone (FSH), ovarian steroids such as estradiol (E2) and progesterone (P4), thyroid hormones namely triiodothyronine (T3) and thyroxine (T4) and melatonin levels along with their respective receptors (ERα, TRα, and MT-1) thereby reducing ovarian folliculogenesis. BPS exposure also led to ovarian oxidative stress/inflammation by increasing reactive oxygen species and metabolic disturbances. However, melatonin supplementation to BPS restored ovarian folliculogenesis/steroidogenesis as indicated by increased number of growing follicles/corpora lutea and E2/P4 levels. Further, melatonin also stimulated key redox/survival markers such as silent information regulator of transcript-1 (SIRT-1), forkhead box O-1 (FOXO-1), nuclear factor E2-related factor-2 (Nrf2), and phosphoinositide 3-kinase/protein kinase B (PI3K/pAkt) expressions along with enhanced ovarian antioxidant capacity. Moreover, melatonin treatment reduced inflammatory load including ovarian nuclear factor kappa-B (NFĸB), cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS) expressions, serum tumor necrosis factor α (TNFα), C-reactive protein (CRP) and nitrite-nitrate levels as well as upregulated ovarian insulin receptor (IR), glucose uptake transporter-4 (GLUT-4), connexin-43, and proliferating cell nuclear antigen (PCNA) expressions in ovary thereby ameliorating inflammatory and metabolic alterations due to BPS. In conclusion, we found severe deleterious impact of BPS on ovary while melatonin treatment protected ovarian physiology from these detrimental changes suggesting it to be a potential preemptive candidate against environmental toxicant-compromised female reproductive health.

Seasonal plasticity in immunocompetent cytokines (IL-2, IL-6, and TNF-α), myeloid progenitor cell (CFU-GM) proliferation, and LPS-induced oxido-inflammatory aberrations in a tropical rodent Funambulus pennanti: role of melatonin.

In seasonal breeders, photoperiods regulate the levels of circulatory melatonin, a well-known immunomodulator and an antioxidant. Melatonin is known to play a complex physiological role in maintaining the immune homeostasis by affecting cytokine production in immunocompetent cells. In this study, we have quantified seasonal and temporal variations in immunocompetent cytokines-IL-2, IL-6, and TNF-α-and circulatory corticosterone along with in- vitro proliferation of bone marrow-derived granulocyte macrophage-colony forming unit (CFU-GM) progenitor cells of a tropical seasonal breeder Funambulus pennanti (northern palm squirrel). Transient variations in antioxidant status of seasonal breeders might be due to the fluctuations associated with immunity and inflammation. Further, to establish a direct immunomodulatory effect of photoperiod, we recorded the LPS-induced oxidative and inflammatory responses of squirrels by housing them in artificial photoperiodic chambers mimicking summer and winter seasons respectively. We observed a marked variation in cytokines level, melatonin, and corticosterone , and CFU-GM cell proliferation during summer and winter seasons. High Peripheral melatonin levels directly correlated with cytokine IL-2 levels, and inversely correlated with TNF-α, and circulatory corticosterone level. LPS-challenged squirrels housed in short photoperiod (10L:14D; equivalent to winter days) showed a marked reduction in the components of the inflammatory cascade, CRP, TNF-α, IL-6, NOx, NF-κB, Cox-2, and PGES, with an overall improvement in antioxidant status when compared to squirrels maintained under a long photoperiod (16L:8D; equivalent to summer days). Our results underline the impact of seasonality, photoperiod, and melatonin in maintaining an intrinsic redox-immune homeostasis which helps the animal to withstand environmental stresses.

Impact of photoperiod on uterine redox/inflammatory and metabolic status of golden hamster, Mesocricetus auratus.

Photoperiod modulates reproductive physiology at multiple levels in seasonally breeding animals. Golden hamsters are long-day breeders that diminish their fertility during the short days. Photoperiod is known to regulate hormonal milieu and uterus is a hormone-sensitive dynamic tissue. However, there is lack of molecular insight regarding the impact of photoperiod on uterine physiology with respect to redox and metabolic status in Mesocricetus auratus. We evaluated the impact of photoperiod on circulatory hormonal parameters (triiodothyronine [T3], thyroxin [T4], estradiol [E2], progesterone [P4], melatonin, and insulin), their receptor expressions and key markers associated with redox (SIRT-1/FOXO-1), inflammatory (NFĸB/COX-2) and metabolic (IR/GLUT4) status in uterus. Adult female golden hamsters were exposed to different photoperiodic regimes, that is, short photoperiod (SP; 8L:16D) and long photoperiod (LP; 16L:8D) for 12 weeks. SP drastically decreased peripheral hormone profiles (T3, T4, E2, and P4) and compromised uterine histoarchitecture when compared with LP-exposed hamsters. Further, SP markedly decreased thyroid hormone receptor-α (TRα), insulin receptor, and glucose uptake transporter-4 (GLUT-4) expressions in uterus. We noted enhanced uterine oxidative (increased MDA and decreased SOD/CAT levels), SIRT-1/FOXO-1 expression and inflammatory (NFĸB/COX-2) load in SP condition. Further, elevated levels of circulatory insulin, melatonin, and its receptor (MT-1) expression in uterus was noted under SP condition. Thus, we may suggest that photoperiod might regulate uterine seasonality through modulation of local hormonal and redox/metabolic homeostasis thereby may restrict offspring bearing capacity under short days.

Melatonin attenuates LPS-induced ovarian toxicity via modulation of SIRT-1, PI3K/pAkt, pErk1/2 and NFĸB/COX-2 expressions.

The association between inflammation and metabolic disturbances leads to various female pathophysiological conditions. Bacterial lipopolysaccharide (LPS), found in the outer membrane of gram-negative bacteria, elicits an oxidative and inflammatory response that profoundly interferes with female reproductive health. We investigated the ameliorative action of melatonin on LPS-induced ovarian pathophysiology in golden hamsters, Mesocricetus auratus. Hamsters were administered with exogenous melatonin (5 mg/kg BW) and LPS (100 µg/kg BW) intraperitoneally for 7 days. LPS treatment impaired ovarian folliculogenesis as evident by histoarchitecture (elevated number of atretic follicles, reduced number of growing follicles and corpus luteum) and steroidogenesis (decreased aromatase/ERα, estradiol and progesterone). On the other hand, LPS administration also perturbed thyroid hormone (T3 and T4) homeostasis, ovarian melatonin receptor (MT-1) expression, antioxidant potential (SOD and catalase) and concomitantly elevated nitro-oxidative stress (decreased SOD, catalase and elevated CRP, TNFα and nitrate/nitrite level) and inflammatory load (NFĸB and COX-2) which culminated into ovarian follicular apoptosis (elevated caspase-3). LPS also disrupted metabolic homeostasis as indicated by hyperinsulinemia with a simultaneous decrease in ovarian IR/GLUT-4 and glucose content. Moreover, LPS treatment decreased expressions of key markers of ovarian physiology (SIRT-1, pErk1/2, PI3K and pAkt). Melatonin co-treatment with LPS improve these detrimental changes proposing melatonin as a potent therapeutic candidate against ovarian dysfunction induced by endotoxin.

Circadian desynchronization in pregnancy of Golden hamster following long time light exposure: Involvement of Akt/FoxO1 pathway.

Coordination between central and peripheral reproductive clocks in females is poorly understood. Long light is having a hazardous effect on reproductive health. Hence, explored the effect of long-time light exposure (LLD; 16L:8D) on the central and peripheral reproductive (ovary and uterus) clock genes (Bmal1, Clock, Per1, Per2, Cry1 and Cry2) and its downstream regulators (Aanat, Egf, Cx26, Cx43, ERα, pAktS-473, pAktT-308, pFoxO1T-24, 14-3-3, HoxA10, HoxA11 and Pibf) expression in non-pregnant and pregnant Golden hamster. Young adult Golden hamsters were exposed to LLD for 30 days and then were mated. We observed that LLD exposure increased the thickness of the endometrium and reduced myometrium thickness, resembling uterine adenomyosis. In non-pregnant females LLD altered the expressions of clock genes in suprachiasmatic nuclei (SCN), ovary and the uterus along with serum estradiol rhythm. LLD upregulated Egf and downregulated Aanat, Cx26, and Cx43 mRNA levels in uterus. LLD upregulated Akt/FoxO1 phosphorylation and 14-3-3 expressions in the uterus of nonpregnant females. LLD exposure to pregnant females lowered serum progesterone, Aanat, Pibf, Hoxa10, and Hoxa11 mRNA expressions on D4 (peri-implantation) and D8 (post-implantation) resulting in a low implantation rate on D8 (post-implantation). Hence it is evident that the frequent pregnancy anomalies noted under a long light schedule might be due to desynchronization in Aanat, Pibf, Hoxa10, and Hoxa11 as well as the central and peripheral clock genes (Bmal1, Clock, Per1, Per2, Cry1 and Cry2). LLD exposure desynchronized the central and peripheral reproductive clock affecting uterine physiology via Akt/FoxO1 pathway in Golden hamsters. Thus, LLD is a risk factor for female reproductive health and fertility.

Melatonin induces apoptosis and cell cycle arrest in cervical cancer cells via inhibition of NF-κB pathway.

Cervical cancer is the most prevalent cancer in females. Melatonin, a neurohormone has been documented as a promising therapeutic molecule for cervical cancer. However, the underlying molecular mechanism is not known. We explored the dose-dependent anti-tumor response of melatonin against cervical cancer cell lines, HeLa (HPV-18 positive) and SiHa (HPV-16 positive). The anti-cancer effect of melatonin was evaluated by MTT assay, cell imaging, colony formation, DAPI, AO/PI, LDH, Flow cytometry, scratch assay, western blot analysis and real-time PCR. Results of DAPI, AO/PI, LDH, and Annexin/PI staining revealed that melatonin induces apoptosis. The results of cell cycle analysis revealed that melatonin arrests the HeLa and SiHa cells in sub-G1 and G1 phases, respectively. Western blot analysis revealed that melatonin downregulated the expression of pro-inflammatory transcription factor, NF-κB and the expression of COX-2 protein, a key mediator in cell proliferation. In addition, melatonin downregulated the expression of an invasive marker, MMP-9, an antiapoptotic protein, Bcl-2, and upregulated the expression of pro-apoptotic protein, Bax at both transcriptional and translational levels. Overall, the results suggest that melatonin exhibited strong anti-cancer therapeutic potential against human cervical cancer cell line progression possibly through inhibition of NF-κB signalling pathway.

Continuous artificial light potentially disrupts central and peripheral reproductive clocks leading to altered uterine physiology and reduced pregnancy success in albino mice.

AIMS: The mechanism behind clock coordination in female reproductive disorders is poorly understood despite the known importance of coordinated and synchronized timing of central and clocks in reproductive organs. We investigated the effect of continuous artificial light (LL) on the central and peripheral reproductive clock gene (Bmal1, Clock, Per1, Per2 and Cry1) and its downstream regulators (Hgf, PR-A and HOXA10) during non-pregnancy and pregnancy phases of female mice. MAIN METHODS: Mice (n = 60) in two sets, were maintained under continuous light (LL) and natural day cycle (LD;12L: 12D) for both non-pregnant and pregnant study. Tissues from hypothalamus-containing SCN, ovary, uterus and serum were collected at different zeitgeber time points (ZT; at 4-h intervals across 24-h periods). KEY FINDINGS: LL exposure desynchronized the expressions of the clock mRNAs (Bmal1, Clock, Per1, Per2 and Cry1) in SCN, ovary, and uterus along with Hgf mRNA rhythm. LL significantly increased the thickness of endometrial tissues. Furthermore, the pregnant study revealed lower serum progesterone level during peri- and post-implantation under LL along with downregulated expression of progesterone receptor (PR) as well as progesterone dependent uterine Homeobox A-10 (Hoxa10) proteins with lowered pregnancy outcomes. SIGNIFICANCE: Our result suggests that LL disrupted the circadian coordination between central and clock genes in reproductive tissue leading to interrupted uterine physiology and altered pregnancy in mice. This led us to propose that duration of light exposure at work-places or home for females is very important in prevention of pregnancy anomalies.

Lactate-Dependent Cross-Talk Between Astrocyte and GnRH-I Neurons in Hypothalamus of Aged Brain: Decreased GnRH-I Transcription.

GnRH-I produced by hypothalamic neurosecretory cells is considered a master regulator of mammalian reproduction. Although GnRH-I transcription is well studied, the effect of ageing on transcriptional regulation of GnRH-I has not yet been explored. Here, we elucidate the effects of ageing on the metabolic environment like lactate level and TNF-α and how these affect GnRH-I transcription. Using pathway analysis of transcriptomic data, we found that lactate is upregulated in ageing astrocytes due to the downregulation of cellular respiration pathways possibly resulting in greater pyruvate concentration for lactate production. This lactate could then be shuttled into neurons where it would affect GnRH-I transcription. We showed that supra-physiological level of lactate in young mouse brain can mimic metabolic disturbances in the old brain and cause downregulation in GnRH-I transcription at a young age. In particular, we found upregulation of GnRH-I repressors in the young brain treated with high levels of lactate similar to old brain. Hence, this confirmed that aged metabolic environment can affect GnRH-I transcription even in the young brain. Further downstream analysis using the TRUST database showed NF-Kb signalling which lies downstream of both lactate and TNF-α as being capable of upregulating GnRH-I repressors. Since NF-Kb signalling has been shown in our study as well as others to be induced by TNF-α during ageing, it is likely that GnRH-I transcriptional regulation is mediated through these pathways. Thus, we formed a model for explaining the downregulation of GnRH-I transcription during ageing through differential expression of its TFs in an aged metabolic environment.

Photoperiodic modulation of ovarian metabolic, survival, proliferation and gap junction markers in adult golden hamster, Mesocricetus auratus.

Female reproductive physiology is greatly dependent on tight regulation of metabolic and survival factors. Photoperiod regulates female reproductive rhythms but very less information exists explaining whether photoperiod could modulate thyroid hormone homeostasis, metabolic/energy parameters along with survival, proliferation and gap junction proteins in the ovary of a long-day breeder, Mesocricetus auratus. Adult female hamsters were exposed to different photoperiodic regimes i.e., critical photoperiod (CP; 12.5L:11.5D), short photoperiod (SP; 8L:16D) and long photoperiod (LP; 16L:8D) for 12 weeks. LP upregulated thyroidal and gonadal activity as apparent by histoarchitecture, thyroid hormone profile [triiodothyronine (T3), thyroxin (T4) and thyroid stimulating hormone (TSH)], luteinizing hormone (LH), follicle stimulating hormone (FSH), estradiol (E2) and progesterone (P4) levels when compared with SP exposed hamsters. Further, LP increased thyroid hormone receptor-α/deiodinase-2 (TRα/Dio-2), estrogen receptor-α (ERα)/aromatase and insulin receptor/glucose transporter-4 (IR/GLUT-4) expressions in ovary. Interestingly, ovarian sirtuin-1 (SIRT-1) expression was also upregulated under LP condition along with cell proliferation (proliferating cell nuclear antigen or PCNA), survival (B cell lymphoma-2 or Bcl-2) and gap junction (connexin-43) markers when compared to SP exposed hamsters. We also noted elevated levels of circulatory leptin, insulin along with melatonin and its receptor (MT-1) in ovary under SP condition. Thus, we suggest that photoperiod plays a vital role in regulation of thyroid and reproductive hormone homeostasis along with key metabolic and survival markers in the ovary of adult golden hamsters, M. auratus providing further insight into the regulation of female reproductive seasonality in a long-day breeder.

Stress associated ovarian dysfunctions in a seasonal breeder Funambulus pennanti: Role of glucocorticoids and possible amelioration by melatonin.

Studies have shown that stress caused by lack of physical activity disrupts the normal pattern of glucocorticoid secretion which adversely affects the reproductive axis. We studied the effect of chronic movement restriction on ovarian responses in the Indian Palm Squirrel Funambulus pennanti, a highly active diurnal rodent. Physical restraint of squirrels induced stress that led to a significant increase in plasma cortisol, corticosterone and decreased 17ß-estradiol level leading to follicular atresia. Ovarian Reactive Oxygen Species (ROS) content, lipid peroxidation (LPO), activities of superoxide dismutase (SOD) and catalase (CAT) enzymes increased in restrained squirrels. Elevated ROS increased the oxidative load that led to ovarian cell death as evidenced by increased Bax and decreased Bcl2 expression causing further decline in Aromatase and ERα proteins. To elaborate the mechanism(s) involved in stress induced glucocorticoid mediated oxidative damages to the ovary we extended our study by exposing ovaries in vitro to the synthetic glucocorticoid dexamethasone (200 µM). We observed that glucocorticoid receptor (GR) expression was significantly increased in dexamethasone treated ovaries in vitro with a decrease in expression of Nrf2 and HO-1 proteins. Melatonin supplementation (10 nM) along with dexamethasone significantly decreased ovarian ROS production, lipid peroxidation and increased antioxidant enzyme activities by improving the expression of Nrf2 and HO-1, reinstating the cellular redox homeostasis. Therefore, it can be suggested that physical restraint induced glucocorticoid and its receptor activation interfered with the ovarian antioxidant defense mechanism. Melatonin via its receptor MT1 significantly alleviated ovarian damages acting as a cytoprotective agent.

Uterine anomalies in cell proliferation, energy homeostasis and oxidative stress in PCOS hamsters, M. auratus: Therapeutic potentials of melatonin.

AIMS: Polycystic ovarian syndrome (PCOS) is a reproductive, endocrine and metabolic disorder. Less is known about the mechanism of its effect on uterine function and therapeutic potential of melatonin. Our aim was to evaluate uterine dysfunction(s) in letrozole induced PCOS and its possible rectification by melatonin. MAIN METHODS: Adult female golden hamsters were divided into groups of Control (C), Melatonin (M; 1 mg/kg b.w.), Letrozole (L; 3 mg/kg b.w.) and combination of Letrozole+Melatonin (L + M; 3 mg/kg b.w. + 1 mg/kg b.w.) which were treated for 40 days. Analysis of serum testosterone/estradiol/progesterone/leptin/insulin, uterine histomorphometry, immunohistochemistry for proliferation cell nuclear antigen (PCNA), homeostatic assessment model of insulin resistance (HOMA-IR), western blotting for PCNA, androgen receptor (AR), insulin receptor (InsR), glucose tansporter-4 (GLUT-4), nuclear factor-kappa B (NFκB), cyclooxygenase-2 (COX-2) and biochemical analysis of superoxide dismutase (SOD)/catalase/lipid peroxidation (LPO) were done. KEY FINDINGS: Serum testosterone, leptin and insulin increased while uterine InsR/GLUT-4 expression decreased in L group indicating metabolic abnormalities. Endometrial hyperplasia, increased expression of PCNA and AR indicated abnormal proliferation in L compared to C. Increased uterine oxidative load (SOD/catalase/LPO) and inflammatory markers NFκB/COX-2 expression in L was responsible for high tissue oxidative stress and inflammation. M administration normalized all the above parameters suggesting its ameliorative effect in L + M group. SIGNIFICANCE: We report PCOS induced uterine dysfunction in Mesocricetus auratus for the first time. M administration restores uterine functions modulating cellular dynamicity, metabolic status, decreased oxidative and inflammatory load in PCOS hamsters. Therefore, we suggest the therapeutic potential of M against PCOS led uterine abnormalities to restore female fertility.

Role of melatonin and HSF-1\HSP-70 in modulating cold stress-induced immunosuppression in a tropical rodent- Funambulus pennanti [Volume 87, January 2020, 102456].

The neuroendocrine hormone melatonin and molecular chaperones (heat shock proteins) are evolutionarily conserved molecules that play an important role in protecting organisms from abiotic and biotic stressors. Environmental temperature and seasonality modulates immunity which impacts the overall health of animals. Most studies in relation to thermal stress are based on animals inhabiting temperate zones however, the substantial effect of climatic stress on tropical animals is less explored. Therefore, in this study we focused on the immunosuppressive effect of cold environment on a seasonally breeding tropical rodent and highlighted the importance of melatonin and HSF-1/Hsp-70 in regulating immunity. Animals were exposed to different temperatures with or without melatonin treatment. Our results suggest that, low temperature elicited cold-associated stress in animals marked by reduced body weight, decreased TLC/LC count in the blood and increased corticosterone production which was central to all immune alterations. Cold temperature also increased the oxidative stress which further induced apoptosis in the immune cells and activated stress response molecular chaperones HSF-1/HSP-70. Exogenous melatonin treatment not only ameliorated cold-induced immune suppression but also upregulated the expression of HSF-1 and HSP-70 in the immune cells thereby preventing protein unfolding and cell death. Thus, we conclude that melatonin and molecular chaperones synergistically alleviated immune suppression and could emerge as a promising combination therapy to target temperature stress in animals while boosting immunity.

Melatonin Ameliorates LPS-Induced Testicular Nitro-oxidative Stress (iNOS/TNFα) and Inflammation (NF-kB/COX-2) via Modulation of SIRT-1.

Lipopolysaccharide (LPS) - an endotoxin that is being extensively used in laboratory to mimic microbial infection that adversely affects male fertility. This study investigated the protective effects of melatonin on LPS-induced testicular nitro-oxidative stress, inflammation, and associated damages in the testes of male golden hamsters, Mesocricetus auratus. Hamsters were administered with melatonin and LPS for 7 days. Testes of LPS treated hamsters showed degenerative changes (appearance of vacuoles, exfoliation, and depletion of germ cells in the seminiferous tubules), adverse effects on spermatogenesis (sperm count and viability), and steroidogenesis (declined serum and testicular testosterone). Furthermore, LPS treatment decreased melatonin content, melatonin receptor (MT1), and antioxidant potential (catalase and SOD), and simultaneously increased nitro-oxidative stress (CRP, nitrate, TNFα). LPS upregulated NF-kB, COX-2, and iNOS expressions to increase testicular inflammatory load that resulted in the decrease of germ cell proliferation and survival, thus culminating into germ cell apoptosis as indicated by AO-EB staining and caspase-3 expression. Administration of melatonin with LPS showed improved testicular histoarchitecture, sperm parameters, and testosterone level. Melatonin increased testicular antioxidant status (SOD, catalase) to counteract the LPS-induced testicular ROS and thus reduced testicular nitro-oxidative stress. Furthermore, melatonin treatment upregulated testicular SIRT-1 expression to inhibit LPS-induced inflammatory proteins, i.e., NF-kB/COX-2/iNOS expression. The rescue effect of melatonin was further supported by increased germ cell survival (Bcl-2), proliferation (PCNA), and declined apoptosis (caspase-3). In conclusion, our result demonstrated that melatonin rescued testes from LPS-induced testicular nitro-oxidative stress, inflammation, and associated damages by upregulation of SIRT-1.

Retinal and extra-retinal photoreceptor responses and reproductive performance of Japanese quail (Coturnix coturnix japonica) following exposure to different photoperiodic regime.

Japanese quail is a truly photoperiodic avian species. In general long days are gonado-stimulatory and short days are gonado-inhibitory for this poultry bird. To investigate the correlation of retinal and extra-retinal photoreceptors with different photoperiodic conditions quail were divided into 2 groups and kept under long day (16L: 8D) and short day (8L: 16D) condition separately to develop photosensitivity and scotosensitivity respectively. Transfer of long day quail to intermediate day-length (13.5L: 10.5D) developed photorefractoriness (relative) and prolonged exposure to short photoperiodic conditions led the birds to develop scotorefractoriness. Increased expression of mRNA and immunosignaling of photoreceptors rhodopsin, transducin in eye and hypothalamus while decreased mRNA expression of melatonin receptors (Mel1b, Mel1c) were noted in the eyes of photosensitive (PS) and scotorefractory (SR) quail compared to photorefractory (PR) and scotosensitive (SS) birds respectively. Decreased expression of hypothalamic GnIH and melatonin receptors mRNA was observed in PS and SR birds compared to PR and SS birds respectively. Modulation of retinal and extra retinal photoreceptors leads to increased spermatogenesis as well as mRNA expression of steroidogenic genes and androgen receptor in the testis of sexually active PS and SR quail. These results led us to conclude that gonadal stimulation in PS as well as SR quail is outcome of activated retinal and extra retinal photoreceptors which lowered melatonin receptors and GnIH expression. Contrarily testicular inhibition in PR and SS is the outcome of decreased photoperception. It is suggested that decreased photoperception in SS quail increases after prolong exposure of the short day (in SR) leading to increased activity of HPG axis.

Melatonin ameliorates Bisphenol S induced testicular damages by modulating Nrf-2/HO-1 and SIRT-1/FOXO-1 expressions.

BPS has detrimental effects on human reproductive health and emerged as an environmental contaminant for global health concern. This study deals with the adverse impact of BPS exposure on testicular oxidative stress, inflammation and apoptosis in adult male golden hamster, Mesocricetus auratus and its amelioration by melatonin. BPS (75 mg/kg BW/day) exposure caused testicular impairment as evident by histological degenerative changes, declined sperm quality (viability and motility), serum levels of testosterone and melatonin with a concomitant decrease in testicular androgen receptor (AR) and melatonin receptor (MT1) expression. The BPS exposure caused marked increase in testicular oxidative load, inflammation (NF-kB/COX-2) and apoptosis (caspase-3). Melatonin (10 mg/kg BW/alternate day) pretreatment to BPS exposed hamsters resumed normal testicular histoarchitecture, sperm quality and decreased testicular oxidative load as evident by enhanced antioxidant enzymes (SOD and catalase) activities and decreased lipid peroxidation (LPO) level. Further, melatonin also stimulated the testicular antioxidant proteins Nrf-2/HO-1, SIRT-1/FOXO-1 and reduced inflammatory proteins NF-kB/COX-2 expression to counteract BPS induced testicular damages. Melatonin administration to the BPS treated hamsters resulted in increased testicular cell proliferation (PCNA), survival (Bcl-2), gap junction (connexin-43) and decreased apoptosis (caspase-3). In conclusion, our study documented the detrimental effects of BPS on testes that compromises male fertility. Further, melatonin was found as a potent molecule that rescued the BPS induced testicular damages in male golden hamster Mesocricetus auratus.

Trade-off expression of melatonin receptor subtypes (Mel1a and Mel1b) and androgen receptor in lung of a tropical bird, Perdicula asiatica.

The role of circulatory steroid hormone along with melatonin in lung of any seasonally breeding bird has never been explored so far. This could be interesting because steroid hormones are immunosuppressive while melatonin is immunostimulatory in nature. In our present study, we report the effect of exogenous melatonin and testosterone on expression of melatonin receptor subtypes (Mel1a and Mel1b ) and androgen receptor in lung of a tropical bird Perdicula asiatica. Birds were collected from vicinity of Varanasi and acclimatized in laboratory with sufficient food and water. The birds were treated with melatonin and testosterone at dose of 25 µg/100 g B.wt./day and 1 mg/100 g B.wt./day, respectively, for 28 days. At the end of the experiment, the birds were sacrificed and lung tissue and blood sample were collected for immunohistochemistry, Western blot analysis and hormonal assay. Testosterone treatment increased circulatory testosterone and upregulated expression of androgen receptors whereas downregulated expression of melatonin receptor subtypes Mel1a and Mel1b . Melatonin administration increased peripheral melatonin and upregulated expression of melatonin receptor subtypes Mel1a and Mel1b while downregulated androgen receptor. Thus, our results suggest that a trade-off relationship between melatonin and testosterone exists in regulation of their receptors in lung of Perdicula asiatica.

Fluoride Compromises Testicular Redox Sensor, Gap Junction Protein, and Metabolic Status: Amelioration by Melatonin.

The excess fluoride intake has been shown to adversely affect male reproductive health. The aim of the present study was to investigate the key mechanism underlying fluoride-induced testicular dysfunction and the role of melatonin as a modulator of testicular metabolic, oxidative, and inflammatory load. The present results indicated that sodium fluoride (NaF) exposure to adult male golden hamsters severely impairs reproductive physiology as evident from markedly reduced sperm count/viability, testosterone level, androgen receptor (AR), testicular glucose transporter (GLUT-1), gap junction (connexin-43), and survival (Bcl-2) protein expression. NaF exposure markedly increased testicular oxidative load, inflammatory (NF-kB/COX-2), and apoptotic (caspase-3) protein expression. However, melatonin treatment remarkably restored testicular function as evident by normal histoarchitecture, increased sperm count/viability, enhanced antioxidant enzyme activities (SOD and Catalase), and decreased lipid peroxidation (LPO) level. In addition, melatonin treatment upregulated testicular Nrf-2/HO-I, SIRT-1/ FOXO-1, and downregulated NF-kB/COX-2 expression. Further, melatonin ameliorated NaF-induced testicular metabolic stress by modulating testicular GLUT-1expression, glucose level, and LDH activity. Furthermore, melatonin treatment enhanced testicular PCNA, Bcl-2, connexin-43, and reduced caspase-3 expression. In conclusion, we propose the molecular mechanism of fluoride-induced testicular damages and ameliorative action(s) of melatonin.

Paraquat induced impaired reproductive function and modulation of retinal and extra-retinal photoreceptors in Japanese quail (Coturnix coturnix japonica).

Paraquat (PQ) being a potent herbicide, causes toxic effect on growth, development and reproduction of plant as well as in animals. In this study we have mainly focused on the toxic effect of PQ on photoperception via different photoreceptors present in retina, pineal and hypothalamus and thereby its effect on hypothalamic - pituitary - gonadal (HPG) axis. PQ was administered i.p.10 mg/kg body weight daily for 1 week in poultry birds Japanese quail (Coturnix coturnix japonica). Our findings clearly indicated decrease in immunoreactivity of retinal and extra retinal photoreceptors (Iodopsin, rhodopsin and transducin) following PQ treatment in comparison to control group. Increased immunoreactivity of GnIH was observed in testis and epididymis of PQ treated group. Decreased mRNA expression of photoreceptors (rhodopsin and melanopsin), steroidogenic genes, androgen receptor, GnRH-I were found in PQ treated group while increased mRNA expression of melatonin receptors (Mel 1a R, Mel 1b R, Mel 1c R) and GnIH were found in PQ treated group. Thus, from the present results it may be concluded that PQ treatment alters the photoperception via altering the expression of photoreceptors and also modulates the HPG axis thereby alters the reproductive functions in Japanese quails.