Physiological Concentrations Of Melatonin Research Articles

Reassessment of oxidative stress in idiopathic sudden hearing loss and preliminary exploration of the effect of physiological concentration of melatonin on prognosis.

The pathogenesis of idiopathic sudden sensorineural hearing loss (ISSNHL) is still unclear, and there is no targeted treatment. This research aimed to verify the role of oxidative stress in ISSNHL and explore whether melatonin has a protective effect on hearing. A total of 43 patients with ISSNHL and 15 healthy controls were recruited to detect the level of melatonin, reactive oxygen species (ROS), and total antioxidant capacity (TAC) in the blood and compared before and after treatment. Multivariate logistic regression models were performed to assess the factors relevant to the occurrence and improvement of ISSNHL. The patients with ISSNHL showed significantly higher ROS levels than controls (4.42 ± 4.40 vs. 2.30 ± 0.59; p = 0.031). The levels of basal melatonin were higher (1400.83 ± 784.89 vs. 1095.97 ± 689.08; p = 0.046) and ROS levels were lower (3.05 ± 1.81 vs. 5.62 ± 5.56; p = 0.042) in the effective group as compared with the ineffective group. Logistic regression analysis showed that melatonin (OR = 0.999, 95% CI 0.997-1.000, p = 0.049), ROS (OR = 1.154, 95% CI 1.025-2.236, p = 0.037), and vertigo (OR = 3.011, 95% CI 1.339-26.983, p = 0.019) were independent factors associated with hearing improvement. Besides, the level of melatonin (OR = 0.999, 95% CI 0.998-1.000, p = 0.023) and ROS (OR = 3.248, 95% CI 1.109-9.516, p = 0.032) were associated with the occurrence of ISSNHL. Our findings may suggest oxidative stress involvement in ISSNHL etiopathogenesis. The level of melatonin and ROS, and vertigo appear to be predictive of the effectiveness of hearing improvement following ISSNHL treatment.

Photoperiod and Melatonin Supplementation: Variable Effects on the Quality of Chilled Dog Semen.

The addition of melatonin in seminal extenders due to its antioxidant properties and its beneficial role in sperm preservation has been previously described, especially in seasonal species. The aim of this study was to study a potential seasonal effect based on photoperiod duration when adding a physiological concentration of melatonin in the canine ejaculate. A total of 24 ejaculates were obtained from 10 healthy dogs during the increasing photoperiod (from December 21 to June 21), whereas 12 ejaculates were collected from five healthy individuals during the decreasing photoperiod (from June 22 to December 20). Each ejaculate was separated into two aliquots, and one of them remained as a control, whereas melatonin (100 pM) was added to the other one (C and M treatment groups, respectively). Diluted semen was refrigerated at 5°C. On days 0, 1, 2, 3, and 6, sperm motility analyses were performed using a CASA system and hypoosmotic swelling test (HOST), osmotic resistance test (ORT), and flow cytometry analysis. No effect of melatonin on motility was detected in either photoperiod. Negative effects of melatonin were found for acrosomal defects, apoptosis, and viability in the decreasing photoperiod. The addition of melatonin to sperm in the decreasing photoperiod could create such a high level that it would cause the described negative effects. We found a beneficial effect of melatonin in the increasing photoperiod on acrosomal defects and apoptosis during 0–6 days. Melatonin treatment also increased viability in the short term (days 1 and 2) for both photoperiods. Also, melatonin can provide certain beneficial effects on mitochondrial activity in the medium term (days 2 and 3) in the decreasing photoperiod.

Possible effects of melatonin on reperfusion injury following coronary artery bypass graft surgery.

Although coronary artery bypass graft (CABG) surgery has been reported to be one of the most effective internentions in terms of myocardial salvage, reperfusion itself can cause additional damage to the myocardium. Since there is strong evidence that free radicals are the principal offender in ischemia-reperfusion (I/R) injury, it has been suggested that treatment with antioxidant agents can be protective. Investigations have shown that melatonin secretion is partially disturbed in CABG patients. The aim of this study was to evaluate the protective effect of melatonin as an antioxidant agent on I/R injury. 164 elective CABG candidates participated in this randomized clinical trial during the preoperative period. The candidates were randomized to receive 3 mg of melatonin tablets (physiologic dose) from 3 days before surgery until the day of discharge. Cardiac biomarkers [troponin and creatine kinase myocardial band (CKMB)] were assessed once before surgery (24 hours before surgery), and 8 and 24 hours after surgery. Finally, 130 patients, 65 (50%) patients in the melatonin group and 65 (50%) in the control arm finished our study. Mean age of melatonin and control groups was 59.90 ± 9.59 and 60.80 ± 8.00 years, respectively; moreover, 47 (72.30%) in melatonin and 45 (69.23%) in control group were men. No significant difference was seen in baseline cardiac biomarkers between two groups (P > 0.05). In both groups, cardiac biomarkers (CKMB and troponin) elevated after surgery in comparison to their preoperative values. There was no statistically significant difference between the control and melatonin groups regarding the 8-hour and 24-hour troponin and CKMB when adjusted for interacting factors (P > 0.05). Although physiological concentration of melatonin is protective against I/R injury, substitution of endogenous melatonin with the oral supplement which creates physiologic concentration may not prevent I/R injury. In order to have antioxidant effect, pharmacologic doses of melatonin should be employed.

Melatonin enhances hydrogen peroxide-induced apoptosis in human dental pulp cells

Background/purposeMelatonin, at physiological concentrations, was previously found to inhibit proliferation and promote odontogenic differentiation in human dental pulp cells (hDPCs), but its effect on apoptosis is unclear. Our study aimed to investigate the effect of melatonin on the H2O2-mediated viability reduction and apoptosis in hDPCs. Materials and methodshDPCs were treated with H2O2 (0, 250, 500, 1000 μmol/L), melatonin (0, 10−12, 10−10, 10−8 mol/L), and melatonin with H2O2 for 24 h. CCK-8 assays were performed to evaluate cell viability. Apoptosis was measured by DAPI and Annexin V/propidium iodide staining. Intracellular reactive oxygen species (ROS) were measured by CellROX® staining and mitochondrial membrane potential (ΔΨm) was examined by JC-1 staining. ResultsH2O2 obviously decreased the viability of hDPCs in a concentration-dependent manner and melatonin alone also reduced viability by 16–20%. Melatonin was also found to enhance H2O2-induced toxicity in a concentration-dependent manner, and the highest physiological concentration of melatonin (10−8 mol/L) had the most obvious effect (P < 0.001). Treating H2O2-exposed hDPCs with melatonin significantly increased the ratio of apoptotic cells with condensed and deformed nuclei (P < 0.001), as well as the percentage of Annexin V-positive cells (P < 0.01). Furthermore, melatonin significantly increased intracellular ROS levels and induced the loss of ΔΨm in H2O2-exposed cells (P < 0.05). ConclusionOur results indicate that melatonin, at physiological concentrations, can enhance H2O2-induced apoptosis in hDPCs and increase H2O2-mediated ROS production and ΔΨm loss. Further studies are needed to investigate whether melatonin targets the mitochondrial death pathway during the process.

Melatonin promotes neuroblastoma cell differentiation by activating hyaluronan synthase 3-induced mitophagy.

Neuroblastoma is the second most common pediatric malignancy and has a high rate of spontaneous remission. Uncovering the mechanisms underlying neuroblastoma cell differentiation is critical for therapeutic purposes. A neuroblastoma cell line (N2a) treated with either serum withdrawal (<2.5%) or melatonin (>0.1 nmol/L) for 24 hours was used as a cell differentiation research model. Interestingly, the hyaluronan synthase 3 (HAS3) protein was induced in differentiated N2a cells. N2a‐allografted nude mice received an intraperitoneal injection of melatonin (40 or 80 mg/kg/day for 3 weeks). The mean tumor volume in mice treated with 80 mg/kg melatonin was smaller than that in PBS‐treated mice (1416.3 and 3041.3 mm3, respectively, difference = 1625 mm3, *P = 0.0003, n = 7 per group). Compared with the vector control group, N2a cells with forced HAS3 overexpression showed significantly increased neuron length (*P = 0.00082) and neurite outgrowth (*P = 0.00059). Intracellular changes in autophagy, including distorted mitochondria with abnormal circular inner membranes, were detected by transmission electron microscopy (TEM). Our study demonstrated that HAS3‐mediated signaling activated by physiological concentrations of melatonin (>0.1 nmol/L) triggered significant N2a cell differentiation. These results provide molecular data with potential clinical relevance for therapeutic drug development.

Late dinner impairs glucose tolerance in MTNR1B risk allele carriers: A randomized, cross-over study

Late-night dinner eating is associated with increased risk for type-2 diabetes. The underlying mechanism is unclear. One explanatory hypothesis is that the concurrence of elevated circulating melatonin and high glucose concentrations (characterizing late eating) leads to impaired glucose tolerance. However, to date no study has tested the influence of physiological melatonin concentrations on glucose-tolerance. The discovery of melatonin receptor MTNR1B as a diabetes risk gene provides evidence for a role of physiological levels of melatonin in glucose control. The aim of our study was to test the hypothesis that elevated endogenous melatonin concentrations worsen glucose control when eating late. Registered under ClinicalTrials.gov Identifier no. NCT03003936. We performed a randomized, cross-over trial to compare glucose tolerance in the presence (late dinner) or absence (early dinner) of elevated physiological melatonin concentrations and we compared the results between homozygous carriers and non-carriers of the MTNR1B risk allele. The concurrence of meal timing with elevated endogenous melatonin concentrations resulted in impaired glucose tolerance. This effect was stronger in MTNR1B risk-carriers than in non-carriers. Furthermore, eating late significantly impaired glucose tolerance only in risk-carriers and not in the non-risk carriers. The interaction of dinner timing with MTNR1B supports a causal role of endogenous melatonin in the impairment of glucose tolerance. These results suggest that moving the dinner to an earlier time may result in better glucose tolerance specially in MTNR1B carriers. https://clinicaltrials.gov/ct2/show/NCT03003936.

Melatonin potentially acts directly on swine ovary by modulating granulosa cell function and angiogenesis.

Melatonin exerts well-known reproductive effects, mainly acting on hypothalamic gonadotrophin-releasing hormone release. More recent data suggest that melatonin acts directly at the ovarian level, even if, at present, these aspects have been only partly investigated. Swine follicular fluid contains melatonin and its concentration is significantly reduced during follicular growth. Therefore, the present study was undertaken to examine the effects of melatonin, used at physiological concentrations, on cultured swine granulosa cells collected from small (<3mm) and large (>5mm) follicles on the main parameters of granulosa cell function such as proliferation and steroidogenesis, namely oestradiol 17β and progesterone (P4) production. Moreover, the effects of melatonin on superoxide anion and nitric oxide (NO) generation by swine granulosa cells were also investigated. Finally, since angiogenesis is crucial for follicle growth, the effects of melatonin on new vessel growth were studied. Collected data indicate that melatonin interferes with cultured granulosa cell proliferation and steroidogenesis, specifically in terms of P4 production and NO output. In addition, the events of physiological follicular angiogenesis were stimulated by melatonin as evidenced by angiogenesis bioassay. Therefore, we suggest that physiological melatonin concentrations could potentially be involved in local modulation of swine ovarian follicle function.

Melatonin promotes ATO-induced apoptosis in MCF-7 cells: Proposing novel therapeutic potential for breast cancer

Arsenic trioxide (ATO), a traditional Chinese medicine, has long been of biomedical interest and is largely used for treatment of a broad spectrum of cancers. Melatonin, a naturally occurring indoleamine synthesized in the pineal gland, has been considered as a biomarker for endocrine-dependent tumors, particularly breast cancer. An increasing number of studies indicate that melatonin could be an attractive candidate for combined therapy due to its anti-oxidant and cytotoxic activities. The aim of this study was to investigate the potentiating effect of melatonin on ATO-induced apoptosis in estrogen receptor (ER)-positive breast cancer cell line, MCF-7. Our data highlighted for the first time that pre-treating MCF-7 cells with physiological concentration of melatonin substantially augmented the cytotoxic effects of ATO as compared with either agent alone. Real-time PCR analysis revealed that apoptosis induction by the drugs combination was associated with increased p53 transcriptional activity followed by the elevated molecular ratio of Bax/Bcl-2. Moreover, induced p21, subsequent G1 cell cycle arrest and transcriptional suppression of survivin-mediated c-Myc and hTERT expression may contribute in the enhanced growth suppressive effect of ATO-plus-melatonin. Due to the safety profile of melatonin, our study suggests that using melatonin in combination with ATO might provide insight into a novel adjuvant therapy and may confer advantages for breast cancer treatment.

Abstract B01: Mutagenic and tumorigenic effects of follicular fluid in the context of p53 loss: Initiation of fimbria carcinogenesis.

Abstract Ovulation is the strongest risk factor for ovarian high-grade serous carcinoma (HGSC) that largely originates from the fallopian tube fimbriae and always carries loss-of-function mutations of TP53 in both early and late lesions. Mature ovarian follicle contains high level of reactive oxygen species (ROS). When released from ovulation, follicular fluid (FF) bathes the fimbriae and may lead to DNA double strand break (DSB) and neoplastic transformation. In this study, we examined the mutagenic and tumorigenic activities of human pre-ovulatory FFs. A subset (6/11) of FFs was found with high levels of ROS whereas the antioxidant capacities were indifferent. These ROS-high FFs induced intracellular ROS and DSBs in the secretory cell population of fimbriae epithelium. In the context of p53 and Rb deficiency in an E6/E7 immortalized fimbria secretory cell line, FF-exposed secretory cells overcame apoptosis and maintained the population carrying ROS and DSB. The cancer initiation effects of FF were further recapitulated in Trp53-/- mice. When introduced into the mammary fat pad, ROS-high but not ROS-low FFs induced early-onset B-cell lymphoma. Cotreatment with physiological concentration of melatonin, a potent antioxidant, ameliorated the mutagenic and tumorigenic effect of ROS-high FF in vitro and in vivo. The study revealed ROS and mitogens in mature ovarian follicles could initiate the transformation of fimbria epithelium in the context of p53 loss and melatonin is a potent preventive agent. Citation Format: Tang-Yuan Chu, Hsuan-Shun Huang, Che-Fang Hsu. Mutagenic and tumorigenic effects of follicular fluid in the context of p53 loss: Initiation of fimbria carcinogenesis. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: Exploiting Vulnerabilities; Oct 17-20, 2015; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(2 Suppl):Abstract nr B01.

Melatonin suppression of aerobic glycolysis (Warburg effect), survival signalling and metastasis in human leiomyosarcoma.

Leiomyosarcoma (LMS) represents a highly malignant, rare soft tissue sarcoma with high rates of morbidity and mortality. Previously, we demonstrated that tissue-isolated human LMS xenografts perfused in situ are highly sensitive to the direct anticancer effects of physiological nocturnal blood levels of melatonin which inhibited tumour cell proliferative activity, linoleic acid (LA) uptake and metabolism to 13-hydroxyoctadecadienoic acid (13-HODE). Here, we show the effects of low pharmacological blood concentrations of melatonin following oral ingestion of a melatonin supplement by healthy adult human female subjects on tumour proliferative activity, aerobic glycolysis (Warburg effect) and LA metabolic signalling in tissue-isolated LMS xenografts perfused in situ with this blood. Melatonin markedly suppressed aerobic glycolysis and induced a complete inhibition of tumour LA uptake, 13-HODE release, as well as significant reductions in tumour cAMP levels, DNA content and [(3) H]-thymidine incorporation into DNA. Furthermore, melatonin completely suppressed the phospho-activation of ERK 1/2, AKT, GSK3β and NF-kB (p65). The addition of S20928, a nonselective melatonin antagonist, reversed these melatonin inhibitory effects. Moreover, in in vitro cell culture studies, physiological concentrations of melatonin repressed cell proliferation and cell invasion. These results demonstrate that nocturnal melatonin directly inhibited tumour growth and invasion of human LMS via suppression of the Warburg effect, LA uptake and other related signalling mechanisms. An understanding of these novel signalling pathway(s) and their association with aerobic glycolysis and LA metabolism in human LMS may lead to new circadian-based therapies for the prevention and treatment of LMS and potentially other mesenchymally derived solid tumours.

Mutagenic, surviving and tumorigenic effects of follicular fluid in the context of p53 loss: initiation of fimbria carcinogenesis.

Ovulation is the strongest risk factor for ovarian high-grade serous carcinoma (HGSC) that largely originates from the fallopian tube fimbriae and always carries loss-of-function mutations of TP53 in both early and late lesions. Mature ovarian follicle contains high level of reactive oxygen species (ROS). When released from ovulation, follicular fluid (FF) bathes the fimbriae and may lead to DNA double-strand break (DSB) and neoplastic transformation. In this study, we examined the mutagenic and tumorigenic activities of human pre-ovulatory FFs. A subset (6/11) of FFs was found with high levels of ROS whereas the antioxidant capacities were indifferent. These ROS(high) FFs induced intracellular ROS and DSBs in the secretory cell population of fimbriae epithelium. When p53 and Rb were turned down, the FF-exposed secretory cells overcame apoptosis and expanded the population carrying ROS and DSB. The cancer initiation and promotion effects of FF were further recapitulated in Trp53 (-/-) mice. When introduced into the mammary fat pad, ROS(high) but not ROS(low) FFs induced early-onset B-cell lymphoma. Cotreatment with physiological concentration of melatonin, a potent antioxidant, ameliorated the mutagenic and tumorigenic effect of ROS(high) FF in vitro and in vivo. The study revealed ROS and mitogens in mature ovarian follicles could initiate the transformation of fimbria epithelium in the context of p53 loss and melatonin is a potent preventive agent.

Impact of melatonin receptor deletion on intracellular signaling in spleen cells of mice after polymicrobial sepsis.

Melatonin is known to influence immune functions and to ameliorate outcome after septic challenge but it is unknown whether this is mediated by melatonin receptor activation. This study aimed to elucidate molecular differences in spleen and ex vivo splenocytes of wild-type (WT) and melatonin receptor double knockout mice (KO) after polymicrobial sepsis. C3H/HeN wild-type and MT1-/-/MT2-/- mice underwent sham operation or cecum ligation and incision (CLI) and remained anesthetized for 1 h. Splenocytes were isolated and treated in culture with physiological melatonin concentrations (1 nM). Plasma TNFα levels were consistently high after 1 h of CLI. Basal circulating leukocyte numbers were slightly higher in KO animals. We detected transcriptional differences in splenocytes of the knockout strain concerning proinflammatory mediators. Expression levels of IL-1β, IL-2, CXCR2, L-Selectin, TNFα, CXCL2 and ICAM-1 were strongly increased in splenocytes of KO mice. Splenocytes of KO mice displayed reduced ERK and p38 as well as increased JNK phosphorylation. None of the analyzed factors were influenced by melatonin in the culture medium. The results of this study indicate an increased proinflammatory status of mice deficient in both membrane-bound melatonin receptors reflected by altered activation of MAPK cascades and transcriptional activation of proinflammatory mediators.

Seasonal Changes and Regulation of the Glucocorticoid Receptor in the Testis of the Toad Rhinella arenarum

Several studies indicate that wild free-living vertebrates seasonally regulate plasma glucocorticoids. However, not only glucocorticoids but also the amount of receptors is important in determining biological responses. In this context, seasonal regulation of glucocorticoid receptor (GR) is crucial to modulate the response to glucocorticoids. Rhinella arenarum is an anuran exhibiting seasonal variations in plasma glucocorticoids and also in the number of binding sites (B(max)) of the testicular cytosolic GR. In this work, we evaluated if the annual pattern of GR protein in the testis varies seasonally and, by an in vitro approach, the role of glucocorticoids, androgens, and melatonin in the regulation of the GR B(max) and protein level. For this purpose, testes were treated with two physiological concentrations of melatonin (40 and 200 pg/ml), with or without luzindole (melatonin-receptor antagonist); with testosterone, cyanoketone (inhibitor of steroidogenesis) or casodex (androgen-receptor antagonist); or with dexamethasone or RU486 (GR antagonist). After treatments, B(max) and protein level were determined by the binding of [(3)H]dexamethasone and Western blot, respectively. Results showed that GR protein decreases in the winter. The in vitro treatment with melatonin produced a biphasic effect on the B(max) with the lowest concentration decreasing this parameter by a receptor-mediated mechanism. However, melatonin had no effect on the GR protein level. Conversely, a high concentration of dexamethasone up-regulated the GR protein and androgens neither changed the B(max) nor the protein level. These findings suggest that seasonal changes in plasma melatonin and glucocorticoids modulate the effect of glucocorticoids in the testis of R. arenarum.

Melatonin inhibits glucocorticoid-dependent GR–TIF2 interaction in newborn hamster kidney (BHK) cells

The antagonism exerted by melatonin on the glucocorticoid response has been well established, being strongly dependent on the cellular context. Previously, we found that melatonin inhibits glucocorticoid receptor (GR) dissociation from the chaperone hetero-complex and nuclear translocation on mouse thymocytes. Here, by performing confocal fluorescence microscopy and the Number and Brightness assay we show that in newborn hamster kidney cells (BHK21) melatonin neither affects GR nuclear translocation nor GR homodimerization. Instead, co-immunoprecipitation studies suggest that physiological concentrations of melatonin impair GR interaction with the transcriptional intermediary factor 2 (TIF2). This melatonin effect was not blocked by the MT1/MT2 receptor antagonist luzindole. Curiously, luzindole behaved as an antiglucocorticoid per se by impairing the glucocorticoid-dependent MMTV-driven gene expression affecting neither GR translocation nor GR–TIF2 interaction.

Melatonin in plants.

Once thought to be exclusively a molecule of the animal kingdom, melatonin has now been found to exist in plants as well. Among a number of actions, melatonin is a direct free radical scavenger and an indirect antioxidant. Melatonin directly detoxifies the hydroxyl radical (OH), hydrogen peroxide, nitric oxide, peroxynitrite anion, peroxynitrous acid, and hypochlorous acid. The products from each of these reactions have been identified in pure chemical systems and in at least one case in vivo; the interaction product of melatonin with the OH, ie., cyclic 3-hydroxymelatonin, is found in the urine of humans and rats. Some of the products that are produced when melatonin detoxifies reactive species are also highly efficient scavengers. As a result, a cascade of scavenging reactions may enhance the antioxidant capacity of melatonin. Additionally, melatonin increases the activity of several antioxidative enzymes, thereby improving its ability to protect macromolecules from oxidative stress. Melatonin is endogenously produced and is also consumed in edible plants. In animal experiments, feeding melatonin-containing foods raised blood levels of the indole. Because physiologic concentrations of melatonin in the blood are known to correlate with the total antioxidant capacity of the serum, consuming food-stuffs containing melatonin may be helpful in lowering oxidative stress.

Melatonin is up‐regulator on small G‐protein mediated apoptotic cell death in cervical and endometrial cancer cells of human

Many gynecologic tumors are associated with infertility and spontaneous abortion that constitute a public problem. Recent studies have shown the main hormone of the pineal gland inhibit directly many different types of human tumor cells and naturally secreted cytotoxin, which induces tumor cell death(apoptosis). Additionally, activated Ras proteins have important roles on the regulation of apoptosis depending on cell type and other factors. Ras protein, a small GTP binding protein, send growth stimulating message to the nucleus through downstream small G protein such as Rac1 and Cdc42. Ras protein is an initiator of many tumors, and melatonin shows oncostatic action in various tumors cells. Whether melatonin can counteract the Ras‐involved functions remains unknown and is worth exploring. Thus, our overall hypothesis is that natural hormone melatonin regulates small GTP binding protein mediated apoptosis and we would demonstrate anti‐tumor function of melatonin in primary cultured human uterus cervix and endometrial cancer cells. What Ras gene transfected into cells were treated with melatonin, then observed increasing caspase‐3 expression suggesting turn of the apoptotic signal machinery. At physiological concentration of melatonin (1 nM), cell proliferation was decreased, suggesting cell cycle arrest through increasing p21 expression, which is mediated by p53(tumor suppressor gene). These observations indicate that melatonin, an endogenous hormone, inhibits the tumor cell proliferation suggesting melatonin should be developed as a therapeutic reagent.Supported by BioGreen21 Program (#20070401034006) of Rural Development Administration, South Korea

Epigenetic targets for melatonin: induction of histone H3 hyperacetylation and gene expression in C17.2 neural stem cells

We have reported the induction of glial cell line-derived neurotrophic factor, a potent survival factor for dopaminergic neurons, in the C17.2 neural stem cell line following in vitro treatment with melatonin. Furthermore, we have detected the melatonin MT(1) receptor in these cells. Given these findings and recent evidence that melatonin may play a role in cellular differentiation, we examined whether this indoleamine induces morphological and transcriptional changes suggestive of a neuronal phenotype in C17.2 cells. Moreover, in order to extend preliminary evidence of a potential role for melatonin in epigenetic modulation, its effects on the mRNA expression of several histone deacetylase (HDAC) isoforms and on histone acetylation were examined. Physiological concentrations of melatonin (nanomolar range) increased neurite-like extensions and induced mRNA expression of the neural stem cell marker, nestin, the early neuronal marker beta-III-tubulin and the orphan nuclear receptor nurr1 in C17.2 cells. The indoleamine also significantly increased mRNA expression for various HDAC isoforms, including HDAC3, HDAC5, and HDAC7. Importantly, treatment with melatonin for 24 hr caused a significant increase in histone H3 acetylation, which is associated with chromatin remodeling and gene transcription. Since the melatonin MT(2) receptor was not detected in C17.2 cells, it is likely that the MT(1) receptor is involved in mediating these physiological effects of melatonin. These findings suggest novel roles for melatonin in stem cell differentiation and epigenetic modulation of gene transcription.

Selective estrogen enzyme modulator actions of melatonin in human breast cancer cells

Melatonin exerts oncostatic effects on different kinds of neoplasias, especially on estrogen-dependent mammary tumors. Current knowledge about the mechanisms by which melatonin inhibits the growth of breast cancer cells point to an interaction of melatonin with estrogen-responsive pathways. The intratumoral production of estrogens in breast carcinoma tissue plays a pivotal role in the proliferation of mammary tumoral cells and its blockade is one of the main objectives of the treatment of breast cancer. The aim of the present work is centered on the study of the role of melatonin in the control of some enzymes involved in the formation and transformation of estrogens in human breast cancer cells. The present study demonstrates that melatonin, at physiologic concentrations, modulates the synthesis and transformation of biologically active estrogens in MCF-7 cells, through the inhibition of sulfatase (STS) and 17beta-hydroxysteroid dehydrogenase type 1 (17beta-HSD1) activity and expression, enzymes involved in the estradiol formation in breast cancer cells. Physiologic concentrations of melatonin also stimulate the activity and expression of estrogen sulfotransferase (EST), the enzyme responsible for the formation of the biologically inactive estrogen sulfates. The level of EST mRNA steady-state of cells treated with melatonin was three times higher than that in control cells. These findings which document that melatonin has an inhibitory effect on STS and 17beta-HSD1 and a stimulatory effect on EST, in combination with its previously described antiaromatase effect, can open up new and interesting possibilities in clinical applications of melatonin in breast cancer.

Effects of melatonin on the proliferation and differentiation of rat adipose-derived stem cells.

Osteogenesis driven by adipose-derived stem cells (ADSCs) is regulated by physiological and pathological factors. Accumulating evidence from in vitro and in vivo experiments suggests that melatonin may have an influence on bone formation. However, little is known about the effects of melatonin on osteogenesis, which thus remains to be elucidated. This study was performed to determine whether melatonin at physiological concentrations (0.01-10 nM) could affect the in vitro proliferation and osteogenic differentiation of rat ADSCs. ADSCs were isolated from the fat of adult rats. After cell expansion in culture media and through three passages, osteogenesis was induced in a monolayer culture using osteogenic medium with or without melatonin at physiological concentrations (0.01-10 nM). After four weeks, the cultures were examined for mineralization by Alizarin Red S and von Kossa staining and for alkaline phosphatase (ALP) activity using an ALP kit. Cell viability and apoptosis were also assayed by 3-(4, 5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTT) assay and flow cytometry, respectively. The results indicated that at physiological concentrations, melatonin suppressed proliferation and differentiation of ADSCs. These data indicate that ADSCs exposed to melatonin, had a lower ALP activity in contrast to the cells exposed to osteogenic medium alone. Similarly, mineral deposition (calcium level) also decreased in the presence of melatonin. Flow cytometry confirmed that cell growth had decreased and that the numbers of apoptotic cells had increased. These results suggest that the physiological concentration of melatonin has a negative effect on ADSC osteogenesis.

Effects of melatonin on the proliferation and differentiation of rat adipose-derived stem cells

Background:Osteogenesis driven by adipose-derived stem cells (ADSCs) is regulated by physiological and pathological factors. Accumulating evidence from in vitro and in vivo experiments suggests that melatonin may have an influence on bone formation. However, little is known about the effects of melatonin on osteogenesis, which thus remains to be elucidated. This study was performed to determine whether melatonin at physiological concentrations (0.01-10 nM) could affect the in vitro proliferation and osteogenic differentiation of rat ADSCs.Materials and Methods:ADSCs were isolated from the fat of adult rats. After cell expansion in culture media and through three passages, osteogenesis was induced in a monolayer culture using osteogenic medium with or without melatonin at physiological concentrations (0.01-10 nM). After four weeks, the cultures were examined for mineralization by Alizarin Red S and von Kossa staining and for alkaline phosphatase (ALP) activity using an ALP kit. Cell viability and apoptosis were also assayed by 3-(4, 5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTT) assay and flow cytometry, respectively.Results:The results indicated that at physiological concentrations, melatonin suppressed proliferation and differentiation of ADSCs. These data indicate that ADSCs exposed to melatonin, had a lower ALP activity in contrast to the cells exposed to osteogenic medium alone. Similarly, mineral deposition (calcium level) also decreased in the presence of melatonin. Flow cytometry confirmed that cell growth had decreased and that the numbers of apoptotic cells had increased.Conclusion:These results suggest that the physiological concentration of melatonin has a negative effect on ADSC osteogenesis.