Melatonin Receptor Research Articles

Potential role of melatonin in prevention and treatment of lung cancer.

Lung cancer is the second most common cancer and the most lethal cancer worldwide. Melatonin, an indoleamine produced in the pineal gland, shows anticancer effects on a variety of cancers, especially lung cancer. Herein, we clarify the pathophysiology of lung cancer, the association of circadian rhythm with lung, and the relationship between shift work and the incidence of lung cancer. Special focus is placed on the role of melatonin receptors in lung cancer, the relationship between inflammation and lung cancer, control of cell proliferation, apoptosis, autophagy, and immunomodulation in lung cancer by melatonin. A review of the drug synergy of melatonin with other anticancer drugs suggests its usefulness in combination therapy. In summary, the information compiled may serve as a comprehensive reference for the various mechanisms of action of melatonin against lung cancer, as a guide for the design of future experimental research and for advancing melatonin as a therapeutic agent for lung cancer.

Effect of a hyaluronic acid-based mesotherapeutic injectable on the gene expression of CLOCK and Klotho proteins, and environmentally induced oxidative stress in human skin cells.

Normal circadian rhythms are essential to the repair mechanisms of oxidative stress implicated in skin aging. Given reports that hyaluronic acid (HA) homeostasis exhibits a different profile in chronological skin aging, as compared to environmental or extrinsic aging, an improved understanding of the way HA interacts with its surroundings, and the impact of HA injectables in replacing lost HA and encouraging rejuvenation, is of key benefit to skin aging treatments. The objectives of these current studies were twofold. Firstly, to demonstrate the in vitro effects of two lightweight hyaluronic-based injectables on the expression of CLOCK protein in human skin fibroblasts, and their effects on Klotho protein expression as a marker for circadian rhythms in a combined human keratinocyte and Merkel cell model. Secondly, to ascertain whether these findings could be correlated with in vitro effects on various environmental oxidative stress aging markers (blue light, UVA/UVB, Urban Dust, and IR exposures). Oxidative stress studies were aimed to highlight possible protective effects through different challenge conditions in two models, ex vivo human skin explants and in vitro monolayer cultures of normal human dermal fibroblasts (NHDF). The protective effects of the test products were evaluated against an increase of cyclobutene pyrimidine dimers (CPDs) abundance within epidermal section of ex vivo skin explants after UVA/UVB radiation; effects of blue light on gene expression from NHDFs fibroblasts; effects of pollutants (Urban dust, UbD) on gene expression in NHDFs fibroblasts; and an increase of reactive oxygen species (ROS) production by NHDFs fibroblasts after infrared-A radiation. Gene expression was assayed and analyzed utilizing microfluidic TaqMan qPCR arrays. CLOCK expression was measured in young and senescing NHDFs by immunostaining, and Klotho and melatonin expression by immunostaining in Merkel cell-enriched normal adult human epidermal cell cultures. In an aging culture of mixed keratinocyte and Merkel skin cells, activation of Klotho expression was induced by the application of both HA test products. Moreover, the HA products increase Klotho protein expression in both Merkel cells and keratinocytes. The observed positive effect of the tested products on melatonin receptors 1A and 1B expression in aging Merkel cell culture and keratinocytes is also interesting. HA-Y (developed for patients 25+ years old) stimulated melatonin receptors type 1B expression in aging cell cultures more strongly than HA-S (developed for patients 35-65 years old). In age (stressed) cells, a lower expression of Klotho protein and melatonin receptors 1A and 1B is apparent. The addition of HA-Y and HA-S stimulates their expression thus providing a "protective" effect. The blue light irradiation at 40 J/cm2 performed in NHDF fibroblast cultures led to a modification of the expression of several genes, all involved in mechanisms known to be modulated in case of solar radiation stress. Although these are preliminary findings, they are the first we know of that demonstrate HA facial injectables having a benefit and possibilities beyond the "physical filling" of the skin. As regards the beneficial effects against blue light-induced oxidative stress, and a return to cellular homeostasis, there is a need to conduct further and more precise investigations into HA-S. Furthermore, the benefit of these HA injectables (Novacutan®) in the modulation of oxidative stressed circadian rhythms widens their potential benefit.

Melatonin Inhibits NF-κB/CREB/Runx2 Signaling and Alleviates Aortic Valve Calcification.

Calcific aortic valve disease (CAVD) is linked to high mortality. Melatonin inhibits nuclear factor-kappa B (NF-κB)/cyclic AMP response element-binding protein (CREB), contributing to CAVD progression. This study determined the role of melatonin/MT1/MT2 signaling in valvular interstitial cell (VIC) calcification. Western blotting and Alizarin red staining were used to analyze NF-κB/CREB/runt-related transcription factor 2 (Runx2) signaling in porcine VICs treated with an osteogenic (OST) medium without (control) or with melatonin for 5 days. Chromatin immunoprecipitation (ChIP) assay was used to analyze NF-κB's transcription regulation of NF-κB on the Runx2 promoter. OST medium-treated VICs exhibited a greater expression of NF-κB, CREB, and Runx2 than control VICs. Melatonin treatment downregulated the effects of the OST medium and reduced VIC calcification. The MT1/MT2 antagonist (Luzindole) and MT1 receptor neutralized antibody blocked the anticalcification effect of melatonin, but an MT2-specific inhibitor (4-P-PDOT) did not. Besides, the NF-κB inhibitor (SC75741) reduced OST medium-induced VIC calcification to a similar extent to melatonin at 10 nmol/L. The ChIP assay demonstrated that melatonin attenuated OST media increased NF-κB binding activity to the promoter region of Runx2. Activation of the melatonin/MT1-axis significantly reduced VIC calcification by targeting the NF-κB/CREB/Runx2 pathway. Targeting melatonin/MT1 signaling may be a potential therapeutic strategy for CAVD.

Regulation of the Cell Cycle, Apoptosis, and Proline Accumulation Plays an Important Role in the Stress Response of the Eastern Oyster Crassostrea Virginica

BackgroundUnderstanding how organisms respond and adapt to environmental changes is central to evolutionary biology. As a sessile organism that has adapted to life in estuaries and intertidal zones, the eastern oyster Crassostrea virginica can tolerate wide fluctuations in temperature and salinity and survive for weeks out of water. To understand the molecular mechanisms underlying the remarkable stress tolerance of the eastern oyster, we studied the transcriptomic changes induced by exposure to air and cold stress. Eastern oysters were maintained for 7 days under four conditions, namely, in seawater (normal) at 22°C, in air at 22°C, in seawater at 5°C and in air at 5°C, and then sampled for RNA sequencing.ResultsTranscriptomic analysis revealed that many genes involved in cell cycle progression and DNA replication were downregulated in oysters exposed to air and cold, which indicates that stress inhibits cell division. Exposure to air at 22°C induced a concerted inhibition of apoptosis through the upregulation of expanded inhibitors of apoptosis and the downregulation of caspases. Interactions between TNF and NF-κB signalling implied a reduction in the inflammatory response and immune functions. Key genes for proline production, fatty acid synthesis and chromosomal proteins were upregulated during exposure to low temperatures, which suggested that proline accumulation, energy conservation, and epigenetic modification of chromosomes are important for coping with cold stress. The upregulation of melatonin, FMRFamide, and neural acetylcholine receptors indicate the significance of the neurohormonal regulation of homeostasis.ConclusionThese results show that air exposure and cold stress alter the expression of key genes for cell division, apoptosis, proline accumulation, fatty acid metabolism, neurohormonal signalling, and epigenetic modifications, suggesting regulation of these processes plays an important role in the stress response of the eastern oyster and possibly other marine molluscs. This study provides new insights into molecular mechanisms of stress response that are essential for understanding the adaptive potential of marine organisms under climate change.

Hypnotics and infections: disproportionality analysis of the U.S. Food & Drug Administration adverse event reporting system database.

There is no consensus information on infections associated with nonbenzodiazepines. Knowledge about infections related to newly marketed hypnotics (orexin receptor antagonists and melatonin receptor agonists) is scarce. The study aimed to detect infection signals for nonbenzodiazepines, orexin receptor antagonists, and melatonin receptor agonists by analyzing data from the U.S. Food & Drug Administration adverse event reporting system. A disproportionality analysis was performed to quantitatively detect infection signals for hypnotics by calculating the reporting odds ratio and the 95% confidence interval. Data registered in the U.S. Food & Drug Administration adverse event reporting system from 2010-2020 were retrieved. A total of 3,092 patients with infection were extracted for the 3 classes of hypnotic drugs. Nonbenzodiazepines were associated with a higher disproportionality of infections (reporting odds ratio: 1.10; 95% confidence interval, 1.06-1.14). The association of infections was not present for melatonin receptor agonists (reporting odds ratio: 0.86; 95% confidence interval, 0.74-1.00) and orexin receptor antagonists (reporting odds ratio: 0.19; 95% confidence interval, 0.15-0.25). Significant reporting associations were identified for nonbenzodiazepines concerning the categories of bone and joint infections, dental and oral soft tissue infections, upper respiratory tract infections, and urinary tract infections. Nonbenzodiazepines had a positive signal for infections, while orexin receptor antagonists and melatonin receptor agonists had a negative signal. More research needs to be conducted to confirm this relationship. Meng L, Huang J, He Q, etal. Hypnotics and infections: disproportionality analysis of the U.S. Food & Drug Administration adverse event reporting system database. J Clin Sleep Med. 2022;18(9):2229-2235.

Melatonin inhibits angiotensin II–induced atrial fibrillation through preventing degradation of Ang II Type I Receptor–Associated Protein (ATRAP)

Angiotensin II (Ang II) induced Atrial fibrillation (AF) often accompanied with reduced ATRAP which is a negative modulator of Ang II type 1 receptor (AT1R). Melatonin can protect against AF, but the underlying molecular mechanism remains poorly understood. In this study, Ang II was used to induce AF, and AF inducibility and duration were documented telemetrically. Ang II-infused mice had a higher AF incidence, which was associated with atrial fibrosis, inflammation, and oxidative stress. Melatonin partially inhibited these effects, and enforced expression of siRNA-ATRAP in atria counteracted the beneficial role of melatonin. Specifically, melatonin inhibited expression of Ang II-induced proteasome and immunoproteasome subunits β2, β2i, β5, and β5i as well as their corresponding trypsin-like and chymotrypsin-like activities and blocked ATRAP degradation. In turn, this inhibited AT1R-mediated NF-κB signaling, transforming growth factor (TGF)-β1/Smad signaling in the atria, and thereby affected atrial remodeling and AF. Melatonin receptor inhibition by the chemical inhibitor luzindole partially inhibited the inhibitory effects of melatonin on proteasome activity and also Ang II-induced pathological changes in the atria. Overall, our study demonstrates that melatonin protects against Ang II-induced AF by inhibiting proteasome activity and stabilizing ATRAP expression, and these effects are partially dependent on melatonin receptor activation.

Melatonergic Receptors (Mt1/Mt2) as a Potential Additional Target of Novel Drugs for Depression.

A complex pathogenesis involving several physiological systems is theorized to underline the development of depressive disorders. Depression is accompanied by circadian regulation disruption and interaction with the functioning of both central and peripheral oscillators. Many aspects of melatonin function unite these systems. The use of drugs for circadian rhythm disorders could inspire a potential treatment strategy for depression. Melatonin plays an essential role in the regulation of circadian rhythms. It exerts effect by activating two types of melatonin receptors, type 1A (MT1) and 1B (MT2). These are G-protein-coupled receptors, predominantly located in the central nervous system. MT1/MT2 agonists could be a useful treatment approach according to all three prevalent theories of the pathogenesis of depression involving either monoamines, synaptic remodeling, or immune/inflammatory events. MT1/MT2 receptors can be a potential target for novel antidepressants with impact on concentrations of neurotrophins or neurotransmitters, and reducing levels of pro-inflammatory cytokines. There is an interesting cross-talk mediated via the physical association of melatonin and serotonin receptors into functional heteromers. The antidepressive and neurogenetic effects of MT1/MT2 agonists can also be caused by the inhibition of the acid sphingomyelinase, leading to reduced ceramide, or increasing monoamine oxidase A levels in the hippocampus. Compounds targeting MT1 and MT2 receptors could have potential for new anti-depressants that may improve the quality of therapeutic interventions in treating depression and relieving symptoms. In particular, a combined effect on MT1 and/or MT2 receptors and neurotransmitter systems may be useful, since the normalization of the circadian rhythm through the melatonergic system will probably contribute to improved treatment. In this review, we discuss melatonergic receptors as a potential additional target for novel drugs for depression.

The rs10830963 Polymorphism of the MTNR1B Gene: Association With Abnormal Glucose, Insulin and C-peptide Kinetics.

BackgroundThe MTNR1B gene encodes a receptor for melatonin, a hormone regulating biorhythms. Disruptions in biorhythms contribute to the development of type 2 diabetes mellitus (T2DM). Genetic studies suggest that variability in the MTNR1B gene affects T2DM development. Our aim was to compare the distribution of the genetic variant rs10830963 between persons differing in glucose tolerance in a sample of the Czech population (N=1206). We also evaluated possible associations of the polymorphism with insulin sensitivity, beta cell function, with the shape of glucose, insulin and C-peptide trajectories measured 7 times during a 3-hour oral glucose tolerance test (OGTT) and with glucagon response. In a subgroup of 268 volunteers we also evaluated sleep patterns and biorhythm.Results13 persons were diagnosed with T2DM, 119 had impaired fasting blood glucose (IFG) and/or impaired glucose tolerance (IGT). 1074 participants showed normal results and formed a control group. A higher frequency of minor allele G was found in the IFG/IGT group in comparison with controls. The GG constellation was present in 23% of diabetics, in 17% of IFG/IGT probands and in 11% of controls. Compared to CC and CG genotypes, GG homozygotes showed higher stimulated glycemia levels during the OGTT. Homozygous as well as heterozygous carriers of the G allele showed lower very early phase of insulin and C-peptide secretion with unchanged insulin sensitivity. These differences remained significant after excluding diabetics and the IFG/IGT group from the analysis. No associations of the genotype with the shape of OGTT-based trajectories, with glucagon or with chronobiological patterns were observed. However, the shape of the trajectories differed significantly between men and women.ConclusionIn a representative sample of the Czech population, the G allele of the rs10830963 polymorphism is associated with impaired early phase of beta cell function, and this is evident even in healthy individuals.

Bos taurus and Cervus elaphus as Non-Seasonal/Seasonal Models for the Role of Melatonin Receptors in the Spermatozoon.

Melatonin is crucial in reproduction due its antioxidant, hormonal, and paracrine action. Melatonin membrane receptors (MT1/MT2) have been confirmed on spermatozoa from several species, but functionality studies are scarce. To clarify their role in ruminants as reproductive models, bull (Bos taurus, non-seasonal) and red deer (Cervus elaphus, highly seasonal) spermatozoa were analyzed after 4 h of incubation (38 °C, capacitating media) in 10 nM melatonin, MT1/MT2 agonists (phenylmelatonin and 8M-PDOT), and antagonists (luzindole and 4P-PDOT). Motility and functionality (flow cytometry: viability, intracellular calcium, capacitation status, reactive oxygen species (ROS) production, and acrosomal and mitochondrial status) were assessed. In bull, MT1 was related to sperm viability preservation, whereas MT2 could modulate cell functionality to prevent excess ROS produced by the mitochondria; this action could have a role in modulating sperm capacitation. Deer spermatozoa showed resistance to melatonin and receptor activation, possibly because the samples were of epididymal origin and collected at the breeding season’s peak, with high circulating melatonin. However, receptors could be involved in mitochondrial protection. Therefore, melatonin receptors are functional in the spermatozoa from bull and deer, with different activities. These species offer models differing from traditional laboratory experimental animals on the role of melatonin in sperm biology.

Melatonin Attenuates Methamphetamine-Induced Alteration of Amyloid β Precursor Protein Cleaving Enzyme Expressions via Melatonin Receptor in Human Neuroblastoma Cells.

Alzheimer's disease (AD) is the most prominent neurodegenerative disease represented by the loss of memory and cognitive impairment symptoms and is one of the major health imperilments among the elderly. Amyloid (Aβ) deposit inside the neuron is one of the characteristic pathological hallmarks of this disease, leading to neuronal cell death. In the amyloidogenic processing, the amyloid precursor protein (APP) is cleaved by beta-secretase and γ-secretase to generate Aβ. Methamphetamine (METH) is a psychostimulant drug that causes neurodegeneration and detrimental cognitive deficits. The analogy between the neurotoxic and neurodegenerative profile of METH and AD pathology necessitates an exploration of the underlying molecular mechanisms. In the present study, we found that METH ineluctably affects APP processing, which might contribute to the marked production of Aβ in human neuroblastoma cells. Melatonin, an indolamine produced and released by the pineal gland as well as other extrapineal, has been protective against METH-induced neurodegenerative processes, thus rescuing neuronal cell death. However, the precise action of melatonin on METH has yet to be determined. We further propose to investigate the protective properties of melatonin on METH-induced APP-cleaving secretases. Pretreatment with melatonin significantly reversed METH-induced APP-cleaving secretases and Aβ production. In addition, pretreatment with luzindole, a melatonin receptor antagonist, significantly prevented the protective effect of melatonin, suggesting that the attenuation of the toxic effect on METH-induced APP processing by melatonin was mediated via melatonin receptor. The present results suggested that melatonin has a beneficial role in preventing Aβ generation in a cellular model of METH-induced AD.

New Uses of Melatonin as a Drug; A Review.

Melatonin is a simple compound called N-acetyl-5-methoxy tryptamine and is known as a hormone controlling the circadian rhythm. Melatonin is produced by the human body naturally. The production of melatonin increases in the evening, causing a state of sleep at night and wakefulness during the day. Melatonin interacts with melatonin receptors MT1 and MT2, but it was revealed that melatonin is a strong antioxidant and it also has a role in the regulation of the cell cycle. Melatonin is now used as a medicine to treat some types of sleep disorders. Moreover, recent research suggests that it can also be utilized for other purposes, such as prevention from chemical exposure and treatment of various diseases, including cancer, neurodegenerative disorders, etc. This review summarizes the basic facts and direction of the current research on melatonin. The existing literature was scrutinized for this review.

Melatonin Receptor 1B Polymorphisms and Reproductive Failure – a Pilot Study

The present study aimed to investigate the possible associations between the melatonin receptor 1B (MTNR1b) genetic polymorphisms and the presence of reproductive disturbances due to spontaneous abortion and implantation failure leading to pregnancy loss. The patients group included 35 women presenting with one or more miscarriages and/or implantation failure. Another 36 healthy women without history of implantation failure or miscarriages and with at least one birth served as a control group. Genomic DNA was extracted and genotyping for the MTNR1b polymorphisms rs1562444, rs10830962 and rs10830963 was performed by PCR-RFLP analysis. The genotype distribution of the investigated MTNR1b polymorphisms in patients with reproductive failure did not differ significantly to the group of women with at least one birth (p > 0.05 for all). Nevertheless, the presence of a haplotype rs1562444 AA rs10830962 GG was significantly less common in women with miscarriages or implantation failure than in women with at least one birth (2.9% vs. 22.2%, p = 0.028). We suggest that the haplotype might be protective against implantation failure and spontaneous miscarriage. Additionally, rs1562444 AA genotype was not found in women with recurrent pregnancy loss. Studies in larger populations are necessary to reveal the proper associations between different MTNR1b haplotypes, ovarian function and reproductive failure.

0446 Trends and Patterns in Prescriptions of Hypnotics for The Treatment of Insomnia in Japan: Analysis of A Nationwide Japanese Claims Database

Abstract Introduction Optimal treatment of insomnia is uncommon, given the lack of awareness regarding insomnia management. While the treatment landscape for insomnia has evolved following the introduction of orexin receptor antagonists (ORA), few studies have examined prescribing patterns of hypnotics. We analyzed data from a claims database to examine real-world use of hypnotics in Japan. Methods Patients (aged ≥20 to <75 years old) with insomnia diagnosis, prescribed ≥1 hypnotic and continuous enrollment for ≥12 months before the index date were extracted from the JMDC claims database between April 1st, 2009 and March 31st, 2020. Patients were classified as new users of hypnotics or long-term users (prescribed the same mechanism of action [MOA] for ≥180 days). Trends (2010–2019) and patterns (2018–2019) in hypnotics prescriptions were analyzed. Results The analysis comprised of 130,177 new users and 91,215 long-term users (2010–2019). Nearly all new users (97.1%–97.9%) were prescribed a single MOA in each year. In 2010, almost all new users (94.0%) of hypnotics were prescribed GABAA-receptor agonists (benzodiazepines [BZD] or z-drugs). The proportions of patients prescribed BZD declined over time (from 54.8% in 2010 to 30.5% in 2019), whereas those prescribed z-drugs remained stable (~40%). The proportion of patients prescribed a melatonin receptor agonist increased slightly (3.2% to 6.3%), while those prescribed ORA increased substantially (0% to 20.2%). Among long-term users, the proportion of patients prescribed BZD steadily declined over time, but more than half were prescribed BZD. Unlike new users, a lower proportion of long-term users were prescribed ORA (0% in 2010, 4.3% in 2019). Analyses using 2018–2019 data showed that a combination of multiple (≥2) MOAs was prescribed to a higher proportion (18.2%) of long-term users than new users (2.8%). The prescription patterns of hypnotics were comparable among patients stratified by age, sex, medical specialty, and psychiatric comorbidities. Conclusion The present study showed distinct characteristics in the patterns and trends of the prescriptions of hypnotics among new users and long-term users in Japan. The high proportion of long-term BZD users suggests the need for educating clinicians about the optimal care pathway for insomnia. Support (If Any) MSD K.K., Tokyo, Japan

0456 Comparative Safety and Efficacy of Hypnotics: A Quantitative Risk-Benefit Analysis

Abstract Introduction Hypnotics continue to be preferentially used in practice for the treatment of chronic insomnia, yet the comparative safety and efficacy of medication options are unclear. While several position statements and network meta-analyses have provided some guidance, none have provided a quantitative assessment of risk-benefit. In the present analysis, each therapeutic class commonly used for chronic insomnia is quantitatively ranked with respect to safety, efficacy, and risk-benefit. Methods Safety data for FDA-approved hypnotics were extracted from the package insert adverse event tables and standardized to form a placebo-adjusted adverse event rate per 1000 (AEr). Efficacy data were extracted from randomized controlled trials identified in professional society position statements and published systematic reviews. The efficacy metric was computed from placebo adjusted pre-post change scores for self-reported sleep continuity (i.e., sleep latency [SL], wake after sleep onset [WASO], and total sleep time [TST]) and represented as a summed composite effect size score (SWT). Risk-benefit (R/B) was represented as a ratio between AEr and SWT. Comparative safety, efficacy, and risk-benefit metrics were calculated for five therapeutic classes: Benzodiazepines (BZs), non-benzodiazepine benzodiazepine receptor agonists (BZRAs), dual orexin receptor antagonists (DORAs), melatonin receptor agonists (MELAs), and sedating antidepressants (SADs). Results With respect to safety, MELAs had the lowest adverse event rate (AEr=43.1) and BZRAs had the highest rate (AEr=255.0). With respect to efficacy, BZs were the most efficacious (SWT=1.94, Mean ES=0.59) and MELAs were least efficacious (SWT=0.11, Mean ES=0.02). Overall, with respect to risk-benefit, SADs had the most favorable profile (R/B=69.5), while MELAs had the least favorable profile (R/B=395.7). Conclusion The optimal selection of hypnotics requires consideration of both risk and benefit. Findings suggest that SADs can be considered the first-line pharmacotherapeutic option due to the superior risk-benefit profile. If treatment responses are inadequate or if SADs are contraindicated, risk-benefit rankings may serve as a decision tree as part of a medical algorithm protocol to guide treatment selection for patients with chronic insomnia. Support (If Any) Sleep Research Society Mentor-Mentee Award (Cheung); K24AG055602 & R01AG054521 (Perlis)

Oncostatic activities of melatonin: Roles in cell cycle, apoptosis, and autophagy [Biochimie 200 (2022) 44–59

Melatonin, the major secretory product of the pineal gland, not only regulates circadian rhythms, mood, and sleep but also has actions in neoplastic processes which are being intensively investigated. Melatonin is a promising molecule which considered a differentiating agent in some cancer cells at both physiological and pharmacological concentrations. It can also reduce invasive and metastatic status through receptors MT1 and MT2 cytosolic binding sites, including calmodulin and quinone reductase II enzyme, and nuclear receptors related to orphan members of the superfamily RZR/ROR. Melatonin exerts oncostatic functions in numerous human malignancies. An increasing number of studies report that melatonin reduces the invasiveness of several human cancers such as prostate cancer, breast cancer, liver cancer, oral cancer, lung cancer, ovarian cancer, etc. Moreover, melatonin's oncostatic activities are exerted through different biological processes including antiproliferative actions, stimulation of anti-cancer immunity, modulation of the cell cycle, apoptosis, autophagy, the modulation of oncogene expression, and via antiangiogenic effects. This review focuses on the oncostatic activities of melatonin that targeted cell cycle control, with special attention to its modulatory effects on the key regulators of the cell cycle, apoptosis, and telomerase activity.

Activation of MT1/MT2 to Protect Testes and Leydig Cells against Cisplatin-Induced Oxidative Stress through the SIRT1/Nrf2 Signaling Pathway.

There is growing concern that chemotherapy drugs can damage Leydig cells and inhibit the production of testosterone. Increasing evidence shows that melatonin benefits the reproductive process. This study mainly explores the protective effect and possible molecular mechanism of melatonin regarding cisplatin-induced oxidative stress in testicular tissue and Leydig cells. We found that there were only Leydig and Sertoli cells in the testes of gastrointestinal tumor patients with azoospermia caused by platinum chemotherapeutic drugs. Melatonin (Mel) receptor 1/melatonin receptor 2 (MT1/MT2) was mainly expressed in human and mouse Leydig cells of the testes. We also observed that the melatonin level in the peripheral blood decreased and oxidative stress occurred in mice treated with cisplatin or gastrointestinal tumor patients treated with platinum-based chemotherapeutic drugs. iTRAQ proteomics showed that SIRT1/Nrf2 signaling and MT1 proteins were downregulated in cisplatin-treated mouse testes. The STRING database predicted that MT1 might be able to regulate the SIRT1/Nrf2 signaling pathway. Melatonin reduced oxidative stress and upregulated SIRT1/Nrf2 signaling in cisplatin-treated mouse testes and Leydig cells. Most importantly, after inhibiting MT1/MT2, melatonin could not upregulate SIRT1/Nrf2 signaling in cisplatin-treated Leydig cells. The MT1/MT2 inhibitor aggravated the cisplatin-induced downregulation of SIRT1/Nrf2 signaling and increased the apoptosis of Leydig cells. We believe that melatonin stimulates SIRT1/Nrf2 signaling by activating MT1/MT2 to prevent the cisplatin-induced apoptosis of Leydig cells.

Therapeutic potential of a single-dose melatonin in the attenuation of cardiac ischemia/reperfusion injury in prediabetic obese rats.

Although acute melatonin treatment effectively reduces cardiac ischemia/reperfusion (I/R) injury in lean rats by modulating melatonin receptor 2 (MT2), there is no information regarding the temporal effects of melatonin administration during cardiac I/R injury in prediabetic obese rats. Prediabetic obese rats induced by chronic consumption of a high-fat diet (HFD) were used. The rats underwent a cardiac I/R surgical procedure (30-min of ischemia, followed by 120-min of reperfusion) and were randomly assigned to receive either vehicle or melatonin treatment. In the melatonin group, rats were divided into 3 different subgroups: (1) pretreatment, (2) treatment during ischemic period, (3) treatment at the reperfusion onset. In the pretreatment subgroup either a nonspecific MT blocker (Luzindole) or specific MT2 blocker (4-PPDOT) was also given to the rats prior to melatonin treatment. Pretreatment with melatonin (10mg/kg) effectively reduced cardiac I/R injury by reducing infarct size, arrhythmia, and LV dysfunction. Reduction in impaired mitochondrial function, mitochondrial dynamic balance, oxidative stress, defective autophagy, and apoptosis were observed in rats pretreated with melatonin. Unfortunately, the cardioprotective benefits were not observed when 10-mg/kg of melatonin was acutely administered to the rats after cardiac ischemia. Thus, we increased the dose of melatonin to 20mg/kg, and it was administered to the rats during ischemia or at the onset of reperfusion. The results showed that 20-mg/kg of melatonin effectively reduced cardiac I/R injury to a similar extent to the 10-mg/kg pretreatment regimen. The MT2 blocker inhibited the protective effects of melatonin. Acute melatonin treatment during cardiac I/R injury exerted protective effects in prediabetic obese rats. However, a higher dose of melatonin is required when given after the onset of cardiac ischemia. These effects of melatonin were mainly mediated through activation of MT2.

Melatonin increases growth properties in human dermal papilla spheroids by activating AKT/GSK3β/β-Catenin signaling pathway.

BackgroundMelatonin, a neurohormone, maybe involved in physiological processes, such as antioxidation, anti-inflammation, and hair growth. In the present study, we investigated the effects of melatonin on proliferation and intracellular signaling in DP cells using a three-dimensional (3D) spheroid culture system that mimics the in vivo hair follicle system.MethodsDP cells were incubated in monolayer (2D) and 3D spheroid culture systems. The expression levels of melatonin receptors in DP cells were analyzed using quantitative reverse transcription polymerase chain reaction (qRT-PCR) and western blotting. The effect of melatonin on the hair-inductive property of DP cells was analyzed using a WST-1-based proliferation assay, determination of DP spheroid size, expression analysis of DP signature genes, and determination of β-catenin stabilization in DP cells. The AKT/GSK3β/β-catenin signaling pathway associated with melatonin-induced β-catenin stabilization in DP cells was investigated by analyzing changes in upstream regulator proteins, including AKT, GSK3β, and their phosphorylated forms.ResultsThe expression levels of the melatonin receptors were higher in human DP cells than in human epidermal keratinocytes and human dermal fibroblast cells. Comparing the expression level according to the human DP cell culture condition, melatonin receptor expression was upregulated in the 3D culture system compared to the traditional two-dimensional monolayer culture system. Cell viability analysis showed that melatonin concentrations up to 1 mM did not affect cell viability. Moreover, melatonin increased the diameter of DP cell 3D spheroids in a dose-dependent manner. Immunoblotting and qRT-PCR analysis revealed that melatonin upregulated the expression of hair growth-related genes, including alkaline phosphatase, bone morphogenetic protein 2, versican, and wingless-int 5A, in a melatonin receptor-dependent manner. Cell fractionation analysis showed that melatonin increased the nuclear localization of β-catenin. This result correlated with the increased transcriptional activation of T-cell factor/lymphoid enhancer factor-responsive luciferase induced by melatonin treatment. Interestingly, melatonin induced the phosphorylation of protein kinase B/AKT at serine 473 residue and GSK-3β at serine 9 residue. To determine whether AKT phosphorylation at serine 473 induced β-catenin nuclear translocation through GSK3β phosphorylation at serine 9, the PI3K/AKT inhibitor LY294002 was cotreated with melatonin. Immunoblotting showed that LY294002 inhibited melatonin-induced phosphorylation of GSK3β at serine 9 residue and β-catenin activation.ConclusionCollectively, this report suggests that melatonin promotes growth properties by activating the AKT/GSK3β/β-catenin signaling pathway through melatonin receptors.

Melatonin alleviates vascular endothelial cell damage by regulating an autophagy-apoptosis axis in Kawasaki disease.

ObjectivesMelatonin has been reported to be an appropriate candidate for mitigating various cardiovascular injuries, owing to its versatility. This study aimed to explore the role of melatonin in Kawasaki disease (KD)‐associated vasculitis and its underlying mechanisms.Material and MethodsThe role of melatonin was evaluated in human coronary artery endothelial cells (HCAECs), peripheral blood mononuclear cells from KD patients, human THP1 cell line in vitro, and a Candida albicans water‐soluble fraction (CAWS)‐induced KD mouse model in vivo. Cell proliferation assay, cell apoptosis assay, cell co‐culture, RNA extraction, RNA sequencing, reverse transcription quantitative PCR, enzyme‐linked immunosorbent assay (ELISA), transwell assay, western blot, dual‐luciferase reporter assay, and autophagic flux assay were performed to investigate the function and regulatory mechanisms of melatonin in vitro, while haematoxylin and eosin staining, Verhoeff's van Gieson staining, ELISA, and immunohistochemical analysis were performed to detect the effect of melatonin in vivo.ResultsMelatonin suppressed cell apoptosis directly reduced the expression of endothelial cell damage markers in HCAECs, and alleviated vasculitis in the CAWS‐induced KD mouse model. Mechanistically, melatonin promoted autophagy by activating the melatonin/ melatonin receptor (MT)/cAMP‐response element binding protein (CREB) pathway and upregulating the expression of autophagy‐related gene‐3, thereby suppressing cell apoptosis in an autophagy‐dependent manner. Additionally, melatonin decreased the production of pro‐inflammatory cytokines in macrophages and indirectly reduced the immunopathological damage of HCAECs.ConclusionsThis study revealed that melatonin protects vascular endothelial cells in KD, by suppressing cell apoptosis in an autophagy‐dependent manner and reducing the immunopathological damage mediated by macrophages.

Melatonin mediates via melatonin receptor 1 in a temperature-dependent manner regulating ATP metabolism and antioxidative enzyme activity of boar spermatozoa in vitro

Researches indicated that melatonin (MLT) could ameliorate oxidative damage and increase sperm motility for conservation sperms, whereas the underlying mechanism remains unknown. And elucidating this, present study was designed to investigate the energy metabolism of sperm throughout preservation at 17 °C and activating with 37 °C. Herein, semen of ten Landrace boars (16∼18 months of age) were diluted and treated with gradient concentrations of MLT (0 nM, 1 nM, 10 nM, 1 μM, 1 mM and 5 mM), and kept away from light in the incubator of 17 °C for 5 days (preservation), and the sperm motility was evaluated after 37 °C incubation for 10 min (activating). The results showed that 1.0 μM MLT treated-sperms had higher motility than other treatments from the 2nd onward. On the 3rd day 17 °C storage, 1.0 μM MLT significantly decreased the ATP content and the mRNA abundance of mitochondrial DNA encoding genes, including NADH dehydrogenase subunits (NDs) 1–4 and 4L, cytochrome b (CYTB), cytochrome c oxidases (COXs) 1 and 3, ATP synthase 6, but increased the hexokinase activity (HK) and the malondialdehyde (MDA) content compared to that of the control. Whereas after 10 min 37 °C incubation, 1.0 μM MLT increased the ATP content, and the mRNA expressions of ND2 and 4L, CYTB, COX1, ATP6, and the activities of mitochondrial respiratory chain complex I and superoxide dismutase (SOD), but depressed the pyruvate kinase (PK) activity in contrast to that at 17 °C. Furthermore, 1.0 μM MLT treated sperms had a higher MLT receptor 1 (MT1) protein level at 17 °C storage than that after 10 min 37 °C incubation, no changes of the MT2 expression were observed at different temperature. This study suggests that MLT might mediate via MT1 in a temperature-dependent manner regulating sperm ATP generation and antioxidative enzyme activity in vitro.