Expression of the MT1 Melatonin Receptor in Ovarian Cancer Cells

Ovarian cancer (OC) is the leading cause of death among women with genital tract disorders. Melatonin exhibits oncostatic properties which it may effect through binding to its membrane receptor, MT1. The aim of this study was to determine the expression of MT1 in OC cells and to correlate this with clinical and pathological data. Immunohistochemistry was performed on 84 cases of OC. Normal ovarian epithelial (IOSE 364) and OC (SK-OV-3, OVCAR-3) cell lines were used to examine the MT1 expression at protein level using the western blot and immunofluorescence technique. The expression of MT1 was observed as cytoplasmic-membrane (MT1_CM) and membrane (MT1_M) reactions. A positive correlation between MT1_CM and MT1_M was found in all the studied cases. There were no significant differences between the expression of MT1_CM, MT1_M, and histological type, staging, grading, presence of residual disease, or overall survival time. Immunofluorescence showed both MT1_M and MT1_CM expression in all the tested cell lines. Western blot illustrated the highest protein level of MT1 in IOSE 364 and the lowest in the OVCAR-3. The results indicate the limited prognostic significance of MT1 in OC cells.     

Melatonin-Induced Postconditioning Suppresses NMDA Receptor through Opening of the Mitochondrial Permeability Transition Pore via Melatonin Receptor in Mouse Neurons

Mitochondrial membrane potential regulation through the mitochondrial permeability transition pore (mPTP) is reportedly involved in the ischemic postconditioning (PostC) phenomenon. Melatonin is an endogenous hormone that regulates circadian rhythms. Its neuroprotective effects via mitochondrial melatonin receptors (MTs) have recently attracted attention. However, details of the neuroprotective mechanisms associated with PostC have not been clarified. Using hippocampal CA1 pyramidal cells from C57BL mice, we studied the involvement of MTs and the mPTP in melatonin-induced PostC mechanisms similar to those of ischemic PostC. We measured changes in spontaneous excitatory postsynaptic currents (sEPSCs), intracellular calcium concentration, mitochondrial membrane potential, and N-methyl-D-aspartate receptor (NMDAR) currents after ischemic challenge, using the whole-cell patch-clamp technique. Melatonin significantly suppressed increases in sEPSCs and intracellular calcium concentrations. The NMDAR currents were significantly suppressed by melatonin and the MT agonist, ramelteon. However, this suppressive effect was abolished by the mPTP inhibitor, cyclosporine A, and the MT antagonist, luzindole. Furthermore, both melatonin and ramelteon potentiated depolarization of mitochondrial membrane potentials, and luzindole suppressed depolarization of mitochondrial membrane potentials. This study suggests that melatonin-induced PostC via MTs suppressed the NMDAR that was induced by partial depolarization of mitochondrial membrane potential by opening the mPTP, reducing excessive release of glutamate and inducing neuroprotection against ischemia-reperfusion injury.     

Therapeutic Effects of Melatonin Receptor Agonists on Sleep and Comorbid Disorders

Several melatonin receptors agonists (ramelteon, prolonged-release melatonin, agomelatine and tasimelteon) have recently become available for the treatment of insomnia, depression and circadian rhythms sleep-wake disorders. The efficacy and safety profiles of these compounds in the treatment of the indicated disorders are reviewed. Accumulating evidence indicates that sleep-wake disorders and co-existing medical conditions are mutually exacerbating. This understanding has now been incorporated into the new Diagnostic and Statistical Manual of Mental Disorders, 5th Edition (DSM-5). Therefore, when evaluating the risk/benefit ratio of sleep drugs, it is pertinent to also evaluate their effects on wake and comorbid condition. Beneficial effects of melatonin receptor agonists on comorbid neurological, psychiatric, cardiovascular and metabolic symptomatology beyond sleep regulation are also described. The review underlines the beneficial value of enhancing physiological sleep in comorbid conditions.     

Transposable Elements and Stress in Vertebrates: An Overview

Since their identification as genomic regulatory elements, Transposable Elements (TEs) were considered, at first, molecular parasites and later as an important source of genetic diversity and regulatory innovations. In vertebrates in particular, TEs have been recognized as playing an important role in major evolutionary transitions and biodiversity. Moreover, in the last decade, a significant number of papers has been published highlighting a correlation between TE activity and exposition to environmental stresses and dietary factors. In this review we present an overview of the impact of TEs in vertebrate genomes, report the silencing mechanisms adopted by host genomes to regulate TE activity, and finally we explore the effects of environmental and dietary factor exposures on TE activity in mammals, which is the most studied group among vertebrates. The studies here reported evidence that several factors can induce changes in the epigenetic status of TEs and silencing mechanisms leading to their activation with consequent effects on the host genome. The study of TE can represent a future challenge for research for developing effective markers able to detect precocious epigenetic changes and prevent human diseases.     

Amyloid Properties of the FXR1 Protein Are Conserved in Evolution of Vertebrates

Functional amyloids are fibrillary proteins with a cross-β structure that play a structural or regulatory role in pro- and eukaryotes. Previously, we have demonstrated that the RNA-binding FXR1 protein functions in an amyloid form in the rat brain. This RNA-binding protein plays an important role in the regulation of long-term memory, emotions, and cancer. Here, we evaluate the amyloid properties of FXR1 in organisms representing various classes of vertebrates. We show the colocalization of FXR1 with amyloid-specific dyes in the neurons of amphibians, reptiles, and birds. Moreover, FXR1, as with other amyloids, forms detergent-resistant insoluble aggregates in all studied animals. The FXR1 protein isolated by immunoprecipitation from the brains of different vertebrate species forms fibrils, which show yellow-green birefringence after staining with Congo red. Our data indicate that in the evolution of vertebrates, FXR1 acquired amyloid properties at least 365 million years ago. Based on the obtained data, we discuss the possible role of FXR1 amyloid fibrils in the regulation of vital processes in the brain of vertebrates.     

Synthesis and SAR Studies of Isoquinoline and Tetrahydroisoquinoline Derivatives as Melatonin Receptor Ligands

In our constant search for new successors of agomelatine, we report herein a new series of compounds resulting from bioisosteric modulation of the naphthalene ring. The isoquinoline and tetrahydroisoquinoline derivatives were synthesized and pharmacologically evaluated. This isosteric replacement of the naphthalene group of agomelatine has led to potent agonist and partial agonist compounds with nanomolar melatonergic binding affinities. Overall, the presence of a nitrogen atom was accompanied with a decrease in the binding affinity toward both MT₁ and MT₂ and the loss of 5HT_2C response, especially for tetrahydroisoquinoline in comparison with the parent compound. Interestingly, due to the presence of this nitrogen atom, a notable improvement in the pharmacokinetic properties was observed for all compounds.     

A Molecular and Chemical Perspective in Defining Melatonin Receptor Subtype Selectivity

Melatonin is primarily synthesized and secreted by the pineal gland during darkness in a normal diurnal cycle. In addition to its intrinsic antioxidant property, the neurohormone has renowned regulatory roles in the control of circadian rhythm and exerts its physiological actions primarily by interacting with the G protein-coupled MT₁ and MT₂ transmembrane receptors. The two melatonin receptor subtypes display identical ligand binding characteristics and mediate a myriad of signaling pathways, including adenylyl cyclase inhibition, phospholipase C stimulation and the regulation of other effector molecules. Both MT₁ and MT₂ receptors are widely expressed in the central nervous system as well as many peripheral tissues, but each receptor subtype can be linked to specific functional responses at the target tissue. Given the broad therapeutic implications of melatonin receptors in chronobiology, immunomodulation, endocrine regulation, reproductive functions and cancer development, drug discovery and development programs have been directed at identifying chemical molecules that bind to the two melatonin receptor subtypes. However, all of the melatoninergics in the market act on both subtypes of melatonin receptors without significant selectivity. To facilitate the design and development of novel therapeutic agents, it is necessary to understand the intrinsic differences between MT₁ and MT₂ that determine ligand binding, functional efficacy, and signaling specificity. This review summarizes our current knowledge in differentiating MT₁ and MT₂ receptors and their signaling capacities. The use of homology modeling in the mapping of the ligand-binding pocket will be described. Identification of conserved and distinct residues will be tremendously useful in the design of highly selective ligands.     

Role of Melatonin in Schizophrenia

Schizophrenia is a chronic mental disease that disturbs several cognitive functions, such as memory, thought, perception and volition. Schizophrenia’s biological etiology is multifactorial and is still under investigation. Melatonin has been involved in schizophrenia since the first decades of the twentieth century. Research into melatonin regarding schizophrenia has followed two different approaches. The first approach is related to the use of melatonin as a biological marker. The second approach deals with the clinical applications of melatonin as a drug treatment. In this paper, both aspects of melatonin application are reviewed. Its clinical use in schizophrenia is emphasized.     

N‐(Anilinoethyl)amides: Design and Synthesis of Metabolically Stable,Selective Melatonin Receptor Ligands

The class of N‐(anilinoethyl)amides includes melatonin receptor ligands with varied subtype selectivity and intrinsic activity. One of these ligands, the MT₂‐selective partial agonist UCM765 (N‐{2‐[(3‐methoxyphenyl)phenylamino]ethyl}acetamide), had evidenced hypnotic effects in rodents at doses ≥40 mg kg^?1 (s.c.), in spite of its sub‐nanomolar affinity for human melatonin receptors. Supposing that its low in vivo potency could be due, at least in part, to metabolic liability in rat liver, UCM765 was incubated with rat liver S9 fraction and rat, mouse, or human microsomes, and the major metabolites were identified by LC–MS, synthesized, and in vitro tested for their affinity toward MT₁ and MT₂ receptors. The obtained information was exploited to design novel analogues of UCM765 that are more resistant to in vitro oxidative degradation, while maintaining a similar binding profile. The analogue UCM924 (N‐{2‐[(3‐bromophenyl)‐(4‐fluorophenyl)amino]ethyl}acetamide) displayed a binding profile similar to that of UCM765 on cloned human receptors (MT₂‐selective partial agonist) and a significantly longer half‐life in the presence of rat liver S9 fraction.     

N-乙酰-5-甲氧基色胺的工艺研究进展

综述了N-乙酰-5-甲氧基色胺主要的合成方法,并对各路线的优缺点进行评价,提出了可行的改进方向.     

Melatonin and Microglia

Melatonin interacts in multiple ways with microglia, both directly and, via routes of crosstalk with astrocytes and neurons, indirectly. These effects of melatonin are of relevance in terms of antioxidative protection, not only concerning free-radical detoxification, but also in prevention of processes that cause, promote, or propagate oxidative stress and neurodegeneration, such as overexcitation, toxicological insults, viral and bacterial infections, and sterile inflammation of different grades. The immunological interplay in the CNS, with microglia playing a central role, is of high complexity and includes signaling toward endothelial cells and other leukocytes by cytokines, chemokines, nitric oxide, and eikosanoids. Melatonin interferes with these processes in multiple signaling routes and steps. In addition to canonical signal transduction by MT₁ and MT₂ melatonin receptors, secondary and tertiary signaling is of relevance and has to be considered, e.g., via the upregulation of sirtuins and the modulation of pro- and anti-inflammatory microRNAs. Many details concerning the modulation of macrophage functionality by melatonin are obviously also applicable to microglial cells. Of particular interest is the polarization toward M2 subtypes instead of M1, i.e., in favor of being anti-inflammatory at the expense of proinflammatory activities, which is well-documented in macrophages but also applies to microglia.     

Mechanisms of Melatonin in Obesity: A Review

Obesity and its complications have become a prominent global public health problem that severely threatens human health. Melatonin, originally known as an effective antioxidant, is an endogenous hormone found throughout the body that serves various physiological functions. In recent decades, increasing attention has been paid to its unique function in regulating energy metabolism, especially in glucose and lipid metabolism. Accumulating evidence has established the relationship between melatonin and obesity; nevertheless, not all preclinical and clinical evidence indicates the anti-obesity effect of melatonin, which makes it remain to conclude the clinical effect of melatonin in the fight against obesity. In this review, we have summarized the current knowledge of melatonin in regulating obesity-related symptoms, with emphasis on its underlying mechanisms. The role of melatonin in regulating the lipid profile, adipose tissue, oxidative stress, and inflammation, as well as the interactions of melatonin with the circadian rhythm, gut microbiota, sleep disorder, as well as the α7nAChR, the opioidergic system, and exosomes, make melatonin a promising agent to open new avenues in the intervention of obesity.     

Nail Melatonin Content: A Suitable Non-Invasive Marker of Melatonin Production

Melatonin plays multiple physiological roles in the human body. Evaluation of melatonin production by the determination of urinary 6-sulfatoxymelatonin in 24-h samples has important drawbacks which hinder the successful evaluation of melatonin production in large cohorts. Here, we evaluated the potential of nail analysis for estimating melatonin production. Firstly, mass spectrometry methodology for the determination of melatonin in nails was optimized and successfully validated. The method was found to be linear in the range 6.5–830 fg/mg with intraday and interday accuracy in the range 100–104 %, precision below 15 % and a LOD of 3.5 fg/mg. Secondly, nail melatonin concentrations from 84 volunteers (age 5–96) were determined. The expected correlation between melatonin and age was obtained (correlation coefficient −0.615; p < 0.001). Additionally, we showed that fingernails are preferable to toenails to determine nail melatonin content. Finally, fingernails collected for 180 days after melatonin administration (two volunteers, 1.9 mg/night during 5 days) were analyzed. Nail melatonin concentrations immediately rose after administration and went back to pre-administration values after ≈100 days in both volunteers. Our results suggest that melatonin determination in nails is a suitable non-invasive tool for the estimation of global melatonin production. Due to the easy collection and storage of nails, the long-term information obtained and the multiple functions of melatonin, nail melatonin content might complement dim light melatonin onset, which is commonly measured from plasma/saliva samples, paving the way for melatonin research.     

松果腺素的合成

以四氢呋喃、4 甲氧基苯胺为主要原料 ,经五步合成了松果腺素。四氢呋喃和氯化氢气体在 1 0 5℃下反应 0 5h ,得到 4 氯丁醇 (Ⅰ) ,产率为 6 8 6 % ;(Ⅰ)用氯铬酸吡啶盐氧化得 4 氯丁醛 (Ⅱ) ,产率为6 0 7% ;4 甲氧基苯胺经重氮化和还原反应得 4 甲氧基苯肼 (Ⅲ) ,产率为 6 0 9% ;(Ⅱ) +(Ⅲ)经环化得 5 甲氧基色胺 (Ⅳ) ,产率为 47 6 % ;(Ⅳ)经乙酰化得松果腺素 (Ⅴ) ,产率为 83 5 %。     

褪黑素的合成研究

通过对褪黑素合成工艺的研究与改进,使氮的烷基化反应中的二溴丙烷消耗降低了３７％;关环反应中１０％氯化氢的乙醇溶液用量减少了３５％;５－甲氧基色胺精制中避免了高成本六甲基二硅胺的使用;乙酰化反应与产品的精制在同一体系中进行,这样既得到了高纯度褪黑素,又简化了工艺操作。使原料总成本降低达２５％左右,生产工艺更适合于工业化生产。产品达到药用标准。     

Melatonin Prevents Chemical-Induced Haemopoietic Cell Death

Melatonin (MEL), a methoxyindole synthesized by the pineal gland, is a powerful antioxidant in tissues as well as within cells, with a fundamental role in ameliorating homeostasis in a number of specific pathologies. It acts both as a direct radical scavenger and by stimulating production/activity of intracellular antioxidant enzymes. In this work, some chemical triggers, with different mechanisms of action, have been chosen to induce cell death in U937 hematopoietic cell line. Cells were pre-treated with 100 μM MEL and then exposed to hydrogen peroxide or staurosporine. Morphological analyses, TUNEL reaction and Orange/PI double staining have been used to recognize ultrastructural apoptotic patterns and to evaluate DNA behavior. Chemical damage and potential MEL anti-apoptotic effects were quantified by means of Tali^® Image-Based Cytometer, able to monitor cell viability and apoptotic events. After trigger exposure, chromatin condensation, micronuclei formation and DNA fragmentation have been observed, all suggesting apoptotic cell death. These events underwent a statistically significant decrease in samples pre-treated with MEL. After caspase inhibition and subsequent assessment of cell viability, we demonstrated that apoptosis occurs, at least in part, through the mitochondrial pathway and that MEL interacts at this level to rescue U937 cells from death.     

一种简便的松果腺素的合成方法

以邻苯二甲酰亚胺钾、１,３二溴丙烷、对甲氧基苯胺为主要原料,经５步化学反应合成了松果腺素：邻苯二甲酰亚胺钾和１,３－二溴丙烷在１５５～１６０℃下反应６ｈ,得到Ｎ－（３－溴丙基）,邻苯二甲酸亚胺（Ⅱ）,产率７７％;Ⅱ在乙醇钠存在下和乙酰乙酸乙酯作用得到２－乙酸基－５－邻苯二甲酰亚氨基戊酸乙酯（Ⅲ）,产率７２％;Ⅲ与对甲氧基苯胺重氮盐偶联后发生环化作用,生成Ｔ２－羧乙基－３－（２－邻苯二甲酰亚氨乙基）－５－甲氧基吲哚（Ⅳ）,产率７４％;Ⅳ经碱皂化、水解、脱羧反萃取纯化,得到５．甲氧基色胺（Ｖ）,产率６６％;Ｖ经乙酸化后得到松果腺素（Ⅵ）,产率８２％。     

褪黑素的化学合成方法

根据褪黑素的结构特点,综述了其不同的化学合成路线,并结合我国国情,提出了比较可行的3种褪黑素合成方法。