Showing papers in "Journal of Pineal Research in 2015"

Melatonin: an ancient molecule that makes oxygen metabolically tolerable.

Abstract: Melatonin is remarkably functionally diverse with actions as a free radical scavenger and antioxidant, circadian rhythm regulator, anti-inflammatory and immunoregulating molecule, and as an oncostatic agent. We hypothesize that the initial and primary function of melatonin in photosynthetic cyanobacteria, which appeared on Earth 3.5-3.2 billion years ago, was as an antioxidant. The evolution of melatonin as an antioxidant by this organism was necessary as photosynthesis is associated with the generation of toxic-free radicals. The other secondary functions of melatonin came about much later in evolution. We also surmise that mitochondria and chloroplasts may be primary sites of melatonin synthesis in all eukaryotic cells that possess these organelles. This prediction is made on the basis that mitochondria and chloroplasts of eukaryotes developed from purple nonsulfur bacteria (which also produce melatonin) and cyanobacteria when they were engulfed by early eukaryotes. Thus, we speculate that the melatonin-synthesizing actions of the engulfed bacteria were retained when these organelles became mitochondria and chloroplasts, respectively. That mitochondria are likely sites of melatonin formation is supported by the observation that this organelle contains high levels of melatonin that are not impacted by blood melatonin concentrations. Melatonin has a remarkable array of means by which it thwarts oxidative damage. It, as well as its metabolites, is differentially effective in scavenging a variety of reactive oxygen and reactive nitrogen species. Moreover, melatonin and its metabolites modulate a large number of antioxidative and pro-oxidative enzymes, leading to a reduction in oxidative damage. The actions of melatonin on radical metabolizing/producing enzymes may be mediated by the Keap1-Nrf2-ARE pathway. Beyond its direct free radical scavenging and indirect antioxidant effects, melatonin has a variety of physiological and metabolic advantages that may enhance its ability to limit oxidative stress.

Functions of melatonin in plants: a review

Abstract: The number of studies on melatonin in plants has increased significantly in recent years. This molecule, with a large set of functions in animals, has also shown great potential in plant physiology. This review outlines the main functions of melatonin in the physiology of higher plants. Its role as antistress agent against abiotic stressors, such as drought, salinity, low and high ambient temperatures, UV radiation and toxic chemicals, is analyzed. The latest data on their role in plant-pathogen interactions are also discussed. Both abiotic and biotic stresses produce a significant increase in endogenous melatonin levels, indicating its possible role as effector in these situations. The existence of endogenous circadian rhythms in melatonin levels has been demonstrated in some species, and the data, although limited, suggest a central role of this molecule in the day/night cycles in plants. Finally, another aspect that has led to a large volume of research is the involvement of melatonin in aspects of plant development regulation. Although its role as a plant hormone is still far of from being fully established, its involvement in processes such as growth, rhizogenesis, and photosynthesis seems evident. The multiple changes in gene expression caused by melatonin point to its role as a multiregulatory molecule capable of coordinating many aspects of plant development. This last aspect, together with its role as an alleviating-stressor agent, suggests that melatonin is an excellent prospect for crop improvement.

Melatonin and endoplasmic reticulum stress: relation to autophagy and apoptosis.

Abstract: Endoplasmic reticulum (ER) is a dynamic organelle that participates in a number of cellular functions by controlling lipid metabolism, calcium stores, and proteostasis Under stressful situations, the ER environment is compromised, and protein maturation is impaired; this causes misfolded proteins to accumulate and a characteristic stress response named unfolded protein response (UPR) UPR protects cells from stress and contributes to cellular homeostasis re-establishment; however, during prolonged ER stress, UPR activation promotes cell death ER stressors can modulate autophagy which in turn, depending of the situation, induces cell survival or death Interactions of different autophagy- and apoptosis-related proteins and also common signaling pathways have been found, suggesting an interplay between these cellular processes, although their dynamic features are still unknown A number of pathologies including metabolic, neurodegenerative and cardiovascular diseases, cancer, inflammation, and viral infections are associated with ER stress, leading to a growing interest in targeting components of the UPR as a therapeutic strategy Melatonin has a variety of antioxidant, anti-inflammatory, and antitumor effects As such, it modulates apoptosis and autophagy in cancer cells, neurodegeneration and the development of liver diseases as well as other pathologies Here, we review the effects of melatonin on the main ER stress mechanisms, focusing on its ability to regulate the autophagic and apoptotic processes As the number of studies that have analyzed ER stress modulation by this indole remains limited, further research is necessary for a better understanding of the crosstalk between ER stress, autophagy, and apoptosis and to clearly delineate the mechanisms by which melatonin modulates these responses

Melatonin delays leaf senescence and enhances salt stress tolerance in rice.

Abstract: Melatonin, an antioxidant in both animals and plants, has been reported to have beneficial effects on the aging process. It was also suggested to play a role in extending longevity and enhancing abiotic stress resistance in plant. In this study, we demonstrate that melatonin acts as a potent agent to delay leaf senescence and cell death in rice. Treatments with melatonin significantly reduced chlorophyll degradation, suppressed the transcripts of senescence-associated genes, delayed the leaf senescence, and enhanced salt stress tolerance. Genome-wide expression profiling by RNA sequencing reveals that melatonin is a potent free radical scavenger, and its exogenous application results in enhanced antioxidant protection. Leaf cell death in noe1, a mutant with over-produced H2O2, can be relieved by exogenous application of melatonin. These data demonstrate that melatonin delays the leaf senescence and cell death and also enhances abiotic stress tolerance via directly or indirectly counteracting the cellular accumulation of H2O2.

Melatonin attenuates D-galactose-induced memory impairment, neuroinflammation and neurodegeneration via RAGE/NF-K B/JNK signaling pathway in aging mouse model.

Abstract: Melatonin acts as a pleiotropic agent in various age-related neurodegenerative diseases. In this study, we examined the underlying neuroprotective mechanism of melatonin against D-galactose-induced memory and synaptic dysfunction, elevated reactive oxygen species (ROS), neuroinflammation and neurodegeneration. D-galactose was administered (100 mg/kg intraperitoneally (i.p.)) for 60 days. After 30 days of D-galactose administration, vehicle (same volume) or melatonin (10 mg/kg, i.p.) was administered for 30 days. Our behavioral (Morris water maze and Y-maze test) results revealed that chronic melatonin treatment alleviated D-galactose-induced memory impairment. Additionally, melatonin treatment reversed D-galactose-induced synaptic disorder via increasing the level of memory-related pre-and postsynaptic protein markers. We also determined that melatonin enhances memory function in the D-galactose-treated mice possibly via reduction of elevated ROS and receptor for advanced glycation end products (RAGE). Furthermore, Western blot and morphological results showed that melatonin treatment significantly reduced D-galactose-induced neuroinflammation through inhibition of microgliosis (Iba-1) and astrocytosis (GFAP), and downregulating other inflammatory mediators such as p-IKKβ, p-NF-K B65, COX2, NOS2, IL-1β, and TNFα. Moreover, melatonin lowered the oxidative stress kinase p-JNK which suppressed various apoptotic markers, that is, cytochrome C, caspase-9, caspase-3 and PARP-1, and prevent neurodegeneration. Hence, melatonin attenuated the D-galactose-induced memory impairment, neuroinflammation and neurodegeneration possibly through RAGE/NF-K B/JNK pathway. Taken together, our data suggest that melatonin could be a promising, safe and endogenous compatible antioxidant candidate for age-related neurodegenerative diseases such as Alzheimer's disease (AD).

Melatonin prevents cell death and mitochondrial dysfunction via a SIRT1-dependent mechanism during ischemic-stroke in mice

Abstract: Silent information regulator 1 (SIRT1), a type of histone deacetylase, is a highly effective therapeutic target for protection against ischemia reperfusion (IR) injury (IRI). Previous studies showed that melatonin preserves SIRT1 expression in neuronal cells of newborn rats after hypoxia-ischemia. However, the definite role of SIRT1 in the protective effect of melatonin against cerebral IRI in adult has not been explored. In this study, the brain of adult mice was subjected to IRI. Prior to this procedure, the mice were given intraperitoneal with or without the SIRT1 inhibitor, EX527. Melatonin conferred a cerebral-protective effect, as shown by reduced infarct volume, lowered brain edema, and increased neurological scores. The melatonin-induced upregulation of SIRT1 was also associated with an increase in the anti-apoptotic factor, Bcl2, and a reduction in the pro-apoptotic factor Bax. Moreover, melatonin resulted in a well-preserved mitochondrial membrane potential, mitochondrial Complex I activity, and mitochondrial cytochrome c level while it reduced cytosolic cytochrome c level. However, the melatonin-elevated mitochondrial function was reversed by EX527 treatment. In summary, our results demonstrate that melatonin treatment attenuates cerebral IRI by reducing IR-induced mitochondrial dysfunction through the activation of SIRT1 signaling.

Melatonin induces nitric oxide and the potential mechanisms relate to innate immunity against bacterial pathogen infection in Arabidopsis

Abstract: Melatonin (N-acetyl-5-methoxytryptamine) is a naturally occurring small molecule, serving as important secondary messenger in the response of plants to various biotic and abiotic stresses. However, the interactions between melatonin and other important molecules in the plant stress response, especially in plant immunity, are largely unknown. In this study, we found that both melatonin and nitric oxide (NO) levels in Arabidopsis leaves were significantly induced by bacterial pathogen (Pst DC3000) infection. The elevated NO production was caused by melatonin as melatonin application enhanced endogenous NO level with great efficacy. Moreover, both melatonin and NO conferred improved disease resistance against Pseudomonas syringe pv. tomato (Pst) DC3000 infection in Arabidopsis. NO scavenger significantly suppressed the rise of NO which was induced by exogenous application of melatonin. As a result, the beneficial effects of melatonin on the expression of salicylic acid (SA)-related genes and disease resistance against bacterial pathogen infection were jeopardized by use of a NO scavenger. Consistently, melatonin application significantly lost its effect on the innate immunity against P. syringe pv. tomato (Pst) DC3000 infection in NO-deficient mutants of Arabidopsis. The results indicate that melatonin-induced NO production is responsible for innate immunity response of Arabidopsis against Pst DC3000 infection.

Melatonin feedback on clock genes: a theory involving the proteasome.

Abstract: The expression of 'clock' genes occurs in all tissues, but especially in the suprachiasmatic nuclei (SCN) of the hypothalamus, groups of neurons in the brain that regulate circadian rhythms. Melatonin is secreted by the pineal gland in a circadian manner as influenced by the SCN. There is also considerable evidence that melatonin, in turn, acts on the SCN directly influencing the circadian 'clock' mechanisms. The most direct route by which melatonin could reach the SCN would be via the cerebrospinal fluid of the third ventricle. Melatonin could also reach the pars tuberalis (PT) of the pituitary, another melatonin-sensitive tissue, via this route. The major 'clock' genes include the period genes, Per1 and Per2, the cryptochrome genes, Cry1 and Cry2, the clock (circadian locomotor output cycles kaput) gene, and the Bmal1 (aryl hydrocarbon receptor nuclear translocator-like) gene. Clock and Bmal1 heterodimers act on E-box components of the promoters of the Per and Cry genes to stimulate transcription. A negative feedback loop between the cryptochrome proteins and the nucleus allows the Cry and Per proteins to regulate their own transcription. A cycle of ubiquitination and deubiquitination controls the levels of CRY protein degraded by the proteasome and, hence, the amount of protein available for feedback. Thus, it provides a post-translational component to the circadian clock mechanism. BMAL1 also stimulates transcription of REV-ERBα and, in turn, is also partially regulated by negative feedback by REV-ERBα. In the 'black widow' model of transcription, proteasomes destroy transcription factors that are needed only for a particular period of time. In the model proposed herein, the interaction of melatonin and the proteasome is required to adjust the SCN clock to changes in the environmental photoperiod. In particular, we predict that melatonin inhibition of the proteasome interferes with negative feedback loops (CRY/PER and REV-ERBα) on Bmal1 transcription genes in both the SCN and PT. Melatonin inhibition of the proteasome would also tend to stabilize BMAL1 protein itself in the SCN, particularly at night when melatonin is naturally elevated. Melatonin inhibition of the proteasome could account for the effects of melatonin on circadian rhythms associated with molecular timing genes. The interaction of melatonin with the proteasome in the hypothalamus also provides a model for explaining the dramatic 'time of day' effect of melatonin injections on reproductive status of seasonal breeders. Finally, the model predicts that a proteasome inhibitor such as bortezomib would modify circadian rhythms in a manner similar to melatonin.

Melatonin alleviates brain injury in mice subjected to cecal ligation and puncture via attenuating inflammation, apoptosis, and oxidative stress: the role of SIRT1 signaling.

Abstract: Sepsis is a systemic inflammatory response to infection that causes severe neurological complications. Previous studies have suggested that melatonin is protective during sepsis. Additionally, silent information regulator 1 (SIRT1) was reported to be beneficial in sepsis. However, the role of SIRT1 signaling in the protective effect of melatonin against septic encephalopathy remains unclear. This study aimed to investigate the role of SIRT1 in the protective effect of melatonin. EX527, a SIRT1 inhibitor, was used to reveal the role of SIRT1 in melatonin's action. Cecal ligation and puncture or sham operation was performed in male C57BL/6J mice. Melatonin was administrated intraperitoneally (30 mg/kg). The survival rate of mice was recorded for the 7-day period following the sham or CLP operation. The blood-brain barrier (BBB) integrity, brain water content, levels of inflammatory cytokines (TNF-α, IL-1β, and HMGB1), and the level of oxidative stress (superoxide dismutase (SOD), catalase (CAT), and malondialdehyde (MDA)) and apoptosis were assessed. The expression of SIRT1, Ac-FoxO1, Ac-p53, Ac-NF-κB, Bcl-2, and Bax was detected by Western blot. The results suggested that melatonin improved survival rate, attenuated brain edema and neuronal apoptosis, and preserved BBB integrity. Melatonin decreased the production of TNF-α, IL-1β, and HMGB1. Melatonin increased the activity of SOD and CAT and decreased the MDA production. Additionally, melatonin upregulated the expression of SIRT1 and Bcl-2 and downregulated the expression of Ac-FoxO1, Ac-p53, Ac-NF-κB, and Bax. However, the protective effects of melatonin were abolished by EX527. In conclusion, our results demonstrate that melatonin attenuates sepsis-induced brain injury via SIRT1 signaling activation.

Arabidopsis serotonin N-acetyltransferase knockout mutant plants exhibit decreased melatonin and salicylic acid levels resulting in susceptibility to an avirulent pathogen.

Abstract: Serotonin N-acetyltransferase (SNAT) is the penultimate enzyme in the melatonin biosynthesis pathway in plants. We examined the effects of SNAT gene inactivation in two Arabidopsis T-DNA insertion mutant lines. After inoculation with the avirulent pathogen Pseudomonas syringe pv. tomato DC3000 harboring the elicitor avrRpt2 (Pst-avrRpt2), melatonin levels in the snat knockout mutant lines were 50% less than in wild-type Arabidopsis Col-0 plants. The snat knockout mutant lines exhibited susceptibility to pathogen infection that coincided with decreased induction of defense genes including PR1, ICS1, and PDF1.2. Because melatonin acts upstream of salicylic acid (SA) synthesis, the reduced melatonin levels in the snat mutant lines led to decreased SA levels compared to wild-type, suggesting that the increased pathogen susceptibility of the snat mutant lines could be attributed to decreased SA levels and subsequent attenuation of defense gene induction. Exogenous melatonin treatment failed to induce defense gene expression in nahG Arabidopsis plants, but restored the induction of defense gene expression in the snat mutant lines. In addition, melatonin caused translocation of NPR1 (nonexpressor of PR1) protein from the cytoplasm into the nucleus indicating that melatonin-elicited pathogen resistance in response to avirulent pathogen attack is SA-dependent in Arabidopsis.

Melatonin induces class A1 heat-shock factors (HSFA1s) and their possible involvement of thermotolerance in Arabidopsis

Abstract: Melatonin (N-acetyl-5-methoxytryptamine) serves as an important signal molecule during plant developmental processes and multiple abiotic stress responses. However, the involvement of melatonin in thermotolerance and the underlying molecular mechanism in Arabidopsis were largely unknown. In this study, we found that the endogenous melatonin level in Arabidopsis leaves was significantly induced by heat stress treatment, and exogenous melatonin treatment conferred improved thermotolerance in Arabidopsis. The transcript levels of class A1 heat-shock factors (HSFA1s), which serve as the master regulators of heat stress responses, were significantly upregulated by heat stress and exogenous melatonin treatment in Arabidopsis. Notably, exogenous melatonin-enhanced thermotolerance was largely alleviated in HSFA1s quadruple knockout (QK) mutants, and HSFA1s-activated transcripts of heat-responsive genes (HSFA2, heat stress-associated 32 (HSA32), heat-shock protein 90 (HSP90), and 101 (HSP101)) might be contributed to melatonin-mediated thermotolerance. Taken together, this study provided direct link between melatonin and thermotolerance and indicated the involvement of HSFA1s-activated heat-responsive genes in melatonin-mediated thermotolerance in Arabidopsis.

Melatonin ameliorates amyloid beta-induced memory deficits, tau hyperphosphorylation and neurodegeneration via PI3/Akt/GSk3β pathway in the mouse hippocampus.

Abstract: Alzheimer's disease (AD) is the most prevalent age-related neurodegenerative disease, pathologically characterized by the accumulation of amyloid beta (Aβ) aggregation in the brain, and is considered to be the primary cause of cognitive dysfunction. Aβ aggregates lead to synaptic disorder, tau hyperphosphorylation, and neurodegeneration. In this study, the underlying neuroprotective mechanism of melatonin against Aβ1-42-induced neurotoxicity was investigated in the mice hippocampus. Intracerebroventricular (i.c.v.) Aβ1-42-injection triggered memory impairment, synaptic disorder, hyperphosphorylation of tau protein, and neurodegeneration in the mice hippocampus. After 24 hr of Aβ1-42 injection, the mice were treated with melatonin (10 mg/kg, intraperitonially) for 3 wks, reversed the Aβ1-42-induced synaptic disorder via increasing the level of presyanptic (Synaptophysin and SNAP-25) and postsynaptic protein [PSD95, p-GluR1 (Ser845), SNAP23, and p-CREB (Ser133)], respectively, and attenuated the Aβ1-42-induced memory impairment. Chronic melatonin treatment attenuated the hyperphosphorylation of tau protein via PI3K/Akt/GSK3β signaling by activating the p-PI3K, p-Akt (Ser 473) and p-GSK3β (Ser9) in the Aβ1-42-treated mice. Furthermore, melatonin decreased Aβ1-42 -induced apoptosis through decreasing the overexpression of caspase-9, caspase-3, and PARP-1 level. Additionally, the evaluation of immunohistochemical analysis of caspase-3, Fluorojade-B, and Nissl staining indicated that melatonin prevented neurodegeneration in Aβ1-42-treated mice. Our results demonstrated that melatonin has neuroprotective effect against Aβ1-42-induced neurotoxicity through decreasing memory impairment, synaptic disorder, tau hyperphosphorylation, and neurodegeneration via PI3K/Akt/GSK3β signaling in the Aβ1-42-treated mouse model of AD. On the basis of these results, we suggest that melatonin could be an effective, promising, and safe neuroprotective candidate for the treatment of progressive neurodegenerative disorders, such as AD.

INDOLE‐3‐ACETIC ACID INDUCIBLE 17 positively modulates natural leaf senescence through melatonin‐mediated pathway in Arabidopsis

Abstract: Melatonin (N-acetyl-5-methoxytryptamine) functions as a ubiquitous modulator in multiple plant developmental processes and various stress responses. However, the involvement of melatonin in natural leaf senescence and the underlying molecular mechanism in Arabidopsis remain unclear. In this study, we found that the endogenous melatonin level was significantly induced in a developmental stage-dependent manner, and exogenous melatonin treatment delayed natural leaf senescence in Arabidopsis. The expression level of AUXIN RESISTANT 3 (AXR3)/INDOLE-3-ACETIC ACID INDUCIBLE 17 (IAA17) was significantly downregulated by exogenous melatonin treatment and decreased with developmental age in Arabidopsis. Further investigation indicated that AtIAA17-overexpressing plants showed early leaf senescence with lower chlorophyll content in rosette leaves compared with wild-type plants, while AtIAA17 knockout mutants displayed delayed leaf senescence with higher chlorophyll content. Notably, exogenous melatonin-delayed leaf senescence was largely alleviated in AtIAA17-overexpressing plants, and AtIAA17-activated senescence-related SENESCENCE 4 (SEN4) and SENESCENCE-ASSOCIATED GENE 12 (SAG12) transcripts might have contributed to the process of natural leaf senescence. Taken together, the results indicate that AtIAA17 is a positive modulator of natural leaf senescence and provides direct link between melatonin and AtIAA17 in the process of natural leaf senescence in Arabidopsis.

Melatonin and its metabolites as copper chelating agents and their role in inhibiting oxidative stress: a physicochemical analysis.

Abstract: The copper sequestering ability of melatonin and its metabolites cyclic 3-hydroxymelatonin (3OHM), N(1) -acetyl-N(2) -formyl-5-methoxykynuramine (AFMK), and N(1) -acetyl-5-methoxykynuramine (AMK) was investigated within the frame of the Density Functional Theory. It was demonstrated that these compounds are capable of chelating copper ions, yielding stable complexes. The most likely chelation sites were identified. Two different mechanisms were modeled, the direct-chelation mechanism (DCM) and the coupled-deprotonation-chelation mechanism (CDCM). It is proposed that, under physiological conditions, CDCM would be the main chelation route for Cu(II). It was found that melatonin and its metabolites fully inhibited the oxidative stress induced by Cu(II)-ascorbate mixtures, via Cu(II) chelation. In the same way, melatonin, AFMK, and 3OHM also prevented the first step of the Haber-Weiss reaction, consequently turning off the ˙OH production via the Fenton reaction. Therefore, it is proposed that, in addition to the previously reported free radical scavenging cascade, melatonin is also involved in a concurrent 'chelating cascade', thereby contributing to a reduction in oxidative stress. 3OHM was identified as the most efficient of the studied compounds for that purpose, supporting the important role of this metabolite in the beneficial effects of melatonin against oxidative stress.

Anatomical and cellular localization of melatonin MT1 and MT2 receptors in the adult rat brain

Abstract: The involvement of melatonin in mammalian brain pathophysiology has received growing interest, but information about the anatomical distribution of its two G-protein-coupled receptors, MT1 and MT2 , remains elusive. In this study, using specific antibodies, we examined the precise distribution of both melatonin receptors immunoreactivity across the adult rat brain using light, confocal, and electron microscopy. Our results demonstrate a selective MT1 and MT2 localization on neuronal cell bodies and dendrites in numerous regions of the rat telencephalon, diencephalon, and mesencephalon. Confocal and ultrastructural examination confirmed the somatodendritic nature of MT1 and MT2 receptors, both being localized on neuronal membranes. Overall, striking differences were observed in the anatomical distribution pattern of MT1 and MT2 proteins, and the labeling often appeared complementary in regions displaying both receptors. Somadendrites labeled for MT1 were observed for instance in the retrosplenial cortex, the dentate gyrus of the hippocampus, the islands of Calleja, the medial habenula, the suprachiasmatic nucleus, the superior colliculus, the substantia nigra pars compacta, the dorsal raphe nucleus, and the pars tuberalis of the pituitary gland. Somadendrites endowed with MT2 receptors were mostly observed in the CA3 field of the hippocampus, the reticular thalamic nucleus, the supraoptic nucleus, the inferior colliculus, the substantia nigra pars reticulata, and the ventrolateral periaqueductal gray. Together, these data provide the first detailed neurocytological mapping of melatonin receptors in the adult rat brain, an essential prerequisite for a better understanding of melatonin distinct receptor function and neurophysiology.

Melatonin induces the transcripts of CBF/DREB1s and their involvement in both abiotic and biotic stresses in Arabidopsis.

Abstract: Melatonin (N-acetyl-5-methoxytryptamine) is a naturally occurring small molecule that acts as an important secondary messenger in plant stress responses. However, the mechanism underlying the melatonin-mediated signaling pathway in plant stress responses has not been established. C-repeat-binding factors (CBFs)/Drought response element Binding 1 factors (DREB1s) encode transcription factors that play important roles in plant stress responses. This study has determined that endogenous melatonin and transcripts level of CBFs (AtCBF1, AtCBF2, and AtCBF3) in Arabidopsis leaves were significantly induced by salt, drought, and cold stresses and by pathogen Pseudomonas syringe pv. tomato (Pst) DC3000 infection. Moreover, both exogenous melatonin treatment and overexpression of CBFs conferred enhanced resistance to both abiotic and biotic stresses in Arabidopsis. Notably, AtCBFs and exogenous melatonin treatment positively regulated the mRNA expression of several stress-responsive genes (COR15A, RD22, and KIN1) and accumulation of soluble sugars content such as sucrose in Arabidopsis under control and stress conditions. Additionally, exogenous sucrose also conferred improved resistance to both abiotic and biotic stresses in Arabidopsis. Taken together, this study indicates that AtCBFs confer enhanced resistance to both abiotic and biotic stresses, and AtCBF-mediated signaling pathway and sugar accumulation may be involved in melatonin-mediated stress response in Arabidopsis, at least partially.

Melatonin uptake through glucose transporters: a new target for melatonin inhibition of cancer.

Abstract: Melatonin is present in a multitude of taxa and it has a broad range of biological functions, from synchronizing circadian rhythms to detoxifying free radicals. Some functions of melatonin are mediated by its membrane receptors but others are receptor-independent. For the latter, melatonin must enter into the cell. Melatonin is a derivative of the amino acid tryptophan and reportedly easily crosses biological membranes due to its amphipathic nature. However, the mechanism by which melatonin enters into cells remains unknown. Changes in redox state, endocytosis pathways, multidrug resistance, glycoproteins or a variety of strategies have no effect on melatonin uptake. Herein, it is demonstrated that members of the SLC2/GLUT family glucose transporters have a central role in melatonin uptake. When studied by docking simulation, it is found that melatonin interacts at the same location in GLUT1 where glucose does. Furthermore, glucose concentration and the presence of competitive ligands of GLUT1 affect the concentration of melatonin into cells. As a regulatory mechanism, melatonin reduces the uptake of glucose and modifies the expression of GLUT1 transporter in prostate cancer cells. More importantly, glucose supplementation promotes prostate cancer progression in TRAMP mice, while melatonin attenuated glucose-induced tumor progression and prolonged the lifespan of tumor-bearing mice. This is the first time that a facilitated transport of melatonin is suggested. In fact, the important role of glucose transporters and glucose metabolism in cell fate might explain some of the diverse functions described for melatonin.

Inter‐relationships of the chronobiotic, melatonin, with leptin and adiponectin: implications for obesity

Abstract: Obesity and its medical complications represent a significant problem throughout the world. In recent decades, mechanisms underlying the progression of obesity have been intensively examined. The involvement of both the behavioral aspects, such as calorie-rich diet, low physical activity and sleep deprivation, and the intrinsic factors, including adipose tissue deregulation, chronic inflammation, oxidative stress, and chronodisruption, has been identified. The circadian disturbances of the adipose tissue endocrine function have been correlated with obesity. Leptin and adiponectin are adipokines strongly associated with glucose and lipid metabolism and with energy balance. Their synthesis and secretion display circadian rhythms that are disturbed in the obese state. Hyperleptinemia resulting in leptin resistance, and hypo-adiponectinemia have been linked to the pathophysiology of the obesity-related disorders. A deficiency of melatonin, one of the consequences of sleep deprivation, has also been demonstrated to correlate with obesity. Melatonin is a pineal secretory product involved in numerous actions, such as regulation of internal biological clocks and energy metabolism, and it functions as an antioxidant and as an anti-inflammatory agent. There exists a substantial amount of evidence supporting the beneficial effects of melatonin supplementation on obesity and its complications. In the current review, the results of studies related to the interactions between melatonin, and both leptin and adiponectin are discussed. Despite the existence of some inconsistencies, melatonin has been found to normalize the expression and secretion patterns of both adipokines. These results support the concept of melatonin as a potential therapeutic agent for obesity and related disorders.

Coordinated regulation of melatonin synthesis and degradation genes in rice leaves in response to cadmium treatment

Abstract: We investigated the expression patterns of genes involved in melatonin synthesis and degradation in rice leaves upon cadmium (Cd) treatment and the subcellular localization sites of melatonin 2-hydroxylase (M2H) proteins. The Cd-induced synthesis of melatonin coincided with the increased expression of melatonin biosynthetic genes including tryptophan decarboxylase (TDC), tryptamine 5-hydroxylase (T5H), and N-acetylserotonin methyltransferase (ASMT). However, the expression of serotonin N-acetyltransferase (SNAT), the penultimate gene in melatonin biosynthesis, was downregulated, suggesting that melatonin synthesis was counter-regulated by SNAT. Notably, the induction of melatonin biosynthetic gene expression was coupled with the induction of four M2H genes involved in melatonin degradation, which suggests that genes for melatonin synthesis and degradation are coordinately regulated. The induced M2H gene expression was correlated with enhanced M2H enzyme activity. Three of the M2H proteins were localized to the cytoplasm and one M2H protein was localized to chloroplasts, indicating that melatonin degradation occurs both in the cytoplasm and in chloroplasts. The biological activity of 2-hydroxymelatonin in the induction of the plant defense gene expression was 50% less than that of melatonin, which indicates that 2-hydroxymelatonin may be a metabolite of melatonin. Overall, our data demonstrate that melatonin synthesis occurs in parallel with melatonin degradation in both chloroplasts and cytoplasm, and the resulting melatonin metabolite 2-hydroxymelatonin also acts as a signaling molecule for defense gene induction.

Melatonin reverses H2 O2 -induced premature senescence in mesenchymal stem cells via the SIRT1-dependent pathway.

Abstract: Mesenchymal stem cells (MSCs) represent an attractive source for stem cell-based regenerative therapy, but they are vulnerable to oxidative stress-induced premature senescence in pathological conditions. We previously reported antioxidant and antiarthritic effects of melatonin on MSCs against proinflammatory cytokines. In this study, we hypothesized that melatonin could protect MSCs from premature senescence induced by hydrogen peroxide (H2 O2 ) via the silent information regulator type 1 (SIRT1)-dependent pathway. In response to H2 O2 at a sublethal concentration of 200 μm, human bone marrow-derived MSCs (BM-MSCs) underwent growth arrest and cellular senescence. Treatment with melatonin before H2 O2 exposure cannot significantly prevent premature senescence; however, treatment with melatonin subsequent to H2 O2 exposure successfully reversed the senescent phenotypes of BM-MSCs in a dose-dependent manner. This result was made evident by improved cell proliferation, decreased senescence-associated β-galactosidase activity, and the improved entry of proliferating cells into the S phase. In addition, treatment with 100 μm melatonin restored the osteogenic differentiation potential of BM-MSCs that was inhibited by H2 O2 -induced premature senescence. We also found that melatonin attenuated the H2 O2 -stimulated phosphorylation of p38 mitogen-activated protein kinase, decreased expression of the senescence-associated protein p16(INK) (4α) , and increased SIRT1. Further molecular experiments revealed that luzindole, a nonselective antagonist of melatonin receptors, blocked melatonin-mediated antisenescence effects. Inhibition of SIRT1 by sirtinol counteracted the protective effects of melatonin, suggesting that melatonin reversed the senescence in cells through the SIRT1-dependent pathway. Together, these findings lay new ground for understanding oxidative stress-induced premature senescence and open perspectives for therapeutic applications of melatonin in stem cell-based regenerative medicine.

Melatonin blunts the mitochondrial/NLRP3 connection and protects against radiation-induced oral mucositis

Abstract: Mucositis is a common and distressing side effect of chemotherapy or radiotherapy that has potentially severe consequences, and no treatment is available. The purpose of this study was to analyze the molecular pathways involved in the development of oral mucositis and to evaluate whether melatonin can prevent this pathology. The tongue of male Wistar rats was subjected to irradiation (X-ray YXLON Y.Tu 320-D03 irradiator; the animals received a dose of 7.5 Gy/day for 5 days). Rats were treated with 45 mg/day melatonin or vehicle for 21 days postirradiation, either by local application into their mouths (melatonin gel) or by subcutaneous injection. A connection between reactive oxygen species, generating mitochondria and the NLRP3 (NLR-related protein 3 nucleotide-binding domain leucine-rich repeat containing receptor-related protein 3) inflammasome, has been reported in mucositis. Here, we show that mitochondrial oxidative stress, bioenergetic impairment and subsequent NLRP3 inflammasome activation are involved in the development of oral mucositis after irradiation and that melatonin synthesized in the rat tongue is depleted after irradiation. The application of melatonin gel restores physiological melatonin levels in the tongue and prevents mucosal disruption and ulcer formation. Melatonin gel protects the mitochondria from radiation damage and blunts the NF-κB/NLRP3 inflammasome signaling activation in the tongue. Our results suggest new molecular pathways involved in radiotherapy-induced mucositis that are inhibited by topical melatonin application, suggesting a potential preventive therapy for mucositis in patients with cancer.

Comparative physiological and proteomic analyses reveal the actions of melatonin in the reduction of oxidative stress in Bermuda grass (Cynodon dactylon (L). Pers.)

Abstract: The fact of melatonin as an important antioxidant in animals led plant researchers to speculate that melatonin also acts in the similar manner in plants. Although melatonin has significant effects on alleviating stress-triggered reactive oxygen species (ROS), the involvement of melatonin in direct oxidative stress and the underlying physiological and molecular mechanisms remain unclear in plants. In this study, we found that exogenous melatonin significantly alleviated hydrogen peroxide (H2O2)-modulated plant growth, cell damage, and ROS accumulation in Bermuda grass. Additionally, 76 proteins significantly influenced by melatonin during mock or H2O2 treatment were identified by gel-free proteomics using iTRAQ (isobaric tags for relative and absolute quantitation). Metabolic pathway analysis showed that several pathways were markedly enhanced by melatonin and H2O2 treatments, including polyamine metabolism, ribosome pathway, major carbohydrate metabolism, photosynthesis, redox, and amino acid metabolism. Taken together, this study provides more comprehensive insights into the physiological and molecular mechanisms of melatonin in Bermuda grass responses to direct oxidative stress. This may relate to the activation of antioxidants, modulation of metabolic pathways, and extensive proteome reprograming.

Melatonin regulates carbohydrate metabolism and defenses against Pseudomonas syringae pv. tomato DC3000 infection in Arabidopsis thaliana.

Abstract: Melatonin has been reported to promote plant growth and development. Our experiments with Arabidopsis thaliana showed that exogenous applications of this molecule mediated invertase inhibitor (C/VIF)-regulated invertase activity and enhanced sucrose metabolism. Hexoses were accumulated in response to elevated activities by cell wall invertase (CWI) and vacuolar invertase (VI). Analyses of sugar metabolism-related genes revealed differential expression during plant development that was modulated by melatonin. In particular, C/VIF1 and C/VIF2 were strongly down-regulated by exogenous feeding. We also found the elevated CWI activity in melatonin-treated Arabidopsis improved the factors (cellulose, xylose, and galactose) for cell wall reinforcement and callose deposition during Pseudomonas syringae pv. tomato DC3000 infection, therefore, partially induced the pathogen resistance. However, CWI did not involve in salicylic acid (SA)-regulated defense pathway. Taken together, this study reveals that melatonin plays an important role in invertase-related carbohydrate metabolism, plant growth, and pathogen defense.

Reduced silent information regulator 1 signaling exacerbates myocardial ischemia-reperfusion injury in type 2 diabetic rats and the protective effect of melatonin.

Abstract: Diabetes mellitus (DM) increases myocardial oxidative stress and endoplasmic reticulum (ER) stress. Melatonin confers cardioprotective effect by suppressing oxidative damage. However, the effect and mechanism of melatonin on myocardial ischemia-reperfusion (MI/R) injury in type 2 diabetic state are still unknown. In this study, we developed high-fat diet-fed streptozotocin (HFD-STZ) rat, a well-known type 2 diabetic model, to evaluate the effect of melatonin on MI/R injury with a focus on silent information regulator 1 (SIRT1) signaling, oxidative stress, and PERK/eIF2α/ATF4-mediated ER stress. HFD-STZ treated rats were exposed to melatonin treatment in the presence or the absence of sirtinol (a SIRT1 inhibitor) and subjected to MI/R surgery. Compared with nondiabetic animals, type 2 diabetic rats exhibited significantly decreased myocardial SIRT1 signaling, increased apoptosis, enhanced oxidative stress, and ER stress. Additionally, further reduced SIRT1 signaling, aggravated oxidative damage, and ER stress were found in diabetic animals subjected to MI/R surgery. Melatonin markedly reduced MI/R injury by improving cardiac functional recovery and decreasing myocardial apoptosis in type 2 diabetic animals. Melatonin treatment up-regulated SIRT1 expression, reduced oxidative damage, and suppressed PERK/eIF2α/ATF4 signaling. However, these effects were all attenuated by SIRT1 inhibition. Melatonin also protected high glucose/high fat cultured H9C2 cardiomyocytes against simulated ischemia-reperfusion injury-induced ER stress by activating SIRT1 signaling while SIRT1 siRNA blunted this action. Taken together, our study demonstrates that reduced cardiac SIRT1 signaling in type 2 diabetic state aggravates MI/R injury. Melatonin ameliorates reperfusion-induced oxidative stress and ER stress via activation of SIRT1 signaling, thus reducing MI/R damage and improving cardiac function.

Unveiling the mechanism of melatonin impacts on maize seedling growth: sugar metabolism as a case

Abstract: Melatonin regulates growth in many plants; however, the mechanism remains unclear. In this study, exogenous melatonin feeding resulted in both promotional (≤10 μm) and inhibitory (≥100 μm) effects on maize seedling growth. Initial analyses suggested positive correlations between the amount of melatonin and sucrose synthesis and hydrolysis-related gene expression, enzyme activities, and sucrose metabolites. However, assays of photosynthetic rate, hexokinase (HxK) activity, expression of photosynthetic marker genes, and HxK-related genes showed opposite effects under 10 μm (positive) and 100 μm (negative) melatonin treatments. Similarly, 10 μm melatonin accelerated starch catabolism at night, whereas 100 μm melatonin significantly decreased this process and led to starch accumulation in photosynthetic tissues. Furthermore, expression analysis of genes related to sucrose phloem loading resulted in a slight upregulation of sucrose transporters (SUT1 and SUT2) when seedlings were induced with 10 μm melatonin, while treatment with 100 μm melatonin resulted in significant downregulation of these sucrose transporter genes (SUT1 and SUT2), as well as tie-dyed2 (Tdy2) and sucrose export defective 1. Taken together, these results suggest that low doses of melatonin benefit maize seedling growth by promoting sugar metabolism, photosynthesis, and sucrose phloem loading. Conversely, high doses of melatonin inhibit seedling growth by inducing the excessive accumulation of sucrose, hexose and starch, suppressing photosynthesis and sucrose phloem loading.

Melatonin improves bone mineral density at the femoral neck in postmenopausal women with osteopenia: a randomized controlled trial

Abstract: Melatonin is known for its regulation of circadian rhythm. Recently, studies have shown that melatonin may have a positive effect on the skeleton. By increasing age, the melatonin levels decrease, which may lead to a further imbalanced bone remodeling. We aimed to investigate whether treatment with melatonin could improve bone mass and integrity in humans. In a double-blind RCT, we randomized 81 postmenopausal osteopenic women to 1-yr nightly treatment with melatonin 1 mg (N = 20), 3 mg (N = 20), or placebo (N = 41). At baseline and after 1-yr treatment, we measured bone mineral density (BMD) by dual X-ray absorptiometry, quantitative computed tomography (QCT), and high-resolution peripheral QCT (HR-pQCT) and determined calciotropic hormones and bone markers. Mean age of the study subjects was 63 (range 56-73) yr. Compared to placebo, femoral neck BMD increased by 1.4% in response to melatonin (P < 0.05) in a dose-dependent manner (P < 0.01), as BMD increased by 0.5% in the 1 mg/day group (P = 0.55) and by 2.3% (P < 0.01) in the 3 mg/day group. In the melatonin group, trabecular thickness in tibia increased by 2.2% (P = 0.04), and volumetric bone mineral density (vBMD) in the spine, by 3.6% (P = 0.04) in the 3 mg/day. Treatment did not significantly affect BMD at other sites or levels of bone turnover markers; however, 24-hr urinary calcium was decreased in response to melatonin by 12.2% (P = 0.02). In conclusion, 1-yr treatment with melatonin increased BMD at femoral neck in a dose-dependent manner, while high-dose melatonin increased vBMD in the spine. Further studies are needed to assess the mechanisms of action and whether the positive effect of nighttime melatonin will protect against fractures.

Melatonin protects porcine oocyte in vitro maturation from heat stress.

Abstract: Melatonin is a pleiotropic molecule which plays an important role in animal reproductive activities. Because of the increased global warming, the impact of heat stress (HS) on stockbreeding has become an inevitable issue to be solved. To investigate the potential effects of melatonin on the in vitro maturation of porcine oocyte under the HS, a HS model for porcine oocyte maturation has been used in this study and the different concentrations of melatonin (10(-6) -10(-9) m) were also tested for their protective effects on oocytes. The polar body rate, the index of the nuclear maturation of the oocytes, and the cleavage rate as well as the blastocyst rate were measured to evaluate the developmental competence of the oocytes after parthenogenetic activation (PA). The results showed that HS [in vitro maturation (IVM) 20-24 hr, 42°C] significantly reduced the polar body rate of oocytes and the blastocyte rate of porcine PA embryos, while melatonin (10(-7) m) application not only improved polar body rate and blastocyte rate, but also preserved the normal levels of steroid hormone which is disrupted by HS. The presence of melatonin (10(-7) m) during the oocyte maturation under the HS reduced reactive oxygen species (ROS) formation, enhanced glutathione (GSH) production, inhibited cell apoptosis, and increased the gene expressions of SIRT1, AKT2, and Polg2. Importantly, the endogenously occurring melatonin of cumulus-oocyte complexes was significantly induced by HS. The results indicated that melatonin application effectively protected the oocytes from HS. These observations warranted the further studies in vivo regarding to improve the reproductive activities of animals under the global warming environment.

Experimental and clinical aspects of melatonin and clock genes in diabetes.

Abstract: The pineal hormone melatonin influences insulin secretion, as well as glucagon and somatostatin secretion, both in vivo and in vitro. These effects are mediated by two specific, high-affinity, seven transmembrane, pertussis toxin-sensitive, Gi-protein-coupled melatonin receptors, MT1 and MT2. Both isoforms are expressed in the β-cells, α-cells as well as δ-cells of the pancreatic islets of Langerhans and are involved in the modulation of insulin secretion, leading to inhibition of the adenylate cyclase-dependent cyclic adenosine monophosphate as well as cyclic guanosine monophosphate formation in pancreatic β-cells by inhibiting the soluble guanylate cyclase, probably via MT2 receptors. In this way, melatonin also likely inhibits insulin secretion, whereas using the inositol triphosphate pathway after previous blocking of Gi-proteins by pertussis toxin, melatonin increases insulin secretion. Desynchrony of receptor signaling may lead to the development of type 2 diabetes. This notion has recently been supported by genomewide association studies pinpointing variances of the MT2 receptor as a risk factor for this rapidly spreading metabolic disturbance. As melatonin is secreted in a clearly diurnal fashion, it is safe to assume that it also has a diurnal impact on the blood-glucose-regulating function of the islet. Observations of the circadian expression of clock genes (Clock, Bmal1, Per1,2,3, and Cry1,2) in pancreatic islets, as well as in INS1 rat insulinoma cells, may indicate that circadian rhythms are generated in the β-cells themselves. The circadian secretion of insulin from pancreatic islets is clock-driven. Disruption of circadian rhythms and clock function leads to metabolic disturbances, for example, type 2 diabetes. The study of melatonin–insulin interactions in diabetic rat models has revealed an inverse relationship between these two hormones. Both type 2 diabetic rats and patients exhibit decreased melatonin levels and slightly increased insulin levels, whereas type 1 diabetic rats show extremely reduced levels or the absence of insulin, but statistically significant increases in melatonin levels. Briefly, an increase in melatonin levels leads to a decrease in stimulated insulin secretion and vice versa. Melatonin levels in blood plasma, as well as the activity of the key enzyme of melatonin synthesis, AA-NAT (arylalkylamine-N-acetyltransferase) in pineal, are lower in type 2 diabetic rats compared to controls. In contrast, melatonin and pineal AA-NAT mRNA are increased and insulin receptor mRNA is decreased in type 1 diabetic rats, which also indicates a close relationship between insulin and melatonin. As an explanation, it was hypothesized that catecholamines, which reduce insulin levels and stimulate melatonin synthesis, control insulin–melatonin interactions. This conviction stems from the observation that catecholamines are increased in type 1 but are diminished in type 2 diabetes. In this context, another important line of inquiry involves the fact that melatonin protects β-cells against functional overcharge and, consequently, hinders the development of type 2 diabetes.

Melatonin attenuates neutrophil inflammation and mucus secretion in cigarette smoke-induced chronic obstructive pulmonary diseases via the suppression of Erk-Sp1 signaling

Abstract: The incidence of chronic obstructive pulmonary disease (COPD) has substantially increased in recent decade. Cigarette smoke (CS) is the most important risk factor in the development of COPD. In this study, we investigated the effects of melatonin on the development of COPD using a CS and lipopolysaccharide (LPS)-induced COPD model and cigarette smoke condensate (CSC)-stimulated NCI-H292 cells, a human mucoepidermoid carcinoma cell. On day 4, the mice were treated intranasally with LPS. The mice were exposed to CS for 1 hr per day (8 cigarettes per day) from day 1 to day 7. Melatonin (10 or 20 mg/kg) was injected intraperitoneally 1 hr before CS exposure. Melatonin markedly decreased the neutrophil count in the BALF, with reduction in the proinflammatory mediators and MUC5AC. Melatonin inhibited Erk phosphorylation and Sp1 expression induced by CS and LPS treatment. Additionally, melatonin decreased airway inflammation with a reduction in myeloperoxidase expression in lung tissue. In in vitro experiments, melatonin suppressed the elevated expression of proinflammatory mediators induced by CSC treatment. Melatonin reduced Erk phosphorylation and Sp1 expression in CSC-stimulated H292 cells. In addition, cotreatment of melatonin and Erk inhibitors significantly limited the proinflammatory mediators with greater reductions in Erk phosphorylation and Sp1 expression than that observed in H292 cells treated with Erk inhibitor alone. Taken together, melatonin effectively inhibited the neutrophil airway inflammation induced by CS and LPS treatment, which was closely related to downregulation of Erk phosphorylation. These findings suggest that melatonin has a therapeutic potential for the treatment of COPD.

Melatonin induces apoptosis of colorectal cancer cells through HDAC4 nuclear import mediated by CaMKII inactivation

Abstract: Melatonin induces apoptosis in many different cancer cell lines, including colorectal cancer. However, the precise mechanisms involved remain largely unresolved. In this study, we provide evidence to reveal a new mechanism by which melatonin induces apoptosis of colorectal cancer LoVo cells. Melatonin at pharmacological concentrations significantly suppressed cell proliferation and induced apoptosis in a dose-dependent manner. The observed apoptosis was accompanied by the melatonin-induced dephosphorylation and nuclear import of histone deacetylase 4 (HDAC4). Pretreatment with a HDAC4-specific siRNA effectively attenuated the melatonin-induced apoptosis, indicating that nuclear localization of HDAC4 is required for melatonin-induced apoptosis. Moreover, constitutively active Ca(2+) /calmodulin-dependent protein kinase II alpha (CaMKIIα) abrogated the melatonin-induced HDAC4 nuclear import and apoptosis of LoVo cells. Furthermore, melatonin decreased H3 acetylation on bcl-2 promoter, leading to a reduction of bcl-2 expression, whereas constitutively active CaMKIIα(T286D) or HDAC4-specific siRNA abrogated the effect of melatonin. In conclusion, the present study provides evidence that melatonin-induced apoptosis in colorectal cancer LoVo cells largely depends on the nuclear import of HDAC4 and subsequent H3 deacetylation via the inactivation of CaMKIIα.