Melatonin Inhibits Hypoxia-Induced Alzheimer's Disease Pathogenesis by Regulating the Amyloidogenic Pathway in Human Neuroblastoma Cells.

Stroke and Alzheimer's disease (AD) are prevalent age-related diseases; however, the relationship between these two diseases remains unclear. In this study, we aimed to investigate the ability of melatonin, a hormone produced by the pineal gland, to alleviate the effects of ischemic stroke leading to AD by observing the pathogenesis of AD hallmarks. We utilized SH-SY5Y cells under the conditions of oxygen-glucose deprivation (OGD) and oxygen-glucose deprivation and reoxygenation (OGD/R) to establish ischemic stroke conditions. We detected that hypoxia-inducible factor-1α (HIF-1α), an indicator of ischemic stroke, was highly upregulated at both the protein and mRNA levels under OGD conditions. Melatonin significantly downregulated both HIF-1α mRNA and protein expression under OGD/R conditions. We detected the upregulation of ß-site APP-cleaving enzyme 1 (BACE1) mRNA and protein expression under both OGD and OGD/R conditions, while 10 µM of melatonin attenuated these effects and inhibited beta amyloid (Aß) production. Furthermore, we demonstrated that OGD/R conditions were able to activate the BACE1 promoter, while melatonin inhibited this effect. The present results indicate that melatonin has a significant impact on preventing the aberrant development of ischemic stroke, which can lead to the development of AD, providing new insight into the prevention of AD and potential stroke treatments.

Characterization of endotoxin free protein production of brain-derived neurotrophic factor (BDNF) for the study of Parkinson model in SH-SY5Y differentiated cells.

Human neuronal cells are a more appropriate cell model for neurological disease studies such as Alzheimer and Parkinson's disease. SH-SY5Y neuroblastoma cells have been widely used for differentiation into a mature neuronal cell phenotype. The cellular differentiation process begins with retinoic acid incubation, followed by incubation with brain-derived neurotrophic factor (BDNF), a recombinant protein produced in E. coli cells. Endotoxin or lipopolysaccharide (LPS) is the major component of the outer membrane of bacterial cells that triggers the activation of pro-inflammatory cytokines and ultimately cell death. Consequently, any endotoxin contamination of the recombinant BDNF used for cell culture experiments would impact on data interpretation. Therefore, in this study, we expressed the BDNF recombinant protein in bacterial endotoxin-free cells that were engineered to modify the oligosaccharide chain of LPS rendering the LPS unable to trigger the immune response of human cells. The expression of DCX and MAP-2 in differentiated cells indicate that in-house and commercial BDNF are equally effective in inducing differentiation. This suggests that our in-house BDNF protein can be used to differentiate SH-SY5Y neuroblastoma cells without the need for an endotoxin removal step.

Novel functions of the ER-located Hsp40s DNAJB12 and DNAJB14 on proteins at the outer mitochondrial membrane under stress mediated by CCCP.

The endoplasmic reticulum (ER) membrane provides infrastructure for intracellular signaling, protein degradation, and communication among the ER lumen, cytosol, and nucleus via transmembrane and membrane-associated proteins. Failure to maintain homeostasis at the ER leads to deleterious conditions in humans, such as protein misfolding-related diseases and neurodegeneration. The ER transmembrane heat shock protein 40 (Hsp40) proteins, including DNAJB12 (JB12) and DNAJB14 (JB14), have been studied for their importance in multiple aspects of cellular events, including degradation of misfolded membrane proteins, proteasome-mediated control of proapoptotic Bcl-2 members, and assembly of multimeric ion channels. This study elucidates a novel facet of JB12 and JB14 in that their expression could be regulated in response to stress caused by the presence of ER stressors and the mitochondrial potential uncoupler CCCP. Furthermore, JB14 overexpression could affect the level of PTEN-induced kinase 1 (PINK1) expression under CCCP-mediated stress. Cells with genetic knockout (KO) of DNAJB12 and DNAJB14 exhibited an altered kinetic of phosphorylated Drp1 in response to the stress caused by CCCP treatment. Surprisingly, JB14-KO cells exhibited a prolonged stabilization of PINK1 during chronic exposure to CCCP. Cells depleted with JB12 or JB14 also revealed an increase in the mitochondrial count and branching. Hence, this study indicates the possible novel functions of JB12 and JB14 involving mitochondria in nonstress conditions and under stress caused by CCCP.

Melatonin suppresses inflammation and blood‒brain barrier disruption in rats with vascular dementia possibly by activating the SIRT1/PGC-1α/PPARγ signaling pathway.

Chronic cerebral hypoxia (CCH) is caused by a reduction in cerebral blood flow, and cognitive impairment has been the predominant feature that occurs after CCH. Recent reports have revealed that melatonin is proficient in neurodegenerative diseases. However, the molecular mechanism by which melatonin affects CCH remains uncertain. In this study, we aimed to explore the role and underlying mechanism of melatonin in inflammation and blood‒brain barrier conditions in rats with CCH. Male Wistar rats were subjected to permanent bilateral common carotid artery occlusion (BCCAO) to establish the VAD model. Rats were randomly divided into four groups: Sham, BCCAO, BCCAO treated with melatonin (10 mg/kg), and BCCAO treated with resveratrol (20 mg/kg). All drugs were administered once daily for 4 weeks. Our results showed that melatonin attenuated cognitive impairment, as demonstrated by the Morris water maze tests. Furthermore, melatonin reduced the activation of inflammation by attenuating the phosphorylated nuclear factor of kappa light polypeptide gene enhancer in B cells inhibitor alpha (pIκBα), causing the suppression of proteins related to inflammation and inflammasome formation. Moreover, immunohistochemistry revealed that melatonin reduced glial cell activation and proliferation, which were accompanied by Western blotting results. Additionally, melatonin also promoted the expression of sirtuin-1 (SIRT1), peroxisome proliferator-activated receptor-gamma coactivator 1-alpha (PGC-1α), and peroxisome proliferator-activated receptor-gamma (PPARγ), causing attenuated blood‒brain barrier (BBB) disruption by increasing tight junction proteins. Taken together, our results prove that melatonin treatment modulated inflammation and BBB disruption and improved cognitive function in VaD rats, partly by activating the SIRT1/PGC-1α/PPARγ signaling pathway.

Melatonin attenuates reactive astrogliosis and glial scar formation following cerebral ischemia and reperfusion injury mediated by GSK-3ß and RIP1K.

Even though astrocytes have been widely reported to support several brain functions, studies have emerged that they exert deleterious effects on the brain after ischemia and reperfusion (I/R) injury. The present study investigated the neuroprotective effects of melatonin on the processes of reactive astrogliosis and glial scar formation, as well as axonal regeneration after transient middle cerebral artery occlusion. Male Wistar rats were randomly divided into four groups: sham-operated, I/R, I/R treated with melatonin, and I/R treated with edaravone. All drugs were administered via intraperitoneal injection at the onset of reperfusion and were continued until the rats were sacrificed on Day 7 or 14 after the surgery. Melatonin presented long-term benefits on cerebral damage after I/R injury, as demonstrated by a decreased infarct volume, histopathological changes, and reduced neuronal cell death. We also found that melatonin attenuated reactive astrogliosis and glial scar formation and, consequently, enhanced axonal regeneration and promoted neurobehavioral recovery. Furthermore, glycogen synthase kinase-3 beta (GSK-3ß) and receptor-interacting serine/threonine-protein 1 kinase (RIP1K), which had previously been revealed as proteins involved in astrocyte responses, were significantly reduced after melatonin administration. Taken together, melatonin effectively counteracted the deleterious effects due to astrocyte responses and improved axonal regeneration to promote functional recovery during the chronic phase of cerebral I/R injury by inhibiting GSK-3ß and RIP1K activities.

A Synopsis of Multitarget Potential Therapeutic Effects of Huperzine A in Diverse Pathologies-Emphasis on Alzheimer's Disease Pathogenesis.

Numerous challenges are confronted when it comes to the recognition of therapeutic agents for treating complex neurodegenerative diseases like Alzheimer's disease (AD). The perplexing pathogenicity of AD embodies cholinergic dysfunction, amyloid beta (Aß) aggregation, neurofibrillary tangle formation, neuroinflammation, mitochondrial disruption along with vicious production of reactive oxygen species (ROS) generating oxidative stress. In this frame of reference, drugs with multi target components could prove more advantageous to counter complex pathological mechanisms that are responsible for AD progression. For as much as, medicinal plant based pharmaco-therapies are emerging as potential candidates for AD treatment keeping the efficacy and safety parameters in terms of toxicity and side effects into consideration. Huperzine A (Hup A) is a purified alkaloid compound extracted from a club moss called Huperzia serrata. Several studies have reported both cholinergic and non-cholinergic effects of this compound on AD with significant neuroprotective properties. The present review convenes cumulative demonstrations of neuroprotection provided by Hup A in in vitro, in vivo, and human studies in various pathologies. The underlying molecular mechanisms of its actions have also been discussed. However, more profound evidence would certainly promote the therapeutic implementation of this drug thus furnishing decisive insights into AD therapeutics and various other pathologies along with preventive and curative management.

Melatonin Attenuates High Glucose-Induced Changes in Beta Amyloid Precursor Protein Processing in Human Neuroblastoma Cells.

Diabetes mellitus (DM), one of metabolic diseases, has been suggested as a risk factor for Alzheimer's disease (AD). However, how the metabolic pathway activates amyloid precursor protein (APP) processing enzymes then contributes to the increase of amyloid-beta (Aß) production, is not clearly understood. In the present study, we aimed to examine the protective effect of melatonin against hyperglycemia-induced alterations in the amyloidogenic pathway. High concentration of glucose was used to induce hyperglycemia in human neuroblastoma SH-SY5Y cells. We found that 30 mM glucose affected the expression of insulin receptors and glucose transporters, which indicated the disruption of glucose sensing. High glucose induced the activation of the phosphorylated protein kinase B (pAkt)/GSK-3ß signaling pathway and a significant increase in the expression of ß-site beta APP cleaving enzyme (BACE1), presenilin1 (PS1) and Aß42. Pretreatment with melatonin significantly reversed these parameters. We also showed that these effects are similar to those effects in the presence of the GSK-3ß blocker, N-(4-methoxybenyl)-N'-(5-nitro-1,3-thiazol-2-yl) urea (ARA) in glucose-treated hyperglycemic cells. These suggested that melatonin exerted an inhibitory effect on the activation of APP-cleaving enzymes via the GSK-3ß signaling pathway. Pretreatment with luzindole, a melatonin receptor MT1 antagonist, significantly prevented the effect of melatonin on the glucose-induced increase level of APP processing enzymes. This suggested that melatonin attenuated the toxic effect on hyperglycemia involving the amyloidogenic pathway partially mediated via melatonin receptor. Taken together the present results suggested that melatonin has a beneficial role in preventing Aß generation in a cellular model of hyperglycemia-induced DM.

Combination of Melatonin and Small Molecules Improved Reprogramming Neural Cell Fates via Autophagy Activation.

Reprogramming cell fates towards mature cell types are a promising cell supply for treating degenerative diseases. Recently, transcription factors and some small molecules have turned into impressive modulating elements for reprogramming cell fates. Melatonin, a pineal hormone, has neuroprotective functions including neural stem cell (NSC) proliferative and differentiative modulation in both embryonic and adult brain. We developed a protocol that could be implemented in the direct reprogramming of human skin fibroblast towards neural cells by using histone deacetylase (HDAC) inhibitor, glycogen synthase kinase-3 (GSK3) inhibitor (CHIR99021), c-Jun N-terminal kinase (JNK) inhibitor, rho-associated protein kinase inhibitor (Y-27632), cAMP activator, and melatonin treatment. We found that melatonin enhanced neural-transcription factor genes expressions, including brain-specific homeobox/POU domain protein 2 (BRN2), Achaete-Scute Family BHLH transcription Factor 1 (ASCL1), and Myelin Transcription Factor 1 Like (MYT1L). Melatonin also increased the expression of different neural-specific proteins such as doublecortin (DCX), Sex determining region Y-box 2 (Sox2), and neuronal nuclei (NeuN) compared with other five small molecules (valproic acid (VPA), CHIR99021, Forskolin, 1,9 pyrazoloanthrone (SP600125), and Y-27632) combination in the presence and absence of melatonin. A noticeable upregulation of autophagy proteins (microtubule-associated protein 1A/1B-light chain 3 (LC3) and Beclin-1) were seen in the melatonin treatment during the induction period while these were reverted in the presence of L-leucine, an autophagy inhibitor. In addition, the expression of NeuN was also significantly reduced by L-leucine. Collectively, our findings revealed an activation of autophagy during neural induction; melatonin enhanced reprogramming efficiency for neuron induction through the modulation of autophagy activation.

Melatonin and its derivative disrupt cancer stem-like phenotypes of lung cancer cells via AKT downregulation.

Cancer stem cells (CSCs), a small subpopulation of tumour cells, have properties of self-renewal and multipotency, which drive cancer progression and resistance to current treatments. Compounds potentially targeting CSCs have been recently developed. This study shows how melatonin, an endogenous hormone synthesised by the pineal gland, and its derivative suppress CSC-like phenotypes of human non-small cell lung cancer (NSCLC) cell lines, H460, H23, and A549. The effects of MLT and its derivative, acetyl melatonin (ACT), on CSC-like phenotypes were investigated using assays for anchorage-independent growth, three-dimensional spheroid formation, scratch wound healing ability, and CSC marker and upstream protein signalling expression. Enriched CSC spheroids were used to confirm the effect of both compounds on lung cancer cells. MLT and ACT inhibited CSC-like behaviours by suppression of colony and spheroid formation in NSCLC cell lines. Their effects on spheroid formation were confirmed in CSC-enriched H460 cells. CSC markers, CD133 and ALDH1A1, were depleted by both compounds. The behaviour and factors associated to epithelial-mesenchymal transition, as indicated by cell migration and the protein vimentin, were also decreased by MLT and ACT. Mechanistically, MLT and ACT decreased the expression of stemness proteins Oct-4, Nanog, and ß-catenin by reducing active AKT (phosphorylated AKT). Suppression of the AKT pathway was not mediated through melatonin receptors. This study demonstrates a novel role, and its underlying mechanism, for MLT and its derivative ACT in suppression of CSC-like phenotypes in NSCLC cells, indicating that they are potential candidates for lung cancer treatment.

5,6,7,4'-Tetramethoxyflavanone alleviates neurodegeneration in a dexamethasone-induced neurodegenerative mouse model through promotion of neurogenesis via the Raf/ERK1/2 pathway.

Adult neurogenesis plays an important role in improving cognitive functions. Neurogenesis generates new neurons, a process mediated by neural stem cell proliferation, migration, and differentiation. Long-term exposure to high levels of glucocorticoid results in the suppression of neurogenesis pathways and leads to the onset of cognitive impairment. The induction of neurogenesis by a potent bioactive compound is considered the most promising treatment for neurodegenerative disorders. 5,6,7,4'-Tetramethoxyflavanone (TMF) is a flavonoid compound isolated from Chromolaena odorata (L.) R. M. King & H. Rob. Previous study showed that TMF improved cognitive impairment by attenuating Aß production and pTau expression, thereby increased cell survival and promoted synaptic plasticity. The aim of this study was to investigate the effect of TMF on dexamethasone (DEX)-suppressed neurogenesis in mice. Mice received DEX for 28 days before being treated with TMF for additional 30 days. Mice were randomly divided into four groups: control, TMF, DEX, and DEX + TMF. TMF promoted neurogenesis by increasing BrdU-positive cells, Prox1, doublecortin, and Nestin expression. TMF also upregulated the expression of Raf and extracellular-signal-regulated kinase (ERK)1/2, which are pivotal for neurogenesis signaling. In conclusion, TMF promoted neurogenesis-related protein expression in the proliferation, differentiation, and maturation phases via Raf/ERK1/2 signaling pathway.

Liraglutide Suppresses Tau Hyperphosphorylation, Amyloid Beta Accumulation through Regulating Neuronal Insulin Signaling and BACE-1 Activity.

Neuronal insulin resistance is a significant feature of Alzheimer's disease (AD). Accumulated evidence has revealed the possible neuroprotective mechanisms of antidiabetic drugs in AD. Liraglutide, a glucagon-like peptide-1 (GLP-1) analog and an antidiabetic agent, has a benefit in improving a peripheral insulin resistance. However, the neuronal effect of liraglutide on the model of neuronal insulin resistance with Alzheimer's formation has not been thoroughly investigated. The present study discovered that liraglutide alleviated neuronal insulin resistance and reduced beta-amyloid formation and tau hyperphosphorylation in a human neuroblostoma cell line, SH-SY5Y. Liraglutide could effectively reverse deleterious effects of insulin overstimulation. In particular, the drug reversed the phosphorylation status of insulin receptors and its major downstream signaling molecules including insulin receptor substrate 1 (IRS-1), protein kinase B (AKT), and glycogen synthase kinase 3 beta (GSK-3ß). Moreover, liraglutide reduced the activity of beta secretase 1 (BACE-1) enzyme, which then decreased the formation of beta-amyloid in insulin-resistant cells. This indicated that liraglutide can reverse the defect of phosphorylation status of insulin signal transduction but also inhibit the formation of pathogenic Alzheimer's proteins like Aß in neuronal cells. We herein provided the possibility that the liraglutide-based therapy may be able to reduce such deleterious effects caused by insulin resistance. In view of the beneficial effects of liraglutide administration, these findings suggest that the use of liraglutide may be a promising therapy for AD with insulin-resistant condition.

Elevation of Cleaved p18 Bax Levels Associated with the Kinetics of Neuronal Cell Death during Japanese Encephalitis Virus Infection.

Japanese encephalitis virus (JEV) infection induces uncontrolled neuronal apoptosis, leading to irreversible brain damage. However, the mechanism of JEV-induced neuronal apoptosis has not been clearly elucidated. This study aimed to investigate both virus replication and neuronal cell apoptosis during JEV infection in human neuroblastoma SH-SY5Y cells. As a result, the kinetic productions of new viral progeny were time- and dose-dependent. The stimulation of SH-SY5Y cell apoptosis was dependent on the multiplicity of infections (MOIs) and infection periods, particularly during the late period of infection. Interestingly, we observed that of full-length Bax (p21 Bax) level started to decrease, which corresponded to the increased level of its cleaved form (p18 Bax). The formation of p18 Bax resulting in cytochrome c release into the cytosol appeared to correlate with JEV-induced apoptotic cell death together with the activation of caspase-3/7 activity, especially during the late stage of a robust viral infection. Therefore, our results suggest another possible mechanism of JEV-induced apoptotic cell death via the induction of the proteolysis of endogenous p21 Bax to generate p18 Bax. This finding could be a new avenue to facilitate novel drug discovery for the further development of therapeutic treatments that could relieve neuronal damage from JEV infection.

Comparing the Effects of Melatonin with Caloric Restriction in the Hippocampus of Aging Mice: Involvement of Sirtuin1 and the FOXOs Pathway.

It has been suggested that age-related neurodegeneration might be associated with neuropeptide Y (NPY); sirtuin1 (SIRT1) and forkhead box transcription factors O subfamily (FOXOs) pathways. Melatonin, a hormone mainly secreted by the pineal gland, is another anti-aging agent associated with the SIRT1-FOXOs pathway. This study aimed to compare the effects of melatonin (Mel) and caloric restriction (CR) on the expression of Sirt1, FoxO1, FoxO3a and FOXOs target genes in the aging mouse hippocampus. Neuropeptide Y-knockout (NpyKO) and wild-type (WT) male mice aged 19 months were previously treated either with food ad libitum or CR for 16 months. WT old animals were divided into four groups: control, CR, Mel and CR+Mel treated groups. The Mel and CR+Mel were treated with melatonin 10 mg/kg, daily, subcutaneously for 7 consecutive days. Mel treatment upregulated the mRNA expression of Sirt1, FOXOs (FoxO1 and FoxO3a) target genes that regulated the cell cycle [e.g., cyclin-dependent kinase inhibitor 1B (p27)], Wingless and INT-1 (Wnt1) and inducible signaling pathway protein 1 (Wisp1) in the aged mouse hippocampus. CR treatment also showed the similar actions. However, the mRNA expression of Sirt1, FoxO1, FoxO3a, p27 or Wisp1 did not alter in the CR+Mel group when compared with CR or Mel group. Melatonin could not produce any additive effect on the CR treatment group, suggesting that both treatments mimicked the effect, possibly via the same pathway. NPY which mediates physiological adaptations to energy deficits is an essential link between CR and longevity in mice. In order to focus on the role of Npy in mediating the effects of melatonin, the gene expression between NpyKO and WT male mice were compared. Our data showed that, in the absence of Npy, melatonin could not mediate effects on those gene expressions, suggesting that Npy was required for melatonin to mediate the effect, possibly, on life extension.

Melatonin ameliorates Aß42 -induced alteration of ßAPP-processing secretases via the melatonin receptor through the Pin1/GSK3ß/NF-κB pathway in SH-SY5Y cells.

Melatonin is involved in the physiological regulation of the ß-amyloid precursor protein (ßAPP)-cleaving secretases which are responsible for generation of the neurotoxic amyloid beta (Aß) peptide, one of the hallmarks of Alzheimer's disease (AD) pathology. In this study, we aimed to determine the underlying mechanisms of this regulation under pathological conditions. We establish that melatonin prevents Aß42 -induced downregulation of a disintegrin and metalloproteinase domain-containing protein 10 (ADAM10) as well as upregulation of ß-site APP-cleaving enzyme 1 (BACE1) and presenilin 1 (PS1) in SH-SY5Y cell cultures. We also demonstrate that the intrinsic mechanisms of the observed effects occurred via regulation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and glycogen synthase kinase (GSK)-3ß as melatonin reversed Aß42 -induced upregulation and nuclear translocation of NF-κBp65 as well as activation of GSK3ß via its receptor activation. Furthermore, specific blocking of the NF-κB and GSK3ß pathways partially abrogated the Aß42 -induced reduction in the BACE1 and PS1 levels. In addition, GSK3ß blockage affected α-secretase cleavage and modulated nuclear translocation of NF-κB. Importantly, our study for the first time shows that peptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (Pin1) is a crucial target of melatonin. The compromised levels and/or genetic variation of Pin1 are associated with age-dependent tau and Aß pathologies and neuronal degeneration. Interestingly, melatonin alleviated the Aß42 -induced reduction of nuclear Pin1 levels and preserved the functional integrity of this isomerase. Our findings illustrate that melatonin attenuates Aß42 -induced alterations of ßAPP-cleaving secretases possibly via the Pin1/GSK3ß/NF-κB pathway.

Neuroprotective effects of melatonin on amphetamine-induced dopaminergic fiber degeneration in the hippocampus of postnatal rats.

Chronic amphetamine (AMPH) abuse leads to damage of the hippocampus, the brain area associated with learning and memory process. Previous results have shown that AMPH-induced dopamine neurotransmitter release, reactive oxygen species formation, and degenerative protein aggregation lead to neuronal death. Melatonin, a powerful antioxidant, plays a role as a neuroprotective agent. The objective of this study was to investigate whether the protective effect of melatonin on AMPH-induced hippocampal damage in the postnatal rat acts through the dopaminergic pathway. Four-day-old postnatal rats were subcutaneously injected with 5-10 mg/kg AMPH and pretreated with 10 mg/kg melatonin prior to AMPH exposure for seven days. The results showed that melatonin decreased the AMPH-induced hippocampal neuronal degeneration in the dentate gyrus, CA1, and CA3. Melatonin attenuated the reduction in the expression of hippocampal synaptophysin, PSD-95, α-synuclein, and N-methyl-D-aspartate (NMDA) receptor protein and mRNA caused by AMPH. Melatonin attenuated the AMPH-induced reduction in dopamine transporter (DAT) protein expression in the hippocampus and the reduction in mRNA expression in the ventral tegmental area (VTA). Immunofluorescence demonstrated that melatonin not only prevented the AMPH-induced loss of DAT and NMDA receptor but also prevented AMPH-induced α-synuclein overexpression in the dentate gyrus, CA1, and CA3. Melatonin decreased the AMPH-induced reduction in the protein and mRNA of the NMDA receptor downstream signaling molecule, calcium/calmodulin-dependent protein kinase II (CaMKII), and the melatonin receptors (MT1 and MT2). This study showed that melatonin prevented AMPH-induced toxicity in the hippocampus of postnatal rats possibly via its antioxidative effect and mitochondrial protection.

Melatonin as a mitochondrial protector in neurodegenerative diseases.

Mitochondria are crucial organelles as their role in cellular energy production of eukaryotes. Because the brain cells demand high energy for maintaining their normal activities, disturbances in mitochondrial physiology may lead to neuropathological events underlying neurodegenerative conditions such as Alzheimer's disease, Parkinson's disease and Huntington's disease. Melatonin is an endogenous compound with a variety of physiological roles. In addition, it possesses potent antioxidant properties which effectively play protective roles in several pathological conditions. Several lines of evidence also reveal roles of melatonin in mitochondrial protection, which could prevent development and progression of neurodegeneration. Since the mitochondrial dysfunction is a primary event in neurodegeneration, the neuroprotection afforded by melatonin is thereby more effective in early stages of the diseases. This article reviews mechanisms which melatonin exerts its protective roles on mitochondria as a potential therapeutic strategy against neurodegenerative disorders.

Melatonin reverses H2 O2 -induced senescence in SH-SY5Y cells by enhancing autophagy via sirtuin 1 deacetylation of the RelA/p65 subunit of NF-κB.

Autophagy, a degradation mechanism that plays a major role in maintaining cellular homeostasis and diminishes in aging, is considered an aging characteristic. Melatonin is an important hormone that plays a wide range of physiological functions, including the anti-aging effect, potentially via the regulation of the Sirtuin1 (SIRT1) pathway. The deacetylation ability of SIRT1 is important for controlling the function of several transcription factors, including nuclear factor kappa B (NF-ĸB). Apart from inflammation, NF-ĸB can regulate autophagy by inhibiting Beclin1, an initiator of autophagy. Although numerous studies have revealed the role of melatonin in regulating autophagy, very limited experiments have shown that melatonin can increase autophagic activity via SIRT1 in a senescent model. This study focuses on the effect of melatonin on autophagy via the deacetylation activity of SIRT1 on RelA/p65, a subunit of NF-ĸB, to determine whether melatonin can attenuate the aging condition. SH-SY5Y cells were treated with H2 O2 to induce the senescent state. These results demonstrated that melatonin reduced a number of beta-galactosidase (SA-ßgal)-positive cells, a senescent marker. In addition, melatonin increased the protein levels of SIRT1, Beclin1, and LC3-II, a hallmark protein of autophagy, and reduced the levels of acetylated-Lys310 in the p65 subunit of NF-ĸB in SH-SY5Y cells treated with H2 O2 . Furthermore, in the presence of SIRT1 inhibitor, melatonin failed to increase autophagic markers. The present data indicate that melatonin enhances autophagic activity via the SIRT1 signaling pathway. Taken together, we propose that in modulating autophagy, melatonin may provide a therapeutically beneficial role in the anti-aging processes.

Neuritogenic activity of bi-functional bis-tryptoline triazole.

Alzheimer's disease (AD) is a common neurodegenerative disorder, one of the hallmarks of which is the deposition of aggregated ß-amyloid peptides (Aß40,42) as plaques in the brain. Oligomers of these peptides have been reported to be toxic and to inhibit neurite outgrowth, as evidenced by neurite dystrophy and significant loss of synaptic connectivity of neurons in the AD brain resulting in cognitive decline. These peptides also react with biological metal in the brain to generate free radicals, thereby aggravating neuronal cell injury and death. Herein, multifunctional triazole-based compounds acting on multiple targets, namely ß-secretase (BACE1), ß-amyloid peptides (Aß) as well as those possessing metal chelation and antioxidant properties, were developed and evaluated for neuritogenic activity in P19-derived neurons. At the non-cytotoxic concentration (1nM), all multifunctional compounds significantly enhanced neurite outgrowth. New bis-tryptoline triazole (BTT) increased the neurite length and neurite number, by 93.25% and 136.09% over the control, respectively. This finding demonstrates the ability of multifunctional compounds targeting Aß to enhance neurite outgrowth in addition to their neuroprotective action.

Effects of melatonin on severe crush spinal cord injury-induced reactive astrocyte and scar formation.

The present work aimed at analyzing the effects of melatonin on scar formation after spinal cord injury (SCI). Upregulation of reactive astrocyte under SCI pathological conditions has been presented in several studies. It has been proved that the crucial factor in triggering this upregulation is proinflammatory cytokines. Moreover, scar formation is an important barrier to axonal regeneration through the lesion area. Melatonin plays an important role in reducing inflammation, but its effects on scar formation in the injured spinal cord remain unknown. Hence, we used the model of severe crush injury in mice to investigate the effects of melatonin on scar formation. Mice were randomly separated into four groups; SCI, SCI+Melatonin 1 (single dose), SCI+Melatonin 14 (14 daily doses), and control. Melatonin was administered by intraperitoneal injection (10 mg/kg) after injury. Immunohistochemical analysis, Western blot, and behavioral evaluation were used to explore the effects of melatonin after SCI for 14 days. The melatonin-treated mice presented higher expression of neuronal markers (P < 0.001). Remarkably, the inflammatory response appeared to be greatly reduced in the SCI+Melatonin 14 group (P < 0.001), which also displayed less scar formation (P < 0.05). These findings suggest that melatonin inhibits scar formation by acting on inflammatory cytokines after SCI. Overall, our results suggest that melatonin is a promising treatment strategy after SCI that deserves further investigation. © 2016 Wiley Periodicals, Inc.

The Nicotinic α6-Subunit Selective Antagonist bPiDI Reduces Alcohol Self-Administration in Alcohol-Preferring Rats.

Cigarettes and alcohol are the most abused substances in the world and are commonly co-abused. Nicotine primarily acts in the brain on nicotinic acetylcholine receptors (nAChR), which are also a target for alcohol. The alpha6 subunit of nAChR is expressed almost exclusively in the brain reward system and may modulate the rewarding properties of alcohol and nicotine. Recently, N,N-decane-1,10-diyl-bis-3-picolinium diiodide (bPiDI) was synthesized as a selective, brain penetrant α6 subunit antagonist that reduces nicotine self-administration. The current study aimed to examine the effects of bPiDI on alcohol self-administration in inbred alcohol-preferring (iP) rats. Adult, male iP rats were trained to self-administer alcohol or sucrose. Once stable responding was achieved, rats were injected with bPiDI (1, 3 mg/kg, i.p.) and tested for self-administration under fixed and progressive ratio schedules of reinforcement. They subsequently underwent extinction, in which no rewards or cues were presented in the operant chambers. Then, they were injected with bPiDI prior to testing for cue-induced reinstatement of reward seeking. bPiDI (3 mg/kg) significantly reduced alcohol self-administration in both fixed and progressive ratios without any effects on sucrose self-administration or locomotor activity. In contrast, bPiDI (3 mg/kg) did not inhibit cue-induced reinstatement of either alcohol or sucrose seeking. The results support the involvement of α6 containing nAChR in reinforcing effects of alcohol, but not relapse to alcohol-seeking, without any impact on responding for a natural reward or general activity. bPiDI may be a potential lead molecule for a therapeutic strategy to limit nicotine and alcohol consumption.