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1.
Melatonin (N‐acetyl‐5‐methoxytryptamine)/MT2 receptor‐dependent epigenetic modification represents a novel pathway in the treatment of neuropathic pain. Because spinal ten‐eleven translocation methylcytosine dioxygenase 1 (Tet1)‐dependent epigenetic demethylation has recently been linked to pain hypersensitivity, we hypothesized that melatonin/MT2‐dependent analgesia involves spinal Tet1‐dependent demethylation. Here, we showed that spinal Tet1 gene transfer by intrathecal delivery of Tet1‐encoding vectors to naïve rats produced profound and long‐lasting nociceptive hypersensitivity. In addition, enhanced Tet1 expression, Tet1‐metabotropic glutamate receptor subtype 5 (mGluR5) promoter coupling, demethylation at the mGluR5 promoter, and mGluR5 expression in dorsal horn neurons were observed. Rats subjected to spinal nerve ligation and intraplantar complete Freund's adjuvant injection displayed tactile allodynia and behavioral hyperalgesia associated with similar changes in the dorsal horn. Notably, intrathecal melatonin injection reversed the protein expression, protein‐promoter coupling, promoter demethylation, and pain hypersensitivity induced by Tet1 gene transfer, spinal nerve ligation, and intraplantar complete Freund's adjuvant injection. All the effects caused by melatonin were blocked by pretreatment with a MT2 receptor‐selective antagonist. In conclusion, melatonin relieves pain by impeding Tet1‐dependent demethylation of mGluR5 in dorsal horn neurons through the MT2 receptor. Our findings link melatonin/MT2 signaling to Tet1‐dependent epigenetic demethylation of nociceptive genes for the first time and suggest melatonin as a promising therapy for the treatment of pain.  相似文献   

2.
Melatonin (MLT) exerts its physiological effects principally through two high‐affinity membrane receptors MT1 and MT2. Understanding the exact mechanism of MLT action necessitates the use of highly selective agonists/antagonists to stimulate/inhibit a given MLT receptor. The respective distribution of MT1 and MT2 within the CNS and elsewhere is controversial, and here we used a “knock‐in” strategy replacing MT1 or MT2 coding sequences with a LacZ reporter. The data show striking differences in the distribution of MT1 and MT2 receptors in the mouse brain: whereas the MT1 subtype was expressed in very few structures (notably including the suprachiasmatic nucleus and pars tuberalis), MT2 subtype receptors were identified within numerous brain regions including the olfactory bulb, forebrain, hippocampus, amygdala and superior colliculus. Co‐expression of the two subtypes was observed in very few structures, and even within these areas they were rarely present in the same individual cell. In conclusion, the expression and distribution of MT2 receptors are much more widespread than previously thought, and there is virtually no correspondence between MT1 and MT2 cellular expression. The precise phenotyping of cells/neurons containing MT1 or MT2 receptor subtypes opens new perspectives for the characterization of links between MLT brain targets, MLT actions and specific MLT receptor subtypes.  相似文献   

3.
Mammalian cells have developed several mechanisms to sense viruses and initiate adequate responses such as production of interferons. Interferons activate the antiviral response through the Jak‐STAT signalling pathway. To establish a chronic infection, viruses need to counteract this barrier of defence. The hepatitis C and hepatitis B viruses are known to up‐regulate the expression of protein phosphatase 2A (PP2A). In this study, we show that PP2Ac associates with Jak1/Tyk2/STAT1 and reduces Jak1/Tyk2/STAT1 phosphorylation resulting in an impairment of the IFNα‐induced HCV antiviral response. Using the fully infectious HCV cell culture system (HCVcc), we demonstrate that the PP2A catalytic activity is not required to block the antiviral effect of IFNα, although it is needed to support HCVcc replication. Our data suggest an important contribution of virus‐induced PP2Ac up‐regulation in the establishment of a chronic infection.  相似文献   

4.
High‐mobility group box 1 (HMGB1) proteins are substantially up‐regulated in acute and chronic hepatitis. However, the immunopathogenic role of HMGB1 in patients with chronic hepatitis B (CHB) has not been elucidated. In this study, using a cohort of 36 CHB patients, we demonstrated a crucial role for HMGB1 to modulate balance between regulatory T (Treg) and T helper 17 (Th17) cells via the toll‐like receptor (TLR)‐4‐interleukin (IL)‐6 pathway. Serum HMGB1 levels were dramatically higher in CHB patients and increased along with liver injury, inflammation and fibrosis. Notably, HMGB1 increased along with decreased Treg/Th17 cells ratios in the periphery or intrahepatic microenvironment, which provides a clue for HMGB1 to favour Th17 responses whereas inhibit Treg responses. For in vitro studies, serum pools were constructed with serum from CHB patients at an advanced stage, whereas peripheral blood mononuclear cells (PBMC) pools were constructed with cells from those at an early stage. CHB‐serum significantly enhanced retinoic acid‐related orphan receptor‐γt (RORγt), whereas they inhibited forkhead box P3 (Foxp3) expression in CHB‐PBMC, which could be reversed by blocking of HMGB1, TLR4, or IL‐6. Besides, recombinant HMGB1 (rHMGB1) dose‐dependently up‐regulated RORγt whereas down‐regulated Foxp3 expression in CHB‐PBMC, and meanwhile, rHMGB1 enhanced TLR4 and IL‐6 expression in CHB‐PBMC. Moreover, the axis of HMGB1–TLR4‐IL‐6–Treg/Th17 required noncontact interactions between CD4 and non‐CD4 cells. In addition, rHMGB1 down‐regulated anti‐inflammatory proteins on CD4+CD25+ cells whereas up‐regulated pro‐inflammatory cytokines in CD4+CD25 cells. In summary, enriched HMGB1 in CHB patients shifts Treg/Th17 balance to Th17 dominance via the TLR4‐IL‐6 pathway, which exacerbates liver injury and inflammation.  相似文献   

5.
Melatonin is an effector of the diurnal clock on pancreatic islets. The membrane receptor‐transmitted inhibitory influence of melatonin on insulin secretion is well established and contrasts with the reported stimulation of glucagon release from α‐cells. Virtually, nothing is known concerning the melatonin‐mediated effects on islet δ‐cells. Analysis of a human pancreatic δ‐cell model, the cell line QGP‐1, and the use of a somatostatin‐specific radioimmunoassay showed that melatonin primarily has an inhibitory effect on somatostatin secretion in the physiological concentration range. In the pharmacological range, melatonin elicited slightly increased somatostatin release from δ‐cells. Cyclic adenosine monophosphate (cAMP) is the major second messenger dose‐dependently stimulating somatostatin secretion, in experiments employing the membrane‐permeable 8‐Br‐cAMP. 8‐Br‐cyclic guanosine monophosphate proved to be of only minor relevance to somatostatin release. As the inhibitory effect of 1 nm melatonin was reversed after incubation of QGP‐1 cells with the nonselective melatonin receptor antagonist luzindole, but not with the MT2‐selective antagonist 4‐P‐PDOT (4‐phenyl‐2‐propionamidotetraline), an involvement of the MT1 receptor can be assumed. Somatostatin release from the δ‐cells at low glucose concentrations was significantly inhibited during co‐incubation with 1 nm melatonin, an effect which was less pronounced at higher glucose levels. Transient expression experiments, overexpressing MT1, MT2, or a deletion variant as a control, indicated that the MT1 and not the MT2 receptor was the major transmitter of the inhibitory melatonin effect. These data point to a significant influence of melatonin on pancreatic δ‐cells and on somatostatin release.  相似文献   

6.
Neuropathic pain, a highly debilitating pain condition that commonly occurs after nerve damage, is a reflection of the aberrant excitability of dorsal horn neurons. This pathologically altered neurotransmission requires a communication with spinal microglia activated by nerve injury. However, how normal resting microglia become activated remains unknown. Here we show that in naive animals spinal microglia express a receptor for the cytokine IFN-γ (IFN-γR) in a cell-type-specific manner and that stimulating this receptor converts microglia into activated cells and produces a long-lasting pain hypersensitivity evoked by innocuous stimuli (tactile allodynia, a hallmark symptom of neuropathic pain). Conversely, ablating IFN-γR severely impairs nerve injury-evoked microglia activation and tactile allodynia without affecting microglia in the contralateral dorsal horn or basal pain sensitivity. We also find that IFN-γ-stimulated spinal microglia show up-regulation of Lyn tyrosine kinase and purinergic P2X4 receptor, crucial events for neuropathic pain, and genetic approaches provide evidence linking these events to IFN-γR-dependent microglial and behavioral alterations. These results suggest that IFN-γR is a key element in the molecular machinery through which resting spinal microglia transform into an activated state that drives neuropathic pain.  相似文献   

7.
Increasing evidence supports the notion that spinal cord microglia activation plays a causal role in the development of neuropathic pain after peripheral nerve injury; yet the mechanisms for microglia activation remain elusive. Here, we provide evidence that NADPH oxidase 2 (Nox2)-derived ROS production plays a critical role in nerve injury-induced spinal cord microglia activation and subsequent pain hypersensitivity. Nox2 expression was induced in dorsal horn microglia immediately after L5 spinal nerve transection (SNT). Studies using Nox2-deficient mice show that Nox2 is required for SNT-induced ROS generation, microglia activation, and proinflammatory cytokine expression in the spinal cord. SNT-induced mechanical allodynia and thermal hyperalgesia were similarly attenuated in Nox2-deficient mice. In addition, reducing microglial ROS level via intrathecal sulforaphane administration attenuated mechanical allodynia and thermal hyperalgesia in SNT-injured mice. Sulforaphane also inhibited SNT-induced proinflammatory gene expression in microglia, and studies using primary microglia indicate that ROS generation is required for proinflammatory gene expression in microglia. These studies delineate a pathway involving nerve damage leading to microglial Nox2-generated ROS, resulting in the expression of proinflammatory cytokines that are involved in the initiation of neuropathic pain.  相似文献   

8.
Host cell death induced by Entamoeba histolytica is an important mechanism for both host defence and microbial immune evasion during human amoebiasis. However, the signalling pathways underlying cell death induced by E. histolytica are not fully understood. This study investigated the involvement of the protein tyrosine phosphatases (PTPs) SHP‐1 and SHP‐2 in the dephosphorylation associated with E. histolytica‐induced host cell death. Incubation with E. histolytica resulted in a marked decrease in protein tyrosine phosphorylation levels and degradation of SHP‐1 or SHP‐2 in Jurkat cells. Pre‐treatment of cells with a calpain inhibitor, calpeptin, impeded the amoeba‐induced dephosporylation and cleavage of SHP‐1 or SHP‐2. Additionally, inhibition of PTPs with phenylarsine oxide (PAO) attenuated Entamoeba‐induced dephosphorylation and DNA fragmentation in Jurkat T cells. These results suggest that calpain‐dependent cleavage of SHP‐1 and SHP‐2 may contribute to protein tyrosine dephosphorylation in Jurkat T cell death induced by E. histolytica.  相似文献   

9.
10.
Characterization of the melatonin (MLT) biosynthesis pathway in plants is still limited. Additionally, a metabolomic analysis of MLT biosynthesis in plants is still a challenge due to analyte structural and chemical diversity, low analyte abundances, and plant matrix complexities. Herein, a sensitive liquid chromatography‐mass spectrometry (LC‐MS) method enabling the simultaneous determination of seven plant MLT biosynthetic metabolites was developed. In the proposed strategy, the targeted metabolites, which included tryptophan (Trp), tryptamine (TAM), 5‐hydroxytryptophan (5HTP), serotonin (5HT), N‐acetylserotonin (NAS), 5‐methoxytryptamine (5MT), and MLT, were purified from plant extracts using a one‐step dispersive solid‐phase extraction (DSPE). The samples were then chemically labeled with dansyl chloride (DNS‐Cl), followed by analysis using LC‐MS. The limit of detection (LOD) values ranged from 0.03 to 1.36 pg/mL and presented a 22‐ to 469‐fold decrease when compared to the unlabeled metabolites. Due to the high sensitivity of the proposed method, the consumption of plant materials was reduced to 10 mg FW. Ultimately, the established method was utilized to examine the distributions of MLT and its intermediates in rice shoots and roots with or without cadmium (Cd) stress. The results suggested that under normal condition, MLT may also be generated via a Trp/TAM/5HT/5MT/MLT path (Pathway II) in addition to the previously reported Trp/TAM/5HT/NAS/MLT path (Pathway I), although Pathway I was shown to be dominant. During Cd stress, MLT was also shown to be produced through these two pathways, with Pathway II shown to be dominant in rice shoots and roots.  相似文献   

11.
The maintenance of glucose homeostasis is a complex process in which the insulin signalling pathway plays a major role. Disruption of insulin‐regulated glucose homeostasis is frequently observed in chronic hepatitis C (CHC) infection and might potentially contribute to type 2 diabetes mellitus (T2DM) development. Presently, the mechanism that links HCV infection to insulin resistance remains unclear. Previously, we have reported that HCV protein expression in HCV transgenic mice (B6HCV) leads to an overexpression of protein phosphatase 2A (PP2A) through an ER stress response. In the present work, we describe an association of FoxO1 hypophosphorylation and upregulation of both PGC‐1α and G6Pase to phenotypic hyperglycaemia and insulin resistance in B6HCV mice. In vitro, we observed that PGC1α is concomitantly induced with PP2A. Moreover, we show that the enhanced PP2A expression is sufficient to inhibit insulin‐induced FoxO1 phosphorylation via blockade of insulin‐mediated Akt activation or/and through direct association and dephosphorylation of pS‐FoxO1. Consequently, we found that the gluconeogenic gene glucose‐6‐phosphatase is upregulated. These observations were confirmed in liver biopsies obtained from CHC patients. In summary, our results show that HCV‐mediated upregulation of PP2A catalytic subunit alters signalling pathways that control hepatic glucose homeostasis by inhibiting Akt and dephosphorylation of FoxO1.  相似文献   

12.
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.  相似文献   

13.
The functions of melatonin in preovulatory fluid remain elusive. In the current study, we observed that the extremely high level of expression of MT1 in mice granulosa cells was rapidly induced by hCG (equivalent LH) within 2 hours and this was referred as MT1 surge. In cumulus cells, serotonin N‐acetyltransferase (SNAT) was also upregulated by hCG and led to elevated melatonin levels in ovarian follicle fluid. Melatonin application before MT1 surge significantly promoted embryo implantation, and this was probably attributed to a rise in progesterone levels in the serum. The mechanistic studies indicated that melatonin/MT1 (MLT/MT1) signaling remarkably improved the expression of corpus luteum marker genes, that is, Akr1c18 and Cyp11a1. High‐throughput sequencing results suggested that extracellular matrix (ECM) receptor interaction, focal adhesion, and activation of PI3K/Akt pathway which are involved in granulosa cell luteinization might mediate the actions of MLT/MT1 signal. In addition, this effect on luteinization was compared in different species. It was verified that high melatonin levels exist in serum at estrum of cows and help to improve the first estrus fecundation rate. These results suggested that both melatonin and MT1 are involved in the downstream reaction of hCG (LH) and they play important roles in luteinization. These findings provide the novel information on the physiology of melatonin in animal reproduction.  相似文献   

14.
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16.
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 Ca2+/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.  相似文献   

17.
Defective regeneration of small‐for‐size (SFS ) liver remnants and partial grafts remains a key limiting factor in the application of liver surgery and transplantation. Exogenous melatonin (MLT ) has protective effects on hepatic ischemia‐reperfusion injury (IRI ), but its influence on graft regeneration is unknown. The aim of the study is to investigate the role of MLT in IRI and graft regeneration in settings of partial liver transplantation. We established three mouse models to study hepatic IRI and regeneration associated with partial liver transplantation: (I) IR +PH group: 60 minutes liver ischemia (IR ) plus 2/3 hepatectomy (PH ); (II ) IR +exPH group: 60 minutes liver IR plus extended hepatectomy (exPH ) associated with the SFS syndrome; (III ) SFS ‐LT group: Arterialized 30% SFS liver transplant. Each group was divided into MLT or vehicle‐treated subgroups. Hepatic injury, inflammatory signatures, liver regeneration, and animal survival rates were assessed. MLT reduced liver injury, enhanced liver regeneration, and promoted interleukin (IL ) 6, IL 10, and tumor necrosis factor‐α release by infiltrating, inflammatory Ly6C+ F4/80+ monocytes in the IR +PH group. MLT ‐induced IL 6 significantly improved hepatic microcirculation and survival in the IR +exPH model. In the SFS ‐LT group, MLT promoted graft regeneration and increased recipient survival along with increased IL 6/GP 130‐STAT 3 signaling. In IL 6 ?/? mice, MLT failed to promote liver recovery, which could be restored through recombinant IL 6. In the IR +exPH and SFS ‐LT groups, inhibition of the IL 6 co‐receptor GP 130 through SC 144 abolished the beneficial effects of MLT . MLT ameliorates SFS liver graft IRI and restores regeneration through monocyte‐released IL 6 and downstream IL 6/GP 130‐STAT 3 signaling.  相似文献   

18.
19.
Epstein‐Barr virus (EBV) latent proteins exert anti‐apoptotic effects on EBV‐transformed lymphoid cells by down‐regulating BCL2L11 (BIM), CDKN2A (p16INK4A) and CDKN1A (p21WAF1). However, the potential therapeutic effects of targeting these anti‐apoptotic mechanisms remain unexplored. Here, we tested both in vitro and in vivo effects of the combination of histone deacetylase (HDAC) and proteasome inhibitors on the apoptosis of six endemic Burkitt lymphoma (BL) lines of different latency patterns (types I and III and Wp‐restricted) and three lymphoblastoid cell lines (LCLs). We found that the combination of HDAC and proteasome inhibitors (e.g. SAHA/bortezomib) synergistically induced the killing of Wp‐restricted and latency III BL and LCLs but not latency I BL cells. The synergistic killing was due to apoptosis, as evidenced by the high percentage of annexin V positivity and strong cleavage of PARP1 (PARP) and CASP3 (caspase‐3). Concomitantly, SAHA/bortezomib up‐regulated the expression of CDKN2A and CDKN1A but did not affect the level of BCL2L11 or BHRF1 (viral homologue of BCL2). The apoptotic effects were dependent on reactive oxygen species generation. Furthermore, SAHA/bortezomib suppressed the growth of Wp‐restricted BL xenografts in nude mice. This study provides the rationale to test the novel application of SAHA/bortezomib on the treatment of EBV‐associated Wp‐restricted BL and post‐transplant lymphoproliferative disorder.  相似文献   

20.
Hyperglycemia is a representative hallmark and risk factor for diabetes mellitus (DM) and is closely linked to DM‐associated neuronal cell death. Previous investigators reported on a genome‐wide association study and showed relationships between DM and melatonin receptor (MT), highlighting the role of MT signaling by assessing melatonin in DM. However, the role of MT signaling in DM pathogenesis is unclear. Therefore, we investigated the role of mitophagy regulators in high glucose‐induced neuronal cell death and the effect of melatonin against high glucose‐induced mitophagy regulators in neuronal cells. In our results, high glucose significantly increased PTEN‐induced putative kinase 1 (PINK1) and LC‐3B expressions; as well it decreased cytochrome c oxidase subunit 4 expression and Mitotracker? fluorescence intensity. Silencing of PINK1 induced mitochondrial reactive oxygen species (ROS) accumulation and mitochondrial membrane potential impairment, increased expressions of cleaved caspases, and increased the number of annexin V‐positive cells. In addition, high glucose‐stimulated melatonin receptor 1B (MTNR1B) mRNA and PINK1 expressions were reversed by ROS scavenger N‐acetyl cysteine pretreatment. Upregulation of PINK1 expression in neuronal cells is suppressed by pretreatment with MT2 receptor‐specific inhibitor 4‐P‐PDOT. We further showed melatonin stimulated Akt phosphorylation, which was followed by nuclear factor kappa‐light‐chain‐enhancer of activated B cells (NF‐κB) phosphorylation and nuclear translocation. Silencing of PINK1 expression abolished melatonin‐regulated mitochondrial ROS production, cleaved caspase‐3 and caspase‐9 expressions, and the number of annexin V‐positive cells. In conclusion, we have demonstrated the melatonin stimulates PINK1 expression via an MT2/Akt/NF‐κB pathway, and such stimulation is important for the prevention of neuronal cell apoptosis under high glucose conditions.  相似文献   

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