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The mitochondrial K-ATP channel opener diazoxide upregulates STIM1 and Orai1 via ROS and the MAPK pathway in adult rat cardiomyocytes.

ABSTRACT: Background:Openers of mitochondrial adenosine triphosphate-dependent potassium (mKATP) channels like diazoxide increase reactive oxygen species (ROS) production in cardiac cells and reduce Ca2+ elevations produced by ischemia-reperfusion, protecting the heart from damage. In this study we tested the hypothesis that opening mKATP channels regulates expression of the major components of store-operated Ca2+ entry (SOCE) STIM1 and Orai1. Results:Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) and western blot experiments showed that diazoxide increased expression of STIM1 and Orai1 at the mRNA and protein levels, respectively, in adult rat cardiomyocytes. Immunofluorescence analyses revealed that diazoxide also disrupted the striated distribution pattern of STIM1. These effects were prevented by the ROS scavenger N-acetyl cysteine (NAC), the mKATP channel antagonist 5-hydroxydecanoate (5-HD), or the protein synthesis inhibitor cycloheximide (CHX). Confocal microscopy revealed that diazoxide also led to nuclear translocation of the transcription factors c-Fos and NF?B, which was also blocked by NAC or 5-HD. Finally, the MAPK pathway inhibitor UO126 attenuated diazoxide-induced upregulation of STIM1 and Orai1 expression. Conclusions:Our results suggest that opening mitochondrial potassium ATP channels with diazoxide upregulates the expression of STIM1 and Orai1 by de novo synthesis by a mechanism that involves NFkB, c-Fos, and ROS via MAPK/ERK signaling.

SUBMITTER: Gavali JT

PROVIDER: S-EPMC7424994 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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The mitochondrial K-ATP channel opener diazoxide upregulates STIM1 and Orai1 via ROS and the MAPK pathway in adult rat cardiomyocytes.

Gavali Joice T JT Carrillo Elba D ED García María C MC Sánchez Jorge A JA

Cell & bioscience 20200813

<h4>Background</h4>Openers of mitochondrial adenosine triphosphate-dependent potassium (mKATP) channels like diazoxide increase reactive oxygen species (ROS) production in cardiac cells and reduce Ca<sup>2+</sup> elevations produced by ischemia-reperfusion, protecting the heart from damage. In this study we tested the hypothesis that opening mKATP channels regulates expression of the major components of store-operated Ca<sup>2+</sup> entry (SOCE) STIM1 and Orai1.<h4>Results</h4>Quantitative reve ...[more]

PMID: 32817784

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Project description:BackgroundMultiple Sclerosis (MS) is an acquired inflammatory demyelinating disorder of the central nervous system (CNS) and is the leading cause of nontraumatic disability among young adults. Activated microglial cells are important effectors of demyelination and neurodegeneration, by secreting cytokines and others neurotoxic agents. Previous studies have demonstrated that microglia expresses ATP-sensitive potassium (KATP) channels and its pharmacological activation can provide neuroprotective and anti-inflammatory effects. In this study, we have examined the effect of oral administration of KATP channel opener diazoxide on induced experimental autoimmune encephalomyelitis (EAE), a mouse model of MS.MethodsAnti-inflammatory effects of diazoxide were studied on lipopolysaccharide (LPS) and interferon gamma (IFNγ)-activated microglial cells. EAE was induced in C57BL/6J mice by immunization with myelin oligodendrocyte glycoprotein peptide (MOG₃₅₋₅₅). Mice were orally treated daily with diazoxide or vehicle for 15 days from the day of EAE symptom onset. Treatment starting at the same time as immunization was also assayed. Clinical signs of EAE were monitored and histological studies were performed to analyze tissue damage, demyelination, glial reactivity, axonal loss, neuronal preservation and lymphocyte infiltration.ResultsDiazoxide inhibited in vitro nitric oxide (NO), tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) production and inducible nitric oxide synthase (iNOS) expression by activated microglia without affecting cyclooxygenase-2 (COX-2) expression and phagocytosis. Oral treatment of mice with diazoxide ameliorated EAE clinical signs but did not prevent disease. Histological analysis demonstrated that diazoxide elicited a significant reduction in myelin and axonal loss accompanied by a decrease in glial activation and neuronal damage. Diazoxide did not affect the number of infiltrating lymphocytes positive for CD3 and CD20 in the spinal cord.ConclusionTaken together, these results demonstrate novel actions of diazoxide as an anti-inflammatory agent, which might contribute to its beneficial effects on EAE through neuroprotection. Treatment with this widely used and well-tolerated drug may be a useful therapeutic intervention in ameliorating MS disease.

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The mitochondrial K-ATP channel opener diazoxide upregulates STIM1 and Orai1 via ROS and the MAPK pathway in adult rat cardiomyocytes.

Publications

The mitochondrial K-ATP channel opener diazoxide upregulates STIM1 and Orai1 via ROS and the MAPK pathway in adult rat cardiomyocytes.

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