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Histone Deacetylases Enhance Ca2+-Activated K? Channel KCa3.1 Expression in Murine Inflammatory CD4? T Cells.

ABSTRACT: The up-regulated expression of the Ca2+-activated K? channel KCa3.1 in inflammatory CD4? T cells has been implicated in the pathogenesis of inflammatory bowel disease (IBD) through the enhanced production of inflammatory cytokines, such as interferon-? (IFN-?). However, the underlying mechanisms have not yet been elucidated. The objective of the present study is to clarify the involvement of histone deacetylases (HDACs) in the up-regulation of KCa3.1 in the CD4? T cells of IBD model mice. The expression levels of KCa3.1 and its regulators, such as function-modifying molecules and transcription factors, were quantitated using a real-time polymerase chain reaction (PCR) assay, Western blotting, and depolarization responses, which were induced by the selective KCa3.1 blocker TRAM-34 (1 ?M) and were measured using a voltage-sensitive fluorescent dye imaging system. The treatment with 1 ?M vorinostat, a pan-HDAC inhibitor, for 24 h repressed the transcriptional expression of KCa3.1 in the splenic CD4? T cells of IBD model mice. Accordingly, TRAM-34-induced depolarization responses were significantly reduced. HDAC2 and HDAC3 were significantly up-regulated in the CD4? T cells of IBD model mice. The down-regulated expression of KCa3.1 was observed following treatments with the selective inhibitors of HDAC2 and HDAC3. The KCa3.1 K? channel regulates inflammatory cytokine production in CD4? T cells, mediating epigenetic modifications by HDAC2 and HDAC3.

SUBMITTER: Matsui M

PROVIDER: S-EPMC6213394 | biostudies-literature | 2018 Sep

REPOSITORIES: biostudies-literature

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Project description:In inflammatory arthritis, the dysregulation of osteoclast activity by proinflammatory cytokines, including TNF, interferes with bone remodeling during inflammation through Ca2+-dependent mechanisms causing pathological bone loss. Ca2+-dependent CREB/c-fos activation via Ca2+-calmodulin kinase IV (CaMKIV) induces transcriptional regulation of osteoclast-specific genes via NFATc1, which facilitate bone resorption. In leukocytes, Ca2+ regulation of NFAT-dependent gene expression oftentimes involves the activity of the Ca2+-activated K+ channel KCa3.1. In this study, we evaluate KCa3.1 as a modulator of Ca2+-induced NFAT-dependent osteoclast differentiation in inflammatory bone loss. Microarray analysis of receptor activator of NF-?B ligand (RANKL)-activated murine bone marrow macrophage (BMM) cultures revealed unique upregulation of KCa3.1 during osteoclastogenesis. The expression of KCa3.1 in vivo was confirmed by immunofluorescence staining on multinucleated cells at the bone surface of inflamed mouse joints. Experiments on in vitro BMM cultures revealed that KCa3.1-/- and TRAM-34 treatment significantly reduced the expression of osteoclast-specific genes (p < 0.05) alongside decreased osteoclast formation (p < 0.0001) in inflammatory (RANKL+TNF) and noninflammatory (RANKL) conditions. In particular, live cell Ca2+ imaging and Western blot analysis showed that TRAM-34 pretreatment decreased transient RANKL-induced Ca2+ amplitudes in BMMs by ?50% (p < 0.0001) and prevented phosphorylation of CaMKIV. KCa3.1-/- reduced RANKL+/-TNF-stimulated phosphorylation of CREB and expression of c-fos in BMMs (p < 0.01), culminating in decreased NFATc1 protein expression and transcriptional activity (p < 0.01). These data indicate that KCa3.1 regulates Ca2+-dependent NFATc1 expression via CaMKIV/CREB during inflammatory osteoclastogenesis in the presence of TNF, corroborating its role as a target candidate for the treatment of bone erosion in inflammatory arthritis.

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Histone Deacetylases Enhance Ca2+-Activated K? Channel KCa3.1 Expression in Murine Inflammatory CD4? T Cells.

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