Dual action of macrophage miR-204 confines cyclosporine A-induced atherosclerosis.

BACKGROUND AND PURPOSE: Atherosclerosis induced by cyclosporine A (CsA), an inhibitor of the calcineurin/nuclear factor of activated T cells (NFAT) pathway, is a major concern after organ transplantation. However, the atherosclerotic mechanisms of CsA remain obscure. We previously demonstrated that calcineurin/NFAT signalling inhibition contributes to atherogenesis via suppressing microRNA-204 (miR-204) transcription. We therefore hypothesised that miR-204 is involved in the development of CsA-induced atherosclerosis. EXPERIMENTAL APPROACH: ApoE-/- mice with macrophage-miR-204 overexpression were generated to determine the effects of miR-204 on CsA-induced atherosclerosis. Luciferase reporter assays and chromatin immunoprecipitation sequencing were performed to explore the targets mediating miR-204 effects. KEY RESULTS: CsA alone did not significantly affect atherosclerotic lesions or serum lipid levels. However, it exacerbated high-fat diet-induced atherosclerosis and hyperlipidemia in C57BL/6J and ApoE-/- mice, respectively. miR-204 levels decreased in circulating monocytes and plaque lesions during CsA-induced atherosclerosis. The upregulation of miR-204 in macrophages inhibited CsA-induced atherosclerotic plaque formation but did not affect serum lipid levels. miR-204 limited the CsA-induced foam cell formation by reducing the expression of the scavenger receptors SR-BII and CD36. SR-BII was post-transcriptionally regulated by mature miR-204-5p via 3'-UTR targeting. Additionally, nuclear-localised miR-204-3p prevented the CsA-induced binding of Ago2 to the CD36 promoter, suppressing CD36 transcription. SR-BII or CD36 expression restoration dampened the beneficial effects of miR-204 on CsA-induced atherosclerosis. CONCLUSION AND IMPLICATIONS: Macrophage miR-204 ameliorates CsA-induced atherosclerosis, suggesting that miR-204 may be a potential target for the prevention and treatment of CsA-related atherosclerotic side effects.

Cardiac-specific deletion of BRG1 ameliorates ventricular arrhythmia in mice with myocardial infarction.

Malignant ventricular arrhythmia (VA) after myocardial infarction (MI) is mainly caused by myocardial electrophysiological remodeling. Brahma-related gene 1 (BRG1) is an ATPase catalytic subunit that belongs to a family of chromatin remodeling complexes called Switch/Sucrose Non-Fermentable Chromatin (SWI/SNF). BRG1 has been reported as a molecular chaperone, interacting with various transcription factors or proteins to regulate transcription in cardiac diseases. In this study, we investigated the potential role of BRG1 in ion channel remodeling and VA after ischemic infarction. Myocardial infarction (MI) mice were established by ligating the left anterior descending (LAD) coronary artery, and electrocardiogram (ECG) was monitored. Epicardial conduction of MI mouse heart was characterized in Langendorff-perfused hearts using epicardial optical voltage mapping. Patch-clamping analysis was conducted in single ventricular cardiomyocytes isolated from the mice. We showed that BRG1 expression in the border zone was progressively increased in the first week following MI. Cardiac-specific deletion of BRG1 by tail vein injection of AAV9-BRG1-shRNA significantly ameliorated susceptibility to electrical-induced VA and shortened QTc intervals in MI mice. BRG1 knockdown significantly enhanced conduction velocity (CV) and reversed the prolonged action potential duration in MI mouse heart. Moreover, BRG1 knockdown improved the decreased densities of Na+ current (INa) and transient outward potassium current (Ito), as well as the expression of Nav1.5 and Kv4.3 in the border zone of MI mouse hearts and in hypoxia-treated neonatal mouse ventricular cardiomyocytes. We revealed that MI increased the binding among BRG1, T-cell factor 4 (TCF4) and ß-catenin, forming a transcription complex, which suppressed the transcription activity of SCN5A and KCND3, thereby influencing the incidence of VA post-MI.

PFI-3 induces vasorelaxation with potency to reduce extracellular calcium influx in rat mesenteric artery.

Background: PFI-3 is a small-molecule inhibitor that targets the bromodomains (BRDs) of Brahma-related gene 1 (BRG1). This monomeric compound, which has high selectivity and potent cellular effects, has recently been developed. Although PFI-3 has been reported as a potential therapeutic agent targeting thrombomodulin, its role in the regulation of vascular function remains unknown. Therefore, we aimed to investigate the impact of PFI-3 on arterial vessel tone. Methods: A microvascular tension measurement device (DMT) was utilized to identify alterations in vascular tension within the mesenteric artery. To detect variations in cytosolic [Ca2+]i, a Fluo-3/AM fluorescent probe and fluorescence microscope were employed. Additionally, whole-cell patch clamp techniques were utilized to evaluate the activity of L-type voltage-dependent calcium channels (VDCCs) in cultured arterial smooth muscle cells (A10 cells). Results: PFI-3 exerted a dose-dependent relaxation effect on rat mesenteric arteries with both intact and denuded endothelium after phenylephrine (PE)- and high-K+-induced constriction. PFI-3-induced vasorelaxation was not affected by the presence of L-NAME/ODQ or K+ channel blockers (Gli/TEA). PFI-3 abolished Ca2+-induced contraction on endothelium-denuded mesenteric arteries preincubated by PE in Ca2+-free solution. Incubation with TG had no impact on PFI-3-induced vasorelaxation pre-contracted by PE. PFI-3 reduced Ca2+-induced contraction on endothelium-denuded mesenteric arteries pre-incubated by KCl (60 mM) in Ca2+-free solution. PFI-3 declined extracellular calcium influx in A10 cells detected by Fluo-3/AM fluorescent probe and fluorescence microscope. Furthermore, we observed that PFI-3 decreased the current densities of L-type VDCC by whole-cell patch clamp techniques. Conclusions: PFI-3 blunted PE and high K+-induced vasoconstriction independent of endothelium on rat mesenteric artery. The vasodilatory effect of PFI-3 may be attributed to its inhibition of VDCCs and receptor-operated calcium channels (ROCCs) on vascular smooth muscle cells (VSMCs).

MicroRNA­143­3p contributes to the regulation of pain responses in collagen­induced arthritis.

Patients with rheumatoid arthritis (RA) suffer from pain, which is associated with inflammation, peripheral and central pain processing, and joint structure damage. The aim of the present study was to investigate a key microRNA (miR) and its target genes that are involved in the pain responses of RA, and to clarify the mechanism of pain regulation. Collagen­induced arthritis (CIA) was induced in DBA/1 and C57BL/6 mice. The paw swelling, mechanical withdrawal threshold (MWT), thermal withdrawal latency (TWL), and expression levels of tumor necrosis factor (TNF)­α and prostaglandin (PG)E2 in the sera were investigated. Decreased MWT and TWL, and increased TNF­α and PGE2, in the CIA model group were observed in DBA/1 and C57BL/6 mice. DBA/1 mice exhibited greater hyperalgesia and higher levels of inflammatory mediators. miR­143­3p expression in the blood and the dorsal root ganglion (DRG) were detected, and low miR­143­3p expression was demonstrated in the blood and DRG tissue of CIA mice. The target genes of miR­143 were predicted and analyzed. A total of 1,305 genes were predicted and 55 pain­associated genes were obtained. Prostaglandin­endoperoxide synthase 2 (Ptgs2), MAS related GPR family member E (Mrgpre), prostaglandin D2 receptor and Tnf were selected as target genes of miR­143. DRG cells were cultured and transfected with miR­143­3p inhibitor or mimic. The expression of Mrgpre, Ptgs2 and Tnf was significantly inhibited following miR­143­3p mimic transfection, while the expression of Mrgpre, Ptgs2 and Tnf was increased following inhibitor transfection. Additionally, the expression of pain­associated genes in the DRG of mice was investigated and the expression of Ptgs2, Mrgpre and Tnf in the DRG of CIA mice was also significantly upregulated. These results revealed that CIA mice exhibited marked hyperalgesia and high levels of inflammatory pain mediators. Low expression of miR­143­3p negatively regulated the pain­associated target genes, including Mrgpre, Ptgs2 and Tnf, thereby affecting chronic inflammatory pain and neuropathic pain in RA.