Megakaryocytic Leukemia 1 Bridges Epigenetic Activation of NADPH Oxidase in Macrophages to Cardiac Ischemia-Reperfusion Injury

Abstract

-Excessive accumulation of reactive oxygen species (ROS), catalyzed by the NADPH oxidases (NOX), is involved in the pathogenesis of ischemia-reperfusion (I/R) injury. The underlying epigenetic mechanism remains elusive. -We evaluated the potential role of megakaryocytic leukemia 1, or MKL1, as a bridge linking epigenetic activation of NOX to ROS production and cardiac ischemia-reperfusion injury. -Following I/R injury, MKL1 deficient (KO) mice exhibited smaller myocardial infarction along with improved heart function compared to wild type (WT) littermates. Similarly, pharmaceutical inhibition of MKL1 with CCG-1423 also attenuated myocardial infarction and improved heart function in mice. Amelioration of I/R injury as a result of MKL1 deletion or inhibition was accompanied by reduced ROS and In response to I/R, MKL1 levels were specifically elevated in macrophages, but not in cardiomyocytes, in the heart. Of note, macrophage-specific deletion (MφcKO), instead of cardiomyocyte-restricted ablation (CMcKO), of MKL1 in mice led to similar improvements of infarct size, heart function, and myocardial ROS generation. Reporter assay and ChIP assay revealed that MKL1 directly bound to the promoters of NADPH oxidase (NOX) genes to activate NOX transcription. Mechanistically, MKL1 recruited the histone acetyltransferase MOF to modify the chromatin structure surrounding the NOX promoters. Knockdown of MOF in macrophages blocked hyoxia/re-oxygenation-induced NOX transactivation and ROS accumulation. Of importance, pharmaceutical inhibition of MOF with MG-149 significantly down-regulated NOX1/NOX4 expression, dampened ROS production, and normalized myocardial function in mice exposed to I/R injury. Finally, administration of a specific NOX1/4 inhibitor GKT137831 dampened ROS generation and rescued heart function following I/R in mice. -Our data delineate an MKL1-MOF-NOX axis in macrophages that contributes to I/R injury and as such have provided novel therapeutic targets in the treatment of ischemic heart disease.

MKL1 mediates TGF‐β‐induced CTGF transcription to promote renal fibrosis

“…It was also observed that MKL1 overexpression activated the CTGF promoter by 3.2× and TGF‐β treatment activated the CTGF promoter by 1.7×; combined, MKL1 overexpression and TGF treatment synergistically activated the CTGF promoter by 4.8× (Figure b). On the contrary, a dominant negative (DN) form of MKL1, which lacks the trans‐activation domain (Yu et al, ), diminished the activation of the CTGF promoter by TGF‐β treatment by 37% (Figure c). It was noteworthy that when a conserved SMAD3 binding site within the CTGF promoter was mutated, MKL1 was unable to activate the CTGF promoter (Figure d), which indicated that MKL1 binding to the CTGF promoter might depend on SMAD3.…”

“…Megakaryocytic leukemia 1 (MKL1) is a transcriptional modulator known to bridge cytoskeletal reshuffling to stress response. MKL1 orchestrates a myriad of cellular responses to environmental and intrinsic stimuli including inflammation (Yu et al, ; Yu, Fang et al, ), oxidative stress (Yu et al, ), aging (Petrini et al, ), hypoxia (D. Chen et al, ; Yang et al, ), and mechanoresponse (Kuwahara et al, ). Mounting evidence portrays MKL1 as a key regulator of myofibroblast maturation and fibrogenesis (Small, ).…”

Forkhead transcription factor FOXO3a mediates interferon‐γ‐induced MHC II transcription in macrophages

“…Chromatin immunoprecipitation (ChIP) assays were performed essentially as described previously . In brief, chromatin in control and treated cells was cross‐linked with 1% formaldehyde.…”

“…Chromatin immunoprecipitation (ChIP) assays were performed essentially as described previously. [34][35][36][37][38][39][40][41][42] In brief, chromatin in control and treated cells was cross-linked with 1% formaldehyde. Cells were incubated in lysis buffer (150 mM NaCl, 25 mM Tris-HCl pH 7Á5, 1% Triton-X-100, 0Á1% SDS, 0Á5% deoxycholate) supplemented with protease inhibitor tablet and phenylmethylsulphonyl fluoride.…”

Protection From Cardiac Ischemia-Reperfusion Injury by Epigenetic Regulation of NADPH Oxidase

“…This issue of the journal contains a detailed examination of the role of MKL1 in the myocardial response to IR injury 11 . Mice lacking MKL1 were protected against IR injury, manifesting smaller infarcts and better cardiac function measured by echocardiography after 45 minutes of coronary artery occlusion followed by 24 hours reperfusion 11 .…”