Mixed lineage kinase-3 prevents cardiac dysfunction and structural remodeling with pressure overload

Abstract

Myocardial hypertrophy is an independent risk factor for heart failure (HF), yet the mechanisms underlying pathological cardiomyocyte growth are incompletely understood. The c-Jun NH2-terminal kinase (JNK) signaling cascade modulates cardiac hypertrophic remodeling, but the upstream factors regulating myocardial JNK activity remain unclear. In this study, we sought to identify JNK-activating molecules as novel regulators of cardiac remodeling in HF. We investigated mixed lineage kinase-3 (MLK3), a master regulator of upstream JNK-activating kinases, whose role in the remodeling process had not previously been studied. We observed increased MLK3 protein expression in myocardium from patients with nonischemic and hypertrophic cardiomyopathy and in hearts of mice subjected to transverse aortic constriction (TAC). Mice with genetic deletion of MLK3 (MLK3−/−) exhibited baseline cardiac hypertrophy with preserved cardiac function. MLK3−/− mice subjected to chronic left ventricular (LV) pressure overload (TAC, 4 wk) developed worsened cardiac dysfunction and increased LV chamber size compared with MLK3+/+ littermates ( n = 8). LV mass, pathological markers of hypertrophy ( Nppa, Nppb), and cardiomyocyte size were elevated in MLK3−/− TAC hearts. Phosphorylation of JNK, but not other MAPK pathways, was selectively impaired in MLK3−/− TAC hearts. In adult rat cardiomyocytes, pharmacological MLK3 kinase inhibition using URMC-099 blocked JNK phosphorylation induced by neurohormonal agents and oxidants. Sustained URMC-099 exposure induced cardiomyocyte hypertrophy. These data demonstrate that MLK3 prevents adverse cardiac remodeling in the setting of pressure overload. Mechanistically, MLK3 activates JNK, which in turn opposes cardiomyocyte hypertrophy. These results support modulation of MLK3 as a potential therapeutic approach in HF. NEW & NOTEWORTHY Here, we identified a role for mixed lineage kinase-3 (MLK3) as a novel antihypertrophic and antiremodeling molecule in response to cardiac pressure overload. MLK3 regulates phosphorylation of the stress-responsive JNK kinase in response to pressure overload and in cultured cardiomyocytes stimulated with hypertrophic agonists and oxidants. This study reveals MLK3-JNK signaling as a novel cardioprotective signaling axis in the setting of pressure overload.

Pyroptosis and ferroptosis induced by mixed lineage kinase 3 (MLK3) signaling in cardiomyocytes are essential for myocardial fibrosis in response to pressure overload

“…CHF has a similar characteristic that the activation of fibroblasts will produce harmful effects on cardiac myocytes. Calamaras et al 42 reported increased MLK3 expression in human (see figure on previous page) Fig. 5 Inhibition of MLK3 improves cardiac function and inhibits ferroptosis at week 8 of TAC.…”

“…CHF has a similar characteristic that the activation of fibroblasts will produce harmful effects on cardiac myocytes. Calamaras et al 42 reported increased MLK3 expression in human patients with cardiomyopathy. Recently, a report suggested that MLK3-deficient mice are protected against diet-induced NASH and liver fibrosis 43 .…”

“…However, Calamaras et al 42 reported that MLK3 prevented adverse cardiac remodeling in the setting of pressure overload, which partly attributed to MLK3 regulated phosphorylation of the stress-responsive JNK kinase. They also reported that CM-derived MLK3 contributes to the increased MLK3 in HF, basal LV hypertrophy observed in MLK3 depletion mice, although with no changes in cardiac function and structure.…”

JNK and cardiometabolic dysfunction

“…The JNK kinases have been implicated in multiple parts of these pathways (Figure 4) involved in cardiomyocyte growth and fibrosis. JNK activation increases in the human failing heart [99], and MLK an upstream JNK activator expression becomes elevated in end-stage heart failure patients [100]. JNKs are thought to be involved in re-programing gene expression that, in part, results in hypertrophic gene expression through regulation of transcription factors such as NFATs, Stats, Creb, c-jun, c-fos, and Gata4.…”

“…CDC42 binds and induces autoactivation of another MAP3K, MLK3 [117]. MLK3 whole body knockout mice displayed baseline LVH with normal LV function [100]. In response to TAC, however, the MLK3 −/− mice developed more severe LV systolic and diastolic dysfunction, as well increased LV end diastolic pressures, indicating heart failure.…”