Pathophysiology of Macrophage Migration Inhibitory Factor on Electrical and Structural Remodelings in Heart Failure-Induced Atrial Arrhythmogenesis(3/3)

Background: Heart failure (HF) is important in the genesis of atrial fibrillation (AF). However, the mechanisms underlying HF-induced AF were not fully elucidated. HF changes atrial structural (more fibrosis) and electrical remodelings with calcium overload and enhanced triggered activity. Our previous studies found that HF increases pulmonary vein (PV) arrhythmogenesis with dysregulated calcium homeostasis and increased calcium spark. However, the molecular mechanisms and therapeutic strategy underlying HF-induced atrial electrical and structural remodeling are not clear. Macrophage migration inhibitory factor (MIF), a pleotropic inflammation cytokine, plays an important role in the pathophysiology of cardiovascular diseases. Elevated MIF levels were detected in the patients with HF and AF. The pro-inflammatory effects of MIF during atherosclerosis are mediated via a functional CXCR2/CD74 complex. Moreover, MIF plays a vital role in the pathophysiology of obesity, type II diabetes, and rheumatoid arthritis, which are important risk factors of AF genesis. Since MIF has direct effects on cardiac electrical activity, calcium regulation, and fibrosis formation, MIF may contribute to the genesis of AF in HF, obesity or diabetes. Epigenetic regulation through histone deacetylases (HDACs) are vital in the pathophysiology of HF and AF. Currently, our previous studies have found that HDAC inhibition can modulate cardiac calcium homeostasis and reducing PV arrhythmogenesis and AF inducibility. However, the role of MIF in HDAC activity is not clear. In this study, we will examine the role of MIF in HF-induced AF arrhythmogenesis on electrical and structural remodelings (1st year), analyze the effects of MIF signaling deficiency on PV or atrial arrhythmogenesis and AF genesis in HF animal model (2nd year) and also evaluate the effect of alter MIF signaling on the expression of HDAC and non-coding RNA clusters in atrial myocytes and cardiac fibroblasts (3rd year). Methods：In the first year, we will study the effects of administration of recombinant MIF protein (25, 100 ng/ml) or attenuate MIF signaling (e.q. MIF antagonist ISO-1 of 10 M, CD74 neutralized Ab of 10 g/ml, or CXCR2 neutralized Ab of 10 g/ml) on the electrical activity, ionic currents and calcium homeostasis by patch clamp, fluorescence and western blot in atrial myocytes (HL-1), and in cardiac fibroblast activity (migration, proliferation, and collagen production). In the second year, we will monitor the atrial arrhythmogenesis by ECG in vivo in rabbit with and without MIF recombinant protein (0.25 g/kg, three times a week for 2 weeks), and in HF rabbit model (coronary artery ligation for 2 weeks) with and without MIF antagonist (once a week for 2 weeks, ISO-1of 4mg/kg for MIF inhibition). In isolated hearts and tissue preparations, we will study the AF inducibility and atrial and PV arrhythmogenesis by monophasic action potential recordings, multielectrode array, and conventional electrolytes recordings, and study molecular and histological changes by histology and immunohisology and western blot. In the third year, we will study the HDAC activity and evaluate expression of long non-coding RNA (lncRNA) in HL-1 cells and cardiac fibroblasts with and without administration of MIF recombinant protein (100 ng/ml) or MIF antagonist ISO-1(10 M), and study whether HDAC inhibition (MPT0E014 of 1 M, MS-275 of 1 M, and MC-1568 of 1 M) or gain or lose-of functions of differentially expressed lncRNA will change the effect of MIF signaling on the functions of HL-1 or cardiac fibroblast. Preliminary results：MIF and ligands of CXCR2, CXCL8, and CXCL1 were highly expressed in atrium of previous established Isoproterenol-induced HF rat. MIF increased -SMA, collagen production in human cardiac fibroblast cell lines, and increased CaMKII activity in HL-1 cells. Furthermore, CXCL8 treatment could induce distinct patterns of calcium transients in the HL-1 cells. These findings suggest an important role of MIF in atrial and structural remodeling. Thus, further elucidation the role of MIF/CD74/CXCR2 signaling in the genesis of AF in HF is needed.