CD47 antibody promoted autophagy flow and inhibited apoptosis. oxidative stress, apoptosis, autophagy Introduction Cardiac hypertrophy, occurring during the clinical course of stress-induced heart failure [1-3], is usually characterized by an abnormal enlargement of the heart muscle resulting from increased myocyte cell size and abnormal proliferation of non-muscle cells [4]. Cardiac hypertrophy is usually controlled by a complex transmission transduction and gene regulatory network [5,6]. Recently, autophagy, a dynamic process involving the bulk degradation of cytoplasmic organelles and proteins, has been proven to participate in the pathogenesis of cardiac hypertrophy [7-12]. Isoproterenol (ISO), a nonselective b-adrenergic receptor (b-AR) agonist, has been widely used as a stimulus Kitasamycin for cardiac hypertrophy [13, 14] due to its convenience and rapidity in yielding reproducible results Kitasamycin [15]. The pathophysiological and morphological aberrations in the heart of myocardial necrotic Kitasamycin rat model are comparable with those in human myocardial infarction [16,17]. These b-adrenergic effects can result in cardiac infarct-like lesions in experimental animals [18], much like those in patients with myocardial infarction [19]. ISO-induced cardiac hypertrophy is usually accompanied by a significant decrease in autophagy activity [20-22]. CD47 is usually a widely expressed cell receptor [23] and activator of NADPH-oxidase-mediated reactive oxygen species (ROS) production in vascular cells [24]. Previous studies have recognized the role of CD47 in limiting blood flow [23] and metabolism [25], and suggested the additional benefits by therapeutic targeting of CD47 in myocardial infarction [26]. CD47 transcript has been reported to increase in ventricular biopsies from patients of left ventricular heart failure (LVHF) [27]. CD47-knockout mice displayed protection from transverse aortic constriction (TAC)-driven LVHF with enhanced cardiac functions, decreased cellular hypertrophy and fibrosis [26], and CD47 deficiency conferred cell survival through the activation of autophagic flux against radiation injury [28-30]. Moreover, CD47-blocking antibody has been used in research of various diseases including tumor and atherosclerosis [31-36]. So far, the specific effect of CD47 antibody on ISO-induced cardiac hypertrophy remains unclear. In this study, we investigated the effect of CD47 antibody on cardiac hypertrophy, fibrosis and myocyte apoptosis in mouse and cell models with ISO-induced cardiomyocyte hypertrophy. Materials and methods Animals Eighty C57/BL6 male mice, 8-10 weeks aged, weighing 22-28 g, were obtained from Nanjing University or college. The mice were housed in a Specific Pathogen Free (SPF) facility in the Animal Core Facility of Nanjing Medical University or college Kitasamycin under standard heat conditions with a 12 h light/dark cycle and fed ad libitum. All experimental protocols and animal handling procedures were performed according to the Guideline for the Care and Use of Laboratory Animals (National Academic Press, USA, 1996). The animal study was approved by the Institutional Animal Care and Use Committee of the Nanjing Medical University or college. Animal model TLR9 of cardiac hypertrophy Animal model of cardiac remodeling was established by intraperitoneally injection of ISO (I5627, Sigma-Aldrich, USA; 60 mg.kg-1.d-1, dissolved in sterile normal saline) once daily for 14 consecutive days [37]. The animals were then allowed to recover with free access to food and water. At 24 h after the Kitasamycin last administration, the mice were euthanized using intraperitoneal injection of sodium pentobarbital (50 mg/kg) under general anesthesia, and the heart tissues were dissected and weighed. The ratio of heart weight to body weight (relative excess weight of heart) was calculated for each group as index of cardiac hypertrophy. The blood and left ventricles were harvested for subsequent examination. Grouping and experimental protocol Two studies were performed. In study 1, a total of 80 mice were randomly allocated into 4 groups. Group 1 (IgG): Animals received IgG antibody (0.4 g/g body weight in 150 l sterile normal saline, i.p., sc-2026, Santa Cruz Biotechnology, USA). Group 2 (IgG+ISO): Animals received IgG antibody (0.4 g/g body weight in 150 l sterile normal saline, i.p.) treatment twice weekly for 4 weeks after injection of ISO (60 mg.kg-1.d-1 in sterile normal.
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