Effect of Endostatin Overexpression on Angiotensin II-Induced Cardiac Hypertrophy in Rats
Cardiovascular diseases remain a leading cause of death worldwide, with cardiac remodeling being a critical pathophysiological process in their progression. Among the key changes observed during cardiac remodeling are myocardial fibrosis and hypertrophy. Cardiac hypertrophy, characterized by increased heart weight, cell size, and expression of genes such as atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP), is an adaptive response to pathological stimuli like myocardial infarction, hypertension, and activation of the renin-angiotensin system. Endostatin, a 20,000 C-terminal fragment derived from collagen XVIII, is known for its potent anti-angiogenic properties and involvement in various physiological and pathological processes, including sepsis, acute kidney injury, and fibrosis. However, its role in cardiac hypertrophy remains unclear. This study investigates whether endostatin overexpression can attenuate cardiac hypertrophy by inhibiting the cyclic adenosine monophosphate-protein kinase A (cAMP-PKA) signaling pathway.
The study was conducted both in vivo in rats and in vitro in primary neonatal rat cardiomyocytes treated with angiotensin II (Ang II) to model cardiac hypertrophy. Twenty-four male Sprague-Dawley rats were randomized into four groups: adenovirus (Ad)-green fluorescent protein (GFP), Ang II, Ad-endostatin, and Ang II + Ad-endostatin, with six rats in each group. After four weeks, all rats underwent transthoracic echocardiography, were weighed, and then sacrificed. Cardiac function was evaluated using echocardiography, cardiomyocyte size was assessed through hematoxylin-eosin (HE) staining, and levels of ANP and BNP were measured using quantitative reverse-transcription polymerase chain reaction (qRT-PCR) or Western blotting. PKA levels were evaluated by Western blotting, and cAMP levels were determined using an enzyme-linked immunosorbent assay (ELISA).
The results demonstrated that endostatin overexpression significantly reduced the increases in left ventricle (LV) mass, LV mass/body weight (BW), interventricular septal thickness (IVS) in diastole and systole, left ventricular posterior wall thickness (LVPW) in diastole and systole, heart weight (HW), HW/BW, and HW/tibial length in Ang II-treated rats. Additionally, endostatin overexpression reduced cardiomyocyte cross-sectional area expansion and decreased the levels of ANP and BNP in both Ang II-treated rats and primary neonatal rat cardiomyocytes. Furthermore, endostatin overexpression reduced the increase in cAMP and PKA levels in cardiomyocytes treated with Ang II. Treatment with cAMP reversed the effects of endostatin overexpression on ANP and BNP levels in cardiomyocytes induced by Ang II.
In the in vivo experiments, echocardiography revealed that Ang II administration increased LV mass, LV/BW, IVS in diastole and systole, and LVPW in diastole and systole. Endostatin overexpression attenuated these increases, demonstrating its protective effect against cardiac hypertrophy. HE staining showed that Ang II increased the size of cardiomyocytes, which was reduced by endostatin overexpression. The levels of ANP and BNP in rat hearts were elevated after Ang II administration but were significantly reduced by endostatin overexpression.
Masson staining indicated that cardiac fibrosis was significantly increased in Ang II-treated rats, and endostatin overexpression reduced this increase. In primary cardiomyocytes, Ang II administration increased cell size and the levels of ANP and BNP mRNA and protein. Endostatin overexpression reduced these increases, further supporting its role in attenuating cardiac hypertrophy.
The study also explored the underlying mechanisms, focusing on the cAMP-PKA signaling pathway. Ang II treatment increased cAMP and PKA levels in cardiomyocytes, while endostatin overexpression reduced these increases. The reduction in ANP and BNP levels by endostatin overexpression was reversed by cAMP administration, indicating that endostatin’s effects are mediated through the inhibition of the cAMP-PKA pathway.
These findings suggest that endostatin overexpression can alleviate cardiac hypertrophy by inhibiting the cAMP-PKA signaling pathway. The study provides valuable insights into the potential therapeutic applications of endostatin in managing cardiac hypertrophy and fibrosis, offering a new avenue for the treatment of cardiovascular diseases.
doi.org/10.1097/CM9.0000000000000513
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