Apelin-13 attenuates doxorubicin-induced myocardial injury via the PI3K/AKT and ERK/MAPK signaling pathways.

Abstract

Background: Anthracyclines play a crucial role in the chemotherapy of various malignant tumors; however, they may cause myocardial injury, which limits their clinical application. Therefore, it is of great importance to investigate drugs for the prevention and treatment of anthracycline-induced myocardial injury. The aim of this study is to evaluate the cardioprotective effects of Apelin-13 in a murine model of doxorubicin (DOX)-induced myocardial injury and to elucidate its potential underlying mechanisms.

Methods: Male C57BL/6 mice (8 weeks old) were randomly assigned to four groups, with all mice received intraperitoneal injections: Control (normal saline), DOX (with a cumulative dose of 20 mg), Apelin-13 (Apelin-13 0.1 μmol/kg/d), and Apelin-13 + DOX (Apelin-13 0.1 μmol/kg/d; with a cumulative dose of 20 mg). Cardiac function was assessed using echocardiography. Histopathological changes were assessed via hematoxylin and eosin (HE) staining, and myocardial fibrosis was evaluated with Masson's trichrome staining. Immunohistochemical analysis and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling staining were conducted to assess protein expression and cardiomyocyte apoptosis, respectively. Protein expression levels were further validated by western blotting (WB). In vitro, rat H9C2 cardiomyocytes were cultured and divided into four groups. Control, Apelin-13 (Apelin-13 1 μM), DOX (DOX 2 μM), and Apelin-13 + DOX group (first treated with Apelin-13 1 μM, followed by DOX 2 μM after 1-2 hours). Cell viability was measured using the Cell Counting Kit-8 (CCK-8) assay. Flow cytometry was employed to quantify apoptosis, and WB was used to analyze signaling pathway components.

Results: In mice studies, no mice died during or after the drug intervention period. It was found that Apelin-13 could significantly increase left ventricular ejection fraction (LVEF) and fractional shortening (FS) (P<0.05) and decrease left ventricular end-diastolic diameter (LVEDD) and left ventricular end-systolic diameter (LVESD) (P<0.05) in mice with DOX-induced cardiomyopathy, as well as alleviate the disordered arrangement of myocardial cells and alleviate the reduction in cross-sectional area (CSA) induced by DOX. Masson staining and immunohistochemistry showed that Apelin-13 could alleviate myocardial fibrosis. WB indicated that Apelin-13 could reduce apoptosis induced by DOX and change the effects of DOX on the transforming growth factor-β (TGF-β)/mothers against decapentaplegic homolog (Smad) proteins. Further research confirmed that the protective effect of Apelin-13 on DOX-induced cardiomyopathy was related to the upregulation of P-ERK and P-AKT. Similarly, in H9C2 studies, it was found that Apelin-13 could reduce DOX-induced apoptosis, increase cell survival rate, and alleviate the decrease in P-ERK and P-AKT induced by DOX.

Conclusions: Apelin-13 exerted a protective effect against DOX-induced myocardial injury in both C57BL/6 mice and H9C2 cardiomyocytes. This cardio protection was mediated, at least in part, through modulation of the phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT) and extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) signaling pathways.