Protective effects of Apelin-13 on nicotine-induced H9c2 cardiomyocyte apoptosis and oxidative stress.

INTRODUCTION

We aimed to explore the role of Apelin-13 in resisting oxidation, inflammation as well as apoptosis and its underlying mechanisms of action using a model of nicotine-induced H9c2 cardiomyocyte injury.

METHODS

H9c2 cardiomyocytes were randomly divided into control, nicotine, nicotine + Apelin-13, and Apelin-13 groups. Cell counting kit-8 assay was conducted to determine the cell viability. Interleukin (IL)-6, superoxide dismutase, tumor necrosis factor-alpha (TNF-α), glutathione peroxidase (GSH-Px), IL-β, catalase (CAT), IL-8, lactate dehydrogenase (LDH), and malondialdehyde (MDA) levels were examined. A 2',7'-dichlorodihydrofluorescein diacetate assay was conducted to measure the intracellular reactive oxygen species (ROS) level. The morphology of apoptotic cardiomyocytes was observed by 4',6-diamidino-2-phenylindole staining. Western blotting was employed to measure the protein expressions of apoptotic factors B-cell lymphoma-2 (Bcl-2) and Bcl-2-associated X (Bax). Apoptosis was quantified using Annexin V/propidium iodide staining.

RESULTS

Exposure of H9c2 cardiomyocytes to 10 μM nicotine significantly reduced cell viability and increased LDH release, oxidative stress (elevated MDA and ROS levels with decreased superoxide dismutase, GSH-Px, and CAT activities), pro-inflammatory cytokines (IL-6, TNF-α, IL-1β, IL-8), and apoptotic markers (increased Bax with decreased Bcl-2 expression, along with nuclear condensation) (p<0.05). In contrast, treatment with 2 μM Apelin-13 significantly alleviated these deleterious effects, enhancing cell viability, restoring antioxidant enzyme activities, reducing oxidative and inflammatory responses, and inhibiting apoptosis (p<0.05).

CONCLUSIONS

Nicotine induction increases the oxidative stress and apoptotic capacity of H9c2 cardiomyocytes, but Apelin-13 protects H9c2 cardiomyocytes against nicotine-induced apoptosis and oxidative stress.