Nε-(carboxymethyl)lysine-modified albumin exacerbates doxorubicin-induced NOX5-dependent oxidative stress in endothelial cells.

INTRODUCTION

Doxorubicin (Dox), a potent chemotherapeutic agent, is associated with cardiovascular complications. Dox-induced cardiotoxicity involves accumulation of Nε-(carboxymethyl)lysine (CML) and activation of its receptor, RAGE (receptor for advanced glycation end-products) in the cardiac microvasculature, potentially driven by NADPH oxidase (NOX)-mediated oxidative stress. NOX5 is a critical contributor to endothelial dysfunction. But its role, alongside extracellular CML, in Dox-induced vascular dysfunction remains unclear. This study investigated the impact of Dox and CML-bovine serum albumin (CML-BSA) on endothelial oxidative stress, and the involvement of NOX5 therein.

MATERIALS AND

METHODS

Human umbilical vein endothelial cells (HUVECs) were treated with Dox and/or CML-BSA for 6h, with or without NOX5 inhibitor ML090. Cell viability was assessed by MTT assay. Reactive oxygen species (ROS), Nitrotyrosine (NT), NOX5, RAGE and CML were visualized and quantified using confocal immunofluorescence imaging and intensity analysis. Subcellular NOX5 localization was confirmed via immunofluorescence. Golgi apparatus morphology was analyzed using electron microscopy.

RESULTS

Dox induced endogenous CML generation and the cellular CML levels were increased further by CML-BSA. Dox significantly reduced cell viability, which was not affected further by CML-BSA. Combined Dox and CML-BSA treatment increased RAGE expression and nitrosative/oxidative stress, evidenced by increased ROS, NT, and NOX5 levels, which was suppressed by ML090. NOX5 localized primarily to the Golgi apparatus, which was fragmented after Dox exposure.

DISCUSSION AND

CONCLUSION

Extracellular CML-BSA exacerbated Dox-induced CML accumulation and NOX5-related oxidative stress in endothelial cells. These findings suggest elevated circulating CML may worsen Dox-induced cardiovascular toxicity, aligning with clinical observations in diabetic patients.