Potential of fluorescent nano-carriers targeting VCAM-1 for early detection of senescent endothelial cells

Abstract

Endothelial senescence has been identified as an early event promoting the development of endothelial dysfunction (ED), a hallmark of vascular ageing and major cardiovascular diseases. In vivo, senescent endothelial cells (EC) appear initially at arterial sites at risk characterized by disturbed flow and low shear stress (i.e., bifurcations and curvatures) compared to those at low risk characterized by laminar flow and a high level of shear stress (i.e., aorta). The aim is to develop fluorescent nano-carriers (NC) to target a cell surface protein up-regulated in senescent ECs, for diagnostic purposes. NC with a lipid core containing a fluorescent probe (NR668 or Cy5.5 derivative) stabilized by a polymer scaffold and decorated with VCAM-1 antibodies (NC-VCAM-1) were applied to different models of EC senescence in cultured porcine coronary artery (premature senescence induced by angiotensin II and replicative senescence observed at passage P3). The fluorescence signal was visualized by confocal microscopy. Both models of EC senescence demonstrated higher levels of senescence-associated beta-galactosidase activity, senescence markers and VCAM-1 than healthy EC at P1. Incubation of either P3 EC or Ang II-treated P1 EC with fluorescent NC-VCAM-1 showed a higher level of fluorescence than P1 EC. The EC-related NC-VCAM-1 fluorescent signal is abolished by the previous incubation of EC with an antibody directed against VCAM-1, demonstrating the ability to target cell surface VCAM-1. The highest level of NC-VCAM-1 fluorescence was observed with NC with an average diameter below 100 nm and at 37 °C. The present findings indicate that fluorescent NC-VCAM-1 allow the detection of senescent EC. Thus, using fluorescent functionalized NC targeting a senescence-associated cell surface protein appears to be an attractive strategy to evaluate the burden of EC senescence, which promotes ED and, ultimately, cardiovascular events.