Single-Molecule Detection of H2O2 Mediating Angiogenic Redox Signaling on Fluorescent Single-Walled Carbon Nanotube Array

Abstract

Reactive oxygen species, specifically hydrogen peroxide (H2O2), activate signal transduction pathways during angiogenesis and therefore play an Important role in physiological development as well as various pathophysiologies. Herein, we utilize a near-Infrared fluorescent single-walled carbon nanotube (SWNT) sensor array to measure the single-molecule efflux of H2O2 from human umbilical vein endothelial cells (HUVEC) in response to angiogenic stimulation. Two angiogenic agents were investigated: the pro-angiogenic cytokine, vascular endothelial growth factor A (VEGF-A) and the recently identified inorganic pro-angiogenic factor, europium(III) hydroxide in nanorod form. The nanosensor array consists of a SWNT embedded within a collagen matrix that exhibits high selectivity and sensitivity to single molecules of H2O2. A calibration from 123 to 400 nM quantifies the production of H2O2 at nanomolar concentration in HUVEC with 1 s temporal and 300 nm spatial resolutions. We find that the production of H2O2 following VEGF stimulation is elevated outside of HUVEC, but not for stimulation via nanorods, while increased generation is observed in the cytoplasm for both cases, suggesting two distinct signaling pathways.