Intracellular delivery of liposome-encapsulated Finland trityl radicals for EPR oximetry

Abstract

Tetrathiatriarylmethyl (TAM, trityl) radicals have found wide applications in electron paramagnetic resonance (EPR) oximetry. However, the biomedical applications of TAM radicals were exclusively limited to an extracellular region owing to their negatively charged nature. The intracellular delivery of TAM radicals still remains a challenge. In the present work, we report a liposome-based method to encapsulate the water-soluble Finland trityl radical CT-03 for its intracellular delivery. Using the thin lipid film hydration method, CT-03-loaded liposomes were prepared from DSPC/cholesterol/DOTAP with a mean size of 167.5 ± 2.4 nm and a zeta potential of 27.8 ± 0.8 mV. EPR results showed that CT-03 was entrapped into the liposomes and still exhibited good oxygen (O₂) sensitivity. Moreover, CT-03 was successfully delivered into HepG2 cells and HUVECs using the CT-03-loaded liposomes. Importantly, the combination of the liposome-encapsulated radical CT-03 and the other TAM radical CT02-H enabled simultaneous measurements of the intracellular and extracellular O₂ concentrations and O₂ consumption rates in HepG2 cells. Our present study provides a new approach for intracellular delivery of TAM radicals and could significantly expand their biomedical applications.