Cell-assembled (Gd-DOTA)_i-triphenylphosphonium (TPP) nanoclusters as a T₂ contrast agent reveal in vivo fates of stem cell transplants

A simple and straightforward strategy for magnetic resonance imaging (MRI) of stem cell transplants in terms of their viability, migration and homing, and differentiation has been pursued over the years. Herein we couple Gd-DOTA with triphenylphosphonium (TPP) to yield small molecule (Gd-DOTA)_i-TPP (i = 1, 4) agents and show that labeling cells with (Gd-DOTA)_i-TPP via electroporation (EP) results in two distinct cellular distributions of (Gd-DOTA)_i-TPP: freely and evenly distributed in the cytosol or cell-assembled nanoclusters in the cytoplasm. The two distinct cellular distributions contribute in different ways to MRI signals in vitro and in vivo. Importantly, we present a detailed interpretation of MRI results based on the signal intensity equation and cellular longitudinal (T₁-) and transverse (T₂-) relaxation rates of water protons. We demonstrate that cell-assembled (Gd-DOTA)_i-TPP nanoclusters not only promote its intracellular retention time but also induce significant MRI signal reduction, which act as an excellent T₂ contrast agent and allows for unambiguous reporting of in vivo viability and migration of cell transplants under T₂-weighted MRI over a long period. Notably, (Gd-DOTA)_i-TPP agents released as a result of exocytosis or cell death induce signal enhancement in the surrounding tissue such that the labeled cells can be unambiguously discriminated from its host tissue. The labeling and imaging strategy provides abundant information on the in vivo fates of stem cell transplants. The strategy features a single contrast, single imaging mode with dual signal output.