Intercellular Transport of Proteins Carried by Nanodiamonds through Membrane Nanotubes

Abstract

Membrane tunneling nanotubes (TNTs) are one of the intercellular communication pathways through which transport of proteins and other cytoplasmic components occurs. The intercellular transport is related to many diseases, such as human immunodeficiency virus (HIV) infection, prion protein infection, Parkinson's disease, and Alzheimer's disease. These nanotubes are also potentially useful as drug delivery channels for cancer therapy. Here, we apply fluorescent nanodiamond (FND) as a photostable tracker as well as a protein carrier to illustrate the transport events in TNTs of human cells. We first coated 100-nm FNDs with proteins including bovine serum albumin (BSA) and green fluorescent protein (GFP) by physical adsorption and then performed single particle tracking of the bioconjugates in the transient membrane connections by fluorescence microscopy. We observed stop-and-go and to-and-fro motions mediated by molecular motors and determined an average velocity of 0.31 μm/s for the active transport of protein-loaded FNDs trapped in the endosomal vehicles of human embryonic kidney cells (HEK293T). Quantitative analysis of the heterotypical transport between HEK293T and SH-SY5Y neuroblastoma cells by flow cytometry confirmed the formation of open-ended nanotubes between them, even though their TNTs differed in structural components. Our results demonstrate a nanotechnology-enabled new tool for long-term investigation of the intercellular transport of proteins, drugs, or vesicles at the single particle level for potential therapeutic applications.