Tracking Myoblasts in the Development of Laryngeal Cellular Engineering Using Fluorescence Nanoparticles (“Quantum Dots”)

Abstract

Objectives: Stem-cell based techniques have previously been successfully used to replace patients’ tracheae, but using similar techniques to replace an entire larynx remains challenging. Additionally, non-immunogenic labelling techniques are required to track cells in vivo to determine the relative contribution of the implanted/donor stem cells with respect to the regeneration seen. Methods: We have developed and synthesized two quantum-dots (QDs) using gold and mercury-cadmium-telluride (HgCdTe) that emit at near-infrared wavelength (>700nm). Labelled C2C12 myoblasts were characterized with both QDs at different concentrations and incubation times. Labelled cells were visualized by confocal microscopy and transmission electron microscopy (TEM). Toxicity in vitro was determined using Alamar Blue assays and Ki67-immunofluorescence. Myosin heavy chain and MyoD-immunofluorescence were used to determine the effect of QDs on cellular differentiation. Results: We show that myoblasts can be optimally labelled with 800nm-emitting gold-nanoparticles that are non-toxic and do not affect cell proliferation or differentiation in vitro ( P > 0.05). Some toxicity-related effects were seen with HgCdTe QDs, which led to smaller, less-mature myofibers ( P < 0.05). On TEM, QDs were seen early on adhering to cell membranes and within phagosomes at later time points. Conclusions: We present new data demonstrating that cells may be optimally labelled with QDs for tracking cells in organ development using tissue-engineering approaches. Labelling with gold-nanoparticles is associated with minimal associated toxicity and may provide an immunomodulatory influence, thereby preventing cell rejection. Such cell-labelling techniques may be useful in tracking cells in real time in vivo to better understand the contribution of such cells towards regeneration.