Heterogeneity in Dental Pulp Stem Cells

Objectives: Dental pulp stem cells (DPSCs) are heterogeneous in nature and understanding how they behave in their native environment is fundamental to our knowledge of endogenous tissue repair processes. Development of a labelling technique that allows identification and tracking of sub-populations of DPSCs with different characteristics is crucial. Fluorescent nanoparticles, known as Qtracker705® (Qdots), label the late endosomes of cells. Fluorescent signal is diluted as Qdots are segregated between resulting daughter cells, permitting tracking of single cells according to their innate proliferative capacity. The overall aim was to develop a method to identify specific sub-populations of DPSCs based on a simple fluorescent readout.
Methods: After obtaining full ethical approval, DPSCs were isolated and cultured in α-MEM with 10% Foetal Bovine Serum, characterised and labelled with 4nM Qdots. Qdot signal attenuation was measured at 48, 72 and 144 hours using flow cytometry or time-series microscopy. Cells were sorted based on Qdot signal, RNA was extracted and biomarker analysis was performed for DPSC, pluripotency, neural crest and early neural markers to reveal specific sub-populations.
Results: DPSCs demonstrated variation in their initial Qdot loading and subsequent signal loss over time. Parallel variation existed in the expression of DPSC, pluripotency and neural markers between the specific sorted cell populations based on their evolved Qdot signal. Features of proliferation, cell-cell inheritance and cellular location were extracted by signal processing
Conclusions: The Qdot approach enables us to label (encode) cells so their cell division and position can be tracked using time series analysis. Sorting and enrichment of cells allowed us to identify unique sub-populations of cells within heterogeneous environments based on simple biophotonic principles. This is a unified approach where the fluorescent signal readout is embedded into the biological system, providing the scope for translation to ex vivo tooth repair models.