Photobiomodulation of Mineralised Tissue Derived Cellular Responses

Previously, low-level laser therapy has been proposed to improve mineralised tissue healing however the cellular and molecular mechanisms involved in this response remain unclear. Objectives: To investigate the in vitro effect of diffusible red laser light on bone marrow and dental pulp cells. Methods: Bone marrow and pulp cells were derived from freshly sacrificed 6 week old rats. Bone marrow was obtained by flushing of femora with culture medium whilst pulpal cells were harvested from exposed pulp chambers. Cells were grown (passages 1-3) in standard tissue culture plasticware in alpha-MEM medium with 2mM L-Glutamine, 10% FCS and 1% penicillin/streptomycin. Cells were exposed to diffusible 660nm laser light (150-200mW) in a standardised manner. Cell counts were used to determine proliferative response, and RT-PCR was performed on oligo-dT reverse transcribed RNA (normalised to GAPDH). Changes in media temperature following laser exposure were determined using a sensitive (±0.01 °C) calibrated thermocouple. Results: Laser exposure resulted in a non-significant increase in cell adhesion for both cell types at 4 and 24h post exposure. Duration of laser light exposure (0-120s) indicated that pulpal cells exhibited optimal increases in cell number with 60s dose, 3/5-days post-exposure. Bone marrow cells demonstrated a more limited growth response. Transcriptional analyses for several growth factors, including TGF/BMP family members, heat shock proteins and mineralised tissue associated genes demonstrated their differential expression at 3-days post-exposure. There were no significant temperature changes detected within media exposed to laser light. Conclusion: Rapidly proliferating bone marrow cells demonstrated a more limited response to laser light compared with pulpal cells. Molecular changes induced by laser light may elicit cellular responses which contribute to wound repair and this is not temperature dependent.