Date of Award
5-2025
Document Type
Thesis
Degree Name
Master of Science (MS)
Department
Chemistry and Chemical Engineering
First Advisor
Vipuil Kishore
Second Advisor
James R. Brenner
Third Advisor
Nasri Nesnas
Fourth Advisor
Jessica Smeltz
Abstract
Photobiomodulation (PBM) is a low-intensity light therapy that uses non-ionizing forms of energy to induce a directed cellular response. Prior work has investigated the effects of blue light exposure at energy densities ranging from 1 J/cm2 to 10 J/cm2 on osteogenic differentiation of human gingival mesenchymal stem cells (hGMSCs), human dental pulp stem cells (hDPSCs), and stem cells from the apical papilla (SCAPs) and shown promising outcomes at 4 J/cm2 and then having a detrimental impact at higher energy densities. One key caveat in prior studies is that the blue light energy density was modulated by changing the light exposure time, and hence it cannot be determined if the observed effects of blue light-mediated PBM on osteogenesis are due to different light energy densities, exposure times, or a combination of both. In addition, prior PBM studies employed culture media composed of dexamethasone (a synthetic glucocorticoid and stimulant of osteogenesis) and hence, the effects of PBM on osteogenesis in the absence of dexamethasone are unknown. The goal of this study is to investigate the effects of different blue light energy densities (i.e., 1 J/cm2, 2 J/cm2, 4J/cm2, and 8 J/cm2) while keeping the exposure time constant at 1 min. Human osteosarcoma cells (i.e., Saos-2) were cultured in RPMI growth media supplemented with 15% FBS, 10 mM β-glycerophosphate, 50 µg/ml ascorbic acid, and 1% penicillin/streptomycin (without dexamethasone) and the effects of different blue light energy densities on Saos-2 cell viability, differentiation, and mineralization were investigated. Live-Dead and Alamar Blue assays showed that Saos-2 cell viability and metabolic activity is maintained in all light exposure conditions. Results from ALP assay showed a significant increase in ALP activity upon exposure to 1 J/cm2, 2 J/cm2, and 4 J/cm2 compared to the dark and 8 J/cm2 conditions. Qualitative assessment of the ARS-stained images shows that 1 J/cm2, 2 J/cm2, and 4 J/cm2 demonstrate visibly higher amount of cell-mediated mineral-like deposition compared to the dark and 8 J/cm2 conditions. These results are corroborated by tetracycline staining (visual indicator of calcium via green fluorescence). Results showed larger amount of green fluorescence in all blue-light conditions compared to the dark condition, with 1 and 2 J/cm2 displaying the highest amounts of fluorescence. Quantification of green fluorescence from tetracycline images corroborates these results, showing 2 J/cm2 has a significantly higher (p < 0.01) amount of fluorescence indicative of greater calcium deposition compared to the dark control. Together these results suggest that 4 J/cm2 is the upper threshold for seeing osteogenic benefits with blue-light exposure and 1 and 2 J/cm2 displayed the most significant increases in osteogenic differentiation and calcium deposition when compared to the dark condition. In conclusion, results from this work indicate that blue light exposure time is a critical factor that must be considered when developing an optimal dosing regimen. Further, blue light-mediated PBM can induce osteogenic cell differentiation and cell-mediated calcium deposition in the absence of external factors such as dexamethasone. Development of an optimal blue light dosing regimen to promote osteogenesis with consideration to both energy density and exposure time can be a promising non-invasive therapeutic approach for treatment of non-healing bone fractures.
Recommended Citation
Jarvis, Jamielyn Alcantara, "Effects of Different Blue Light Conditions on Saos-2 Cell Viability, Differentiation, and Mineralization" (2025). Theses and Dissertations. 1525.
https://repository.fit.edu/etd/1525