CONICET | Buscador de Institutos y Recursos Humanos

Photobiomodulation (PBM) is the use of light in a specific wavelength and irradiance to induce some process in the cells such as death or division. One possible way to follow the light effects is through the optical redox ratio (ORR). ORR is an established microscope optical technique that uses the endogenous fluorescence of NADH (reduced form of nicotinamide adenine dinucleotide) and FAD (flavin adenine dinucleotide) to verify the metabolic route of a cell. The purpose of this study was to measure the effect of light on the ORR of Candida albicans cells, growing in aerobic and anaerobic conditions. Standardized C. albicans suspensions were adjusted in a spectrophotometer to 10^7 colony forming units per milliliter (CFU/mL) (540 nm) after growing 12 or 24 hours under aerobic and anaerobic conditions. The anaerobic state was achieved using an airtight jar with a candle inside. Then, the PBM was performed using a LED device emitting at 450 nm with an irradiance of 44 mW/cm2 and a total fluence rate of 200 J/cm2. The ORR was evaluated through confocal laser microscopy using laser excitation of 755 nm (NADH excitation) or 860 nm (FAD excitation), and images were acquired in the channel mode of the microscope with 440 - 480 nm (NADH fluorescence) or 500 - 550 nm (FAD fluorescence) wavelength range, respectively. The ORR images were created by computing pixel-wise ratios of FAD/(NADH+FAD) fluorescence. Cell viability was quantified by CFU/mL assay. According to the viability test, the anaerobic suspensions (12 and 24 h) showed lower growth in comparison with the aerobic suspensions (12 and 24 h). The ORR is statistically the same in aerobic and anaerobic conditions, and lower after the illumination. In conclusion, our preliminary study demonstrated that it is possible to measure the ORR in C. albicans cells and the effects of PBM in these cells.