Efeito da condição nutricional e do protocolo de fotobiomodulação na viabilidade e proliferação in vitro de células pré-osteoblásticas murinas

Abstract

Low-level laser irradiation (LLLI) has been used to induce the in vitro proliferation of various cell types. However, certain aspects of experimental protocols, particularly those related to cell culture and irradiation protocols, still require standardization. This study aimed to assess the impact of nutritional conditions and irradiation parameters on the viability and proliferation of murine preosteoblasts. MC3T3-E1 cells were maintained under standard culture conditions (αMEM supplemented with 10% fetal bovine serum) or nutritional deficiency conditions (αMEM without serum) and irradiated or not (control) with an InGaAlP diode laser at wavelengths of 660 nm (red) or 790 nm (infrared), with doses of 1, 4, or 6 J/cm², in a single dose in continuous mode. Cell viability and proliferation were assessed 24, 48, and 72 hours after irradiation using the Alamar blue reduction assay. The cell cycle and events related to cell death were evaluated by the propidium iodide stain and Annexin V/PI assay, respectively, through flow cytometry. The data revealed that in cells cultured with normal nutrition (10% FBS), there was no significant difference (p>0.05) in cell viability or proliferation among the different irradiation protocols. In contrast, in the experiments conducted under nutritional deficiency, the infrared laser at a dose of 6 J/cm² significantly increased (p<0.05) cell viability and proliferation compared with those of the control group at 72 hours. The data were confirmed by cell cycle and cell death events (Annexin V/PI) assays. Together, these results suggest that in vitro photobiomodulation yields more consistent biostimulatory effects on bone cells subjected to nutritional deficiency, highlighting the need for attention to simulate these conditions in studies with laser therapy in in vitro bone disease models and in in vitro experiments using photobiomodulation for bone tissue engineering.