Avaliação do efeito da fotobiomodulação com LED ambar sobre leucócitos humanos polarizados para fenótipos pró ou anti-inflamatório

Abstract

The phenotypic plasticity of leukocytes in the different phases of the inflammation and repair process, is the reason these cells are considered key players in their control. The effects of Photobiomodulation (PBM) on the treatment of tissue lesions have been widely described, but little is known about their role on leukocyte polarization. This project aimed to evaluate the optical absorbance spectrum of monocytes, neutrophils and human lymphocytes as well as the effects of PBM with amber LED with 590 nm on the gene and protein expression of mediators produced by these cells, which have been polarized to either the pro- or anti-inflammatory / regulatory phenotype. Peripheral human blood leukocytes were cultured in RPMI with 5% or 10% fetal bovine serum (to activate the pro- or anti-inflammatory phenotypes respectively), polarized with LPS or IL-4+LPS for 2 hours and irradiated with amber LED 590 nm (4J; 70 mW; 23.3 mW / cm2; 2.66 J / cm2), using one application after 2h or two applications after 2h and 6h. The cultures were collected to evaluate the optical absorbance spectrum (neutrophils after 12 hours and monocytes and lymphocytes after 24 hours) and for the analysis of mRNA expression of the cytokines IL-1β, IL-6, IL-10, IL-17, TNF-α, IFNγ and VEGF using qPCR (after 4 hours for neutrophils and 24 hours for monocytes and lymphocytes). The culture supernatants were used for protein evaluation via enzyme immunoassay (neutrophils: IL-1β, TNFα and IL-10; lymphocytes: IFNγ, IL-17 and IL-10 and monocytes: IL-1β, IL-6, TNFα and IL -10). At least 3 experiments (samples from 3 individuals) were carried out in duplicate. Non-irradiated and non-polarized cells served as controls. Monocytes and neutrophils showed peak absorption between 520-570 nm and lymphocytes peak absorption between 450-570 nm and 770-900 nm. Irradiation with amber LED, applied to cells polarized to the pro-inflammatory profile, generated a decrease in the protein synthesis of the pro-inflammatory cytokines IL-6 and TNFα in the monocytes, a decrease in the production synthesis of TNFα in neutrophils and, in the lymphocytes, an increase in the production of IL- 17. In leukocytes polarized to the anti-inflammatory / regulatory profile, irradiation with amber LED reduced the production of IL-10 in monocytes and lymphocytes. These results show that leukocytes present peak absorption in regions of the spectrum which remain mostly unexplored in PBM. In addition, they show that amber LED is capable of modulating

the expression of cytokines which are important to the modulation of inflammation and repair processes; this may contribute to the optimization of inflammatory processes, as well as immunotherapy treatments.