Low-dose 5-aminolevulinic acid photodynamic therapy of promotes polarity switch of M1 macrophages

Objective To determine the effect of low-dose 5-aminolevulinic acid photodynamic therapy (ALA-PDT) on the polarity switch of M1 macrophages and explore its mechanism primarily. Methods RAW264.7 cells were cultured with 100 ng/mL lipopolysaccharide (LPS) for 24 h to induce M1 macrophages.The control group was cultured in the medium without LPS for 24 h.CCK-8 assay was used to detect the toxic effect of low-dose ALA-PDT therapy with different light doses on M1 macrophages, and the expression of M1 type marker, inducible nitric oxide synthase (iNOS) was measured with Western blotting.Then M1 macrophages were divided into M1 group, ALA group, red light group and ALA-PDT group (ALA treatment followed by red light irradiation).The mRNA expression levels of inducible M1 type markers iNOS and interleukin 6(IL-6), and M2 type markers interleukin 10(IL-10) and arginase 1(Arg-1) were detected by real-time fluorescent quantitative PCR, and the secretions of IL-6 and IL-10 were detected by ELISA.Western blotting was applied to detect the protein levels of iNOS, interleukin 1β(IL-1β), Arg-1, and autophagy-related proteins, ubiquitin-binding protein P62 and tubule-associated protein 1 light chain 3(LC3). Results Compared with the control group, the mRNA levels of iNOS and IL-6 were higher (P < 0.05), the protein levels of iNOS and IL-1β were higher (P < 0.05), and that of Arg-1 was lower (P < 0.05), and the secretion level of IL-6 was higher in the M1 group (P < 0.05).With the increase of light intensity, the toxic effect of ALA-PDT therapy on M1 macrophages was gradually increased (P < 0.05), and when the intensity higher than 2 J/cm2, the expression of iNOS was inhibited.The mRNA levels of Arg-1 and IL-10 were increased, while that of iNOS was decreased (P < 0.05), the protein expression of Arg-1 was elevated while those of iNOS and IL-1β were decreased (P < 0.05), the secretion level of IL-10 was increased (P < 0.05) but that of IL-6 decreased (P < 0.05), and the protein level of P62 was decreased (P < 0.05) and the ratio of LC3Ⅱ/LC3Ⅰ was increased (P < 0.05) in the ALA-PDT group when compared with the M1 group, ALA group and red light group. Conclusion Low-dose ALA-PDT therapy can promote the conversion of M1-type macrophages to M2-type macrophages, and its mechanism may be related to autophagy.