GO nanoparticles increase the efficiency of phagocytosis by macrophages in normal and tumor environment
Abstract
Previous experiments on mice suggested that the antitumor effect of M3 macrophages is based on their increased ability to phagocytize pathogenic cell antigens. However, the use of M3 macrophages for treatment of tumors in humans is complicated by: 1) obtaining a sufficient number of macrophage precursors; 2) differentiation of progenitors into macrophages and their reprogramming into the M3 phenotype in vitro; 3) delivery of macrophages to the tumor. Understanding these problems and the fact that there are numerous macrophages of the M0 and M2 phenotypes in the tumor zone make it promising to develop a new method of therapy for tumors by enhancing the ability of M0 and M2 macrophages to phagocytosis. A literature search for factors that increase the macrophage phagocytic activity has revealed a potential candidate for this role, graphene oxide (GO) nanoparticles.
The aim of this work was to test the hypothesis about a possibility of enhancing the macrophage phagocytic function with GO nanoparticles.
Methods. Murine macrophages isolated from peritoneal lavage were used in this work. To produce the M0 phenotype, macrophages were cultured in the DMEM/F12 medium. To produce the M2 phenotype, macrophages were cultured in the DMEM/F12 medium supplemented with 40% FBS. The M3 macrophage phenotype was obtained by adding IFN-γ and STAT3, STAT6, and SMAD3 inhibitors to the culture medium. Then all macrophage phenotypes were stimulated with lipopolysaccharide (LPS). After the end of LPS stimulation of macrophages, the phagocytic ability of different phenotypes was evaluated using a pHrodo® Red Staphylococcus aureus Bioparticles stained bacteria kit (Invitrogen). GO was obtained by electrochemical stratification of working electrodes from highly oriented pyrolytic graphite. Macrophages were cultured in the presence of GO for 2 hours, then washed, and the phagocytic index was determined according to the standard method with pHrodoTM BioParticles® fluorescent particles. The phagocytic activity was assessed for different macrophage phenotypes in normal and tumor environments.
Results. The addition of GO increased the phagocytic ability of macrophages with the M0 and M2 phenotypes by 20% and 19%, respectively, compared with the control, whereas the phagocytosis coefficient of macrophages with the M3 phenotype practically did not change. In addition, GO nanoparticles reduced the depressive effect of the tumor environment on the phagocytic activity of macrophages. This effect significantly depended on the macrophage phenotype. For M0 macrophages, there was an almost twofold increase in the activity for the M2 phenotype, the phagocytic activity in the tumor medium increased by more than 50% with the addition of GO; and for the M3 phenotype, the addition of GO had practically no effect on its already sufficiently high activity in the tumor medium.
Conclusion. GO nanoparticles were able to increase the baseline activity of M0 and M2 macrophages and to restore their phagocytic capability that had been weakened in the tumor environment.
