The effect of medium and low dose X-ray irradiation on the polarity changes of tumor associated macrophages

Objective To explore the effect and mechanism of medium and low dose X-ray irradiation on the polarity changes of tumor associated macrophages.Methods The mouse RAW264.7 macrophage line was cultured with the supernatant of 4T1 tumor cell culture (4T1-TCS) of mouse breast cancer to induce it to become M2 tumor associated macrophages (TAMs). On this basis, medium and low dose X-ray irradiation was performed, and NO was detected by Griess method, CD86 and CD206 were detected by flow cytometry, cytokines IL-10 and IL-12p70 were detected by ELISA, iNOS and Arg-1 were detected by immunofluorescence, and polarization related pathway proteins were detected by WB method to observe whether medium and low dose irradiation could change the polarity of tumor associated macrophages.Result 4T1-TCS induced TAM secretion of NO decreased, flow cytometry expression of CD206 increased, immunofluorescence expression of Arg-1 increased, and expression of iNOS decreased; Low dose irradiation (0.075Gy, 0.5Gy, 1Gy) TCS-TAM showed no significant changes in all indicators; Moderate doses (2Gy, 4Gy, 6Gy) of irradiation on TCS-TAM resulted in increased secretion of NO by cells. Flow cytometry results showed an increase in CD86 expression and a decrease in CD206 expression; The immunofluorescence results showed that the expression of iNOS in cells increased, while the expression of Arg-1 decreased; The ELISA results showed that the secretion of IL-12p70 by cells increased, while the secretion of IL-10 decreased. The WB results showed that the expression of NOX2, ATM, and IRF5 proteins increased during moderate dose irradiation.Conclusion 4T1-TCS can successfully induce macrophage RAW264.7 cells to polarize into the M2 phenotype, while low-dose irradiation has no significant effect on the polarization phenotype of TCS-TAM. Moderate dose irradiation can induce TCS-TAM polarization into the M1 phenotype that inhibits tumor growth, and the reversal mechanism of polarization is related to the IRF5 pathway.