Down-regulated MiRNA146a Of M2 Macrophages In Glioma Microenvironment Inhibit The Secretion Of IL-6

Object:Tumor microenvironment is a hotspot of current research,tumor immune inflammatory microenvironment in the development of tumor play an important role.In glioma, macrophages is the main cell in glioma immune inflammatory microenvironment play an innate immune role, in addition to the traditional tumor inhibitory M1 macrophages, alsoglioma promote type M2 macrophages.Study found miRNA146 a significantly higher expression in M2 type macrophage, M2 type macrophages can produce IL-6, IL-6 not only can promote the growth of glioma cells directly, but also can stimulate the M1 macrophages into M2 macrophages and indirectly promote tumor growth.In present in M2 macrophages miRNA146 a relations with IL-6 rarely reported.This study intends to establish type M2 macrophages model, through the lentivirus transfection down-regulate miRNA146 a expression in M2 macrophages, determination of type M2 macrophages lentivirus interference before and after the change of the secretion of IL-6, to clarify miRNA146a’s role in M2 macrophages to secrete IL-6.This study intends to establish M2 type macrophages model, through the lentivirus transfection down-regulate miRNA146 a expression in M2 type macrophages, measuring the change of IL-6 secreted by M2 macrophages before and after lentivirus interference, to clarify the role of miRNA146 a function in M2 macrophages to secrete IL-6.Methods:After normal cultivating mice macrophage RAW264.7 cells, using IL- 4 stimulus RAW264.7 into M2 macrophages, measured using immunohistochemical method type M2 macrophages CD206 express specific factors. M2 macrophages can be divided into 3 groups, transfection inhibitory carrier of PC group, transfection empty carrier of NC group completely medium only join with the blank control group.Using inhibitory carrier and empty vector transfection lentivirus M2 type macrophages.Take line type M2 macrophages after transfection to Real-time PCR to detection the change of miRNA146 a expressionand the change of the expression of IL- 6, GraphPad Prism 5 software to statistical processing of experimental data, all data to means + SD said, t test and other related statistical analysis, P < 0.05 statistically significant.Results:StimulateRAW264.7 with IL- 4, using immunohistochemical method, the determination of the M2 macrophages CD206 specific markers, DAB in brown after dyeing, suggest that IL-4 stimulation RAW264.7 type into M2 macrophages successfully.Using lentivirus after transfection, puro processing, transfection control PC group of carrier and transfection empty vector control group both express red fluorescence under the fluorescent microscope.On the PC and NC group and blank control group Real-Time PCR detection, Real-Time PCR results show that inhibitory transfection carrier type M2 macrophages after virus miRNA146 a compared with NC group and the blank control group significantly decreased(P < 0.05), the NC group compared with blank control group no significant change(P > 0.05).REAL-TIME PCR secretion of IL- 6 cases, the results showed that inhibitory transfection carrier virus type M2 macrophages after IL-6 in relative to the NC group and the blank control group significantly decreased(P < 0.05).Results suggest miRNA146 a cut in type M2 macrophages may reduce the expression of IL – 6 mRNA.Conclusions:Down-regulated miRNA146 a expression in type M2 macrophages, which can effectively reduce the M2 macrophages secrete IL- 6, in theory, so as to achieve the purpose of reduce IL- 6 stimulate the growth of tumor cells, and can reduce the M1 macrophages to M2 macrophage polarization.