The Effects Of Different Polaration Phenotype Macrophage On Biological Behaviors Of Oral Squamous Cell Carcinoma

ObjectiveThis project focused on the investigation of the effects of different polarization phenotype macrophage on the proliferation,apoptosis,migration and invasion of SCC-15 cells in vitro.Immunohistochemical staining method detected CD68 positive cells(Tumor-associated macrophages,TAMs)of OSCC samples,which represented the relationship between CD68 expression and OSCC clinicopathological features.Meanwhile,the double immunofluorescence staining method detected the ratio of CD206 positive cells(M2 macrophages)to CD68 positive cells,which showed the relationship between CD206 expression ratio and lymph node metastasis.Finally,the effects of different polarization phenotype macrophage on biological characteristics and possible molecular mechanisms of OSCC were preliminarily explored.This project provided a possible direction for the prognosis of OSCC and biological targets for the treatment of OSCC.Research methods1.THP-1 cells were induced to differentiate into M1 and M2 macrophages in vitro,and the phenotype was detected by flow cytometry and Real-time-qPCR.2.CCK-8 proliferation assay detected the proliferation of SCC-15 cells in M1 and M2 conditioned medium after 24 h,48 h and 72 h.3.Transwell assay detected the number of migration and invation cells.4.After SCC-15 cells were co-cultured with M1 or M2 macrophages for 24 h,Real-time-qPCR assay detected the expression of proliferation-related genes Cyclin D1,Ki-67,and apoptosis-related gene Bcl-2.5.SCC-15 cells non-contacted co-cultured with M1 or M2 macrophages for 24 h,Western Blotting assay detected the expression of Caspase-9,E-Cadherin,N-Cadherin,Vimentin,MMP-2 and MMP-9.6.62 OSCC samples and 10 normal oral mucosa samples were used.Immunohistochemistry staining detected CD68 positive cells.7.Double immunofluorescence staining method detected the ratio of CD206 positive cells(M2 macrophages)to CD68 positive cells,to analyze the relationship between CD206 expression ratio and lymph node metastasis.Results1.The vitro experiments showed that M1 macrophages promoted the apoptosis of SCC-15 cells,and inhibited the proliferation,migration and invasion abilities of SCC-15 cells.M2 macrophages inhibited the apoptosis of SCC-15 cells,increased the proliferation,migration and invasion abilities of SCC-15 cells.2.Immunohistochemical staining experiment showed that a significantly(P < 0.05)increased count of CD68 positive cells in OSCC samples compared to normal oral mucosa samples,and the higher CD68 positive cell infiltration density in OSCC was correlated with lymph node metastasis(P < 0.05).3.Immunofluorescence staining experiment showed that comparing to N0 OSCC,the ratio of CD206 positive cells was significantly(P < 0.05)increased in N+ OSCC samples.ConclusionTAMs are closely related to the occurrence,development and prognosis of OSCC.Different polarization phenotypes of macrophages have different effects on the biological behavior of oral squamous cell carcinoma.M1 macrophages can inhibit the development of OSCC,while M2 macrophages play an important role in promoting the development of OSCC.