The Function Of B7-H3 Molecule In Breast Cancer Cell Line MCF-7 And Its Effect On T Lymphocyte

B7-H3(also known as CD276)is a kind of type I membrane protein which is similar to PD-L1(also known as B7-H1).The molecule has contradictory co-stimulation and co-inhibition in adaptive immunity and anti-tumor immunity.At the same time,the influence of the molecule itself on tumorigenesis and development is also questionable.The expression of B7-H3 molecule is more clear,a variety of malignant tumors have a clear expression of up-regulated,including breast cancer.So we suspect that B7-H3 as a tumor immunosuppressive molecule,inhibit ed anti-tumor immunity.At the same time,the molecule can also affect the malignant biological behavior of tumor cells.Methods:The expression of B7-H3 in breast cancer cell line MCF-7 and hepatoma cell line HepG2 was detected by immunofluorescence labeling flow cytometry,and the B7-H3 molecule in tumor cells was silenced by siRNA transfection.CCK8 proliferation test,colony formation test and wound healing test were used to compare the effects of B7-H3 gene silencing on tumor biological behavior.Tumor cells were co-cultured with peripheral blood PBMC and CD3+T cells from healthy donors,to campare the effects of B7-H3 on T cell proliferation and expression of immune-related inflammatory molecules.Extracting RNA and protein to explore the mechanism of B7-H3 affecting lymphocyte function.Results:the expression of B7-H3 on MCF-7 and HepG2 was identified by flow cytometry.about 99.93%of MCF-7 cells expressed B7-H3 and 58.27%of HepG2 expressed B7-H3.MCF-7 was selected as the study cell.After silencing B7-H3 with siRNA,the expression rate of B7-H3 on MCF-7 decreased from 99.93%to 5.47%.The MCF-7 cells silenced by siRNA were named MCF-7-B7H3-/-,control cells as MCF-7-Gapdh.After silencing B7-H3,the proliferation and migration ability of MCF-7-B7H3-/-was lower than that of MCF-7-Gapdh.After co-culture of MCF-7-B7H3-/-and MCF-7-Gapdh with peripheral blood PBMC and CD3+T cells from healthy donors,we found that in the co-culture system of tumor cells and PBMC,if tumor cells highly expressed B7-H3,It can inhibit the activation,proliferation and function of CD8+T cells.However,the proliferation of CD4+T cells was not affected.when the expression of B7-H3 was inhibited,the proliferation ability of CD4+T cells and CD8+T cells was significantly improved.Subsequent experiments showed that B7H3 increased the expression of CTLA-4 on T cells and inhibited the release of IFN y from T cells.After inhibiting the expression of B7-H3,CTLA-4 decreased and IFN y release increased.at the same time,we found that the inhibition of IFN y release was achieved by inhibiting mTOR phosphorylation in T cells.After blocking mTOR signaling pathway,the inhibitory effect of B7H3 on IFN γ secretion was blocked.Conclusion:Our results suggest that B7-H3 can promote the proliferation and migration of MCF-7 and inhibit the proliferation of CD4+and CD8+T cells,induce the high expression of CTLA-4,in T cells and inhibit the release of IFN y by inhibiting the phosphorylation of mTOR in T cells.