Preliminary Study On The Mechanism Of Bone Sialoprotein Regulating Breast Cancer Metastasis By Integrin αvβ3

Breast cancer is a common female malignant tumour with a high morbidity, which increases on a yearly basis. Featured with high occurrence rate, osseous metastasis of breast cancer is very difficult to treat. In autopsy, over 90% death patients are found with osseous metastasis. although good curative effect is achieved in comprehensive treatment with surgery as the main method, certain limitations still exist because metastasis remains the primary factor that triggers postoperative recurrence. consequently, it’s significant to find out special targeted treatment site through deep study on molecular mechanism of osseous metastasis of breast cancer.Integrin αvβ3 is a kind of specific membrane receptor proteins can recognition RGD sequence, specificity combination with the protein containing RGD(Arg-Gly-Asp) motif, play an important role in tumor cell adhesion, migration, infiltration, and tumor angiogenesis. Bone Sialoprotein(BSP) is a highly phosphorylation and glycosylatiion secreted protein in extracellular matrix. BSP was overexpressed in the tumor, lung cancer, and prostate cancer, since a number of studies indicate that BSP play an important role in the process of tumor.The present study that BSP binding with cell membrane receptor integrins αvβ3 can cause a series of cell biology effect, but the mechanism of BSP mediated regulation of integrin αvβ3 is still not clear. Previous experimental work in our lab, found that BSP through integrin αvβ3 regulated ILK signaling pathways in of breast cancer MDA-MB-231BO(231BO)cells, and affect cell proliferation; Recent laboratory experimental results also showed that BSP and integrins αvβ3 may have a positive correlation in 231 BO cells, that is, the expression of integrin αvβ3 decline in stable BSP gene silence of breast cancer cells.Based on previous experimental studies, we analyzed the correlation between BSP and αvβ3 integrin in clinical breast invasive ductal carcinoma tissues by immunohistochemical method. Build β3 lentivirus expression vector, transfected BSP gene stable silence of breast cancer cells, and the expression level of β3 was detected by immunocytochemistry, flow cytometry and Western Blot. The proliferation, migration and invasion ability of cells were detected by CCK8 assay, scratch test and Transwell assay in vitro. To explore the biological characteristics of BSP-/β3+(231BO-BSP27/β3)cells, and revealed BSP through regulating expression of integrin β3 influence the molecular mechanisms of bone metastasis of breast cancer.The results in brief are as follows:1.Immunohistochemical detection of 61 cases of breast invasive ductal carcinoma, BSP and integrin αv、β3 were positively correlated2. Obtain successfully infected BSP-/β3+( 231BO-BSP27/β3) cells by neomycin selection after using the lentivirus to infect 231BO-BSP27 cells.3. Flow cytometry results showed that the quantity of β3 protein in 231BO-BSP27/β3 cells(89.705%±0.456) is higher than 231BO-BSP27(45.941%±0.396)cells, P<0.05; Western Blot results showed that the quantity of β3 protein in 231BO-BSP27/β3 cells(0.428±0.013) is higher than 231BO-BSP27 cells( 0.237±0.012), P<0.05; Immunocytochemistry in 231BO-BSP27/β3 cells was strongly positive, and β3 expresion was mainly lcated in the cell membrance and cytoplasm.4. Experiment of extracorporal cytobiology shows cell growth rate, capabilities of migration and invasion of 231BO-BSP27/β3 are higher than 231BO-BSP27.