The Study for Functions of Annexin a2 in Human Breast Cancer
|School||Tianjin Medical University|
|Keywords||Breast Cancer Annexin a2 Proliferation Migrate Attack|
Objective: annexin a2 (Annexin a2) is a calcium-dependent phospholipid binding protein, a broad highly expressed in the tumor cells, is one of the significant differences in the protein expression level of tumor cells and normal tissue cells. Signal transducer and activator of transcription 3 (STAT3) has the dual function of signal transduction and transcriptional activation of highly expressed in many human malignancies, regulation and proliferation, invasion protein c-Myc, cyclin D1, MMP-2 and MMP -9 protein expression levels, and therefore its overexpression and tumor cell proliferation, migration and invasion closely related. This study RNA interference to reduce the expression level of Annexin a2 breast cancer cells, and to explore the effects of breast cancer cell proliferation, migration and invasion, and to study the interaction with STAT3 exists to further explore Annexin a2 regulate breast cancer cell mechanisms of proliferation, invasion and metastasis. Methods: Annexin a2 protein expression levels of RNA interference technology to reduce breast cancer MDA-MB-231 cells, and Western blotting technology to verify the expression levels. 2 down using the MTT assay Annexin a2 expression of MDA-MB-231 cell proliferation. 3 colony formation assay inhibition of gene silencing Annexin a2 colony-forming ability of MDA-MB-231 cells. 4 Flow cytometry was used to detect changes in cell cycle distribution of MDA-MB-231 Annexin a2 expression levels. The scratch assay Annexin a2 expression levels decline in the ability to influence the migration of MDA-MB-231 cells. 6. Vitro invasion assay Annexin a2 drop expression on the invasive ability of MDA-MB-231 cells. 7. Immune coprecipitation interacting proteins detected by Annexin a2, and immunofluorescence Annexin a2 protein interactions in intracellular localization. Molecules detected by Western blotting techniques Annexin a2 drop expression and proliferation-related protein c-Myc, cyclin D1, MMP-2 and MMP-9 expression levels to further clarify Annexin a2 regulation of breast cancer cell proliferation, migration and invasion mechanism. Results: 1. Using RNA interference technology transfection of human breast cancer cells MDA-MB-231 screened Annexin a2 protein expression levels of 80% -100% 3 monoclonal cells. MDA-MB-231 cell proliferation 2. Annexin a2 expression levels significantly decreased (P lt; 0.05). 3. Annexin a2 MDA-MB-231 cells after gene silencing clone forming ability was significantly lower (P lt; 0.05). 4. Annexin a2 expression levels led to the proliferation of MDA-MB-231 cells fell, the increase in the proportion of G1 phase of the cell cycle, G2/MS period was significantly decreased (P lt; 0.05). Scratch experiments found that the migration of breast cancer MDA-MB-231 cells was significantly decreased (P lt; 0.05) Annexin a2 drop expression. 6 in vitro invasion assay results showed that the cells in vitro invasion ability Annexin a2 drop expression significantly decreased (P lt; 0.05). 7 coimmunoprecipitation technology found Annexin a2 activation and signal transduction and transcription factor 3 (STAT3) there is interaction. 8. Western blotting to detect Annexin a2 gene silencing c-Myc, cyclin D1MMP-2 and MMP-9 expression levels were significantly decreased. Conclusion: 1. Immunoprecipitation detection of MDA-MB-231 cells discovered protein interaction with Annexin a2 mesh interaction with STAT3 existence; immunofluorescence detected in MDA-MB-231 Field cellular Annexina2 STAT3 mainly located in the cell pulp. Proliferation and colony forming ability of MDA-MB-231 cells after 2. Annexin a2 expression levels decreased, decreased cell proliferation index and increased the proportion of G1 phase of the cell cycle distribution G2/MS period was significantly decreased. Annexin a2 expression levels decline in the level of expression of c-Myc and cyclinDl that Annexin a2 affect the level of expression of c-Myc and cyclinD1 with STAT3 interactions regulate the proliferation of breast cancer cells. 3. Annexin a2 gene silencing migration of MDA-MB-231 cells was significantly decreased in vitro invasive ability was significantly weakened. Annexin a2 expression levels of MMP-2 and MMP-9 expression levels, indicating that Annexin a2 by regulation to STAT3 interaction of MMP-2, MMP-9 expression levels and thus affect breast cancer cell invasion.