Research On The Inhibition Of Proliferation And Metastasis Of Breast Cancer Resulting From MiR-22

Background and purpose:Neurokinin-1 receptor(NK1R),microRNAs(miRNAs)or estrogen receptor alpha(ERα)respective deregulation is frequent in human breast cancer(BCs),but the relationship between the abnormal expressions of NK1 R and ERα,and whether this association is regulated by the specific mi RNA,is rarely reported.This topic aims to clarify,whether miR-22 can targeted downregulate the expression of ERα and NK1R-Tr in the breast cancer tissues and cells,to inhibit the proliferation,invasion and metastasis of breast cancer.Methods:1.At tissue and cell levels: Rt-PCR studies the expression of mi R-22 and NK1R-Tr in breast cancer tissues and the paired adjacent normal tissues;Rt-PCR and Western Blot method detect the expression of miR-22,NK1 R Tr and of ERα in breast cancer cells and the normal breast epithelial cells.2.Luciferase reporter assay and chromatin immune coprecipitation experimental studied regulation network between miR-22,NK1R-Tr and ERα3.Constructed MCF-7-ERαI and MDA-MB-231-ERα cell lines with small interference RNA of ERα and ERα expression plasmid;Knocked out or overexpress NK1R-Tr of MCF-7 and MDA-MB-231 cells with small interfering RNAor NK1R-Tr expression plasmids.4.Biological behavior research of breast cancer cells.5.Western Blot assay analysed the signaling pathways research of SP-NK1R-ERK1/26.The effects of miRNA-22 on cell proliferation and metastasis of breast cancer cells in vivo.In the subcutaneous tumor model,mi RNA-22 overexpressed MCF-7 cells were inoculated from the groin.In the metastases tumor model,miRNA-22 overexpressed MCF-7 cells were inoculated from caudal vein.Results:1.The expression of mi RNA-22 in cancer tissues was significantly less than that in adjacent normal tissues,while the expression of NK1R-Tr is obviously increased.The expression of miRNA-22 significantly lower in breast cancer cell lines than that in mammary epithelial cell HBL-100,while the expression of NK1R-Tr is higher.NK1R-Tr and ERα were both positive in MCF-7 and T47 D cells.2.Luciferase reporter experiment showed when miRNA-22 mimics and ERα/NK1R-Tr expression plasmids were transfected,the luciferase activity was obviously reduced in HEK-293 cells.The expression of ERα and NK1R-Tr were reduced when miRNA-22 mimics were transfected.CHIP experimental proved that NK1R-Tr chromatin fragment could be accumulated by the antibodies of ERα.And the luciferase activity increased obviously when ERα and NK1R-Tr expression plasmid were transfected in HEK-293 cells.3.The decreased degree of NK1R-Tr expression in MCF-7-ERαI was far less than that in the wild MCF-7 cells when overexpressing miR-22 The action that miR-22 downregulated NK1R-Tr was more apparent in MDA-MB-231-ERα cells compared with the wild MDA-MB-231 cells.4.The proliferation ability,the clone formation ability,migration and invasion of the miRNA-22 overexpressed MCF-7 cells were significantly reduced compared with wild type MCF-7 cell.Results were the same in MDA-MB-231 cells.5.The peak of phosphorylation of ERK1/2 was delayed and weakened in the miRNA-22 overexpressed MCF-7 cells compared with the wild type MCF-7 cells,which was in accordance with the effect of NK1R-Tr antagonist.6.miRNA-22 inhibited the growth of MDA-MB-231 and MCF-7 engrafted tumors and repressed the distal pulmonary metastases in vivo.Conclusion:miRNA-22 could directly downregulate the expression of NK1R-Tr,and then delayed and weakened the phosphorylation of ERK1/2 to inhibit the proliferation and metastasis of breast cancer.