Secreted MiRNAs Expression Profiles Of Cultured Human Breast Cancer MCF-7 Cells At Difierent HER-2 Levels

Background:Human epidermal growth factor receptor-2 (HER-2), which belongs to the membrane tyrosine kinase, is a member of epidermal growth factor receptor family. When HER-2 gene is activated, it provides the cell with potent proliferative and antiapoptosis signals and it is the major driver of tumor development and progression in mammary cancer. MicroRNAs (miRNAs) are small (18-24 nucleotides), non-protein encoding, endogenous single-stranded RNAs, which regulate cellular processes mainly involved in tumor biology, such as cell proliferation, differentiation, apoptosis and metastasis. At present, miRNAs are known to serve as either oncogenes or oncosuppressors in the regulation of cells. In recent years, the reports about the relevance between HER-2 and miRNAs were increasing. Some studies suggested that the development of breast cancer were associated with the overexpression of HER-2 and the disorder of miRNAs. Our previous studies also showed that the miRNAs expression profiles of breast cancer cells did have difference at different HER-2 levels. Therefore it is very important to explore the relationship between miRNAs and HER-2 expression. With the clinical significance of liquid biopsy technology being recognized, the study about cancer patients’body fluids in the clinic and tumor cells’secreted substances of tumor cultured supernatant in the laboratory has become a hot research topic.Objective:To screen out tumor-secreted miRNAs expression profiles related to HER-2 gene by detecting the expression of miRNAs in the supernatants of cultured human breast cancer cells at different HER-2 expression levels.Methods:(1) Two human breast cancer cell lines, MCF-7with low level expression of HER-2 and MCF-7/HER-2 stably transfected by HER-2 into MCF-7, were cultured. The expression profiles of miRNAs in the supernatants of cultured MCF-7/HER-2 cells were detected by miRNAs microarray analysis, and compared with those in the corresponding non-HER-2-transfected MCF-7cells.(2) The possible pathways of miRNAs relating to HER-2 signaling were forecasted and analyzed by application of bioinformatics methods.Results:(1) miRNAs microarray analysis showed that 138 miRNAs were related to HER-2, in which 86were up-regulated and 52 were down-regulated, using the criteria of more than 2 folds increasing or decreasing of miRNAs compared with HER-2-negative MCF-7 cells. Among these miRNAs,8 of them were up-regulated more than 5 times such as hsa-miR-92b-5p, hsa-miR-582-3p, hsa-miR-302d-5p, hsa-miR-5703, hsa-miR-583, hsa-miR-4270, hsa-miR-3150b-3p, hsa-miR-4489 and 1 was down-regulated more than 5 times such as hsa-miR-589-3p.(2) Among those differentially expressed miRNAs in the supernatant,7 of them were up-regulated and 7 were down-regulated in concordance with those in the cells. Meanwhile,1 up-regulated miRNA in the supernatant was down-regulated in the cells, and 7 down-regulated miRNAs in the supernatant were up-regulated in cells.(3) Bioinformatics analysis showed that Hsa-miR-5703, hsa-miR-92b-5p and hsa-miR-589-3p might be directly or indirectly related to HER-2.Conclusion:(1) The expression profiles of tumor-secreted miRNAs in the supernatants of cultured human breast cancer MCF-7 cells at different HER-2 levels were successfully obtained.(2) MiRNAs excreted outside human breast cancer cells are not concordant with their levels in the cells.(3) Hsa-miR-5703, hsa-miR-92b-5p and hsa-miR-589-3p may be involved in the breast cancer via HER-2 signaling pathway.