Down-regulation Of CD44via CD44shRNA Modulates The Biological Features Of Hunman Breast Cancer MDA-MB-231Cell Line

[Purpose]1. Investigate the expression of CD44splicing variants in breast cancer, to study the role of CD44mocular in progress of breast cancer.2. To silence the expression of CD44in breast cancer cell line MDA-MB-231by using short hairpin small interfering RNA (shRNA) and observe its effects effects on cell growth, adherence, invasion and migration of MDA-MB-231.[Methods]1. RT-PCR were used to examin the expression levels of CD44splicing variants in the breast cancer cell lines MDA-MB-231, MDA-MB-435and20cases of breast cancer tissue samples. The association between CD44splicing variants expression with clinicopathologic parameters were assessed.2. The CD44splicing variant which with the highest expression level in the two cell lines and breast cancer tissue samples was used as template, we construct specific shRNA expression vector. Recombinant plasmids were transient transferred into human breast cancer MDA-MB-231cells by lipofectamine2000respectively.3. To detect the CD44s expression in MDA-MB-231cells by real-time fluorescence quantitative PCR, and measured cell proliferation, adherence, invasion, migration by MTT cell proliferation assay, adhesion assay, transwell invation assay and motility assay "wound assay"[Results]1. RT-PCR revealed that the CD44splicing variants1,2,3,4,5,6,8were detected in the breast cancer cell lines MDA-MB-231, MDA-MB-435and breast cancer tissue samples in the known eight CD44splicing variants, the expression of CD44splicing variants4,5were higher in all detected CD44splicing variants.CD44splicing variants and clinical pathology parameter analysis showed that the kind of CD44splicing variants have nothing to do with the patients age, TNM staging, lymph node metastasis, ER and PR expression. A high expression of splicing variants1and 2mRNA were correlated to smaller tumor diameter, high expression of splicing variants4mRNA were correlated to higher histology grade, negative HER2status were correlated to expression of splicing variants2,3,6, but strong HER2status were correlated to expression of splicing variants5mRNA.2. shRNA-CD44s significantly suppressed the expression of CD44s gene in MDA-MB-231cells, inhibited the proliferation, declined the attachment and invasiveness of tumor cells to the extracellular matrix. The difference was significant compared with coutrol group (P <0.05).[Conclusion]1. The breast cancer cell lines MDA-MB-231, MDA-MB-435and breast cancer tissue samples showed a heterogeneous expression pattern of CD44splicing variants. CD44splicing variants4(the standard CD44, CD44s) mRNA expressed highest in MDA-MB-231, MDA-MB-435and breast cancer tissue samples.2. CD44s expression inhibited significantly suppressed the expression of CD44s gene in MDA-MB-231cells, inhibited the proliferation, declined the attachment and invasiveness of tumor cells to the extracellular matrix, CD44s gene had important effect on proliferation, adherence and invasion of MDA-MB-231breast cancer cells, strategies designed to CD44s gene silencing may respresent promising new strategy for breast cancer therapy.