The Role Of Nonmuscle Myosin Heavy Chain ⅡA (NMHC ⅡA) Isoforms In Breast Cancer Metastasis

Malignant tumor is serious harm to human health and life. Nearly20years,malignant tumor mortality rates have risen29.42%, and tend to be younger.Breast cancer (mammary carcinoma) is one common kind of female malignanttumors. The main reason of death of breast cancer is recur and metastasis.Invasive and metastasis tumor has become a big obstacle to the treatment oftumors. But tumor invasion and metastasis is primarily the result of cellmotility changes.Cell movement is regulated by actin and myosin[1]. But theprocess is regulated by cytoskeleton and myosin initiator promoter. So myosingradually becomes a research hotspot.The nonmuscle myosinⅡ(NMⅡ) is composition of the cytoskeleton. NMⅡinvolved in cell polarity,cell migration and cell adhesion in this three processwith actin interaction. But the process is the initial stage of tumormetastasis.So we conclude that the NMⅡ plays an important role in theprocess of tumor metastasis. NMⅡ is six polymer structure composed of twoheavy chain (NMHC,230kDa) and two light chain (NMLC17MLC20)[4]inmammals. NMLC has regulation and stable effect, and NMHC determine cellmotility. So NMHC maybe a key role in the process of tumor metastasis.NMHCⅡ has three subtypes, respectively NMHCⅡA, NMHCⅡB andNMHCⅡC, and is coded by three different genes (Myh9, Myh10and Myh14)[9]. NMHCⅡA as a a productivity protern involed in cell migration. In thisresearch we observes breast cancer cells growth, adhesion and invasioncapacity by silence NMHCⅡA gene in MCF-7cells. Objective: We utilize RNAi to silence NMHCⅡA gene in MCF-7cells toobserve the growth and adhesion of breast cancer cell lines MCF-7cell.Methods: We utilize RNAi to inhibit NMHCⅡA gene expression ofMCF-7cell. In the experiments include immunohistochemical staining andWestern Blot to evaluate NMHCⅡA protein expression;CCK-8kit to detectMCF-7cells proliferation; cell scratches experiment, Boyden chamber,immunohistochemical staining of MMP9and tublin to detect cell invasionability. We utilize empty plasmid group as a positive control and untransfectedgroupas a negative control.Results:1. Both Immunofluorescence staining and western blot of NMHCⅡAconfirm transient transfection success. Immunofluorescence abserves siRNAtransfection efficiency up to80%.2. CCK-8kit tests showed that, Each group of MCF-7cell growth situationis consistent.3. Cell scratch test results show that experimental group of cells in vitromovement and migration capable is stronger than empty plasmid group and thenon-transfected group.4. Boyden chamber results show that experimental group of cells which thedepth of invasion and the number of cells to through basement membrane ismuch smaller than empty plasmid group and the non-transfected group.6. Immunohistochemical staining experimental results show thatexperimental group of cells MMP-9expression is weaker than empty plasmidgroup and the non-transfected group.(P <0.001)7. Tublin immunofluorescence staining show that experimental groupcytoskeleton arranges disorderly, some skeleton is missing, but empty plasmidgroup and the non-transfected group are normal.Conclusion: 1. We utilize RNAi to inhibit NMHCⅡA gene expression of MCF-7cellshow that cell proliferation ability is no change.2. We utilize RNAi to inhibit NMHCⅡA gene expression of MCF-7celllead to cell migration and invasion deceeased.3. NMHCⅡA regulates tumor migration and invasion through changing cellmotility and the synthesis ability of MMP9, It relates with tumor metastasis.