Involement of actin regulatory protein expression in tyrosine kinase related cell growth of undifferentiated cultured human umbilical stem cells

• Main Authors: Rui-You Li, 李瑞祐
• Other Authors: 劉英明
• Format: Others
• Language: zh-TW
• Online Access: http://ndltd.ncl.edu.tw/handle/40901592828969302864

碩士 === 國立中興大學 === 生命科學系所 === 96 === It is generally accepted that dynamic reorganization of actin polymerization and depolymerization is the basic mechanism for maintaining cell polarization, cell proliferation and regulating cell division. In contrast to Arp2/3 which generates branched filaments, formins together with profilin nucleate straight actin filaments. Formin-2 is a formin homologous protein that is expressed in the brain, the spinal cord and in oocytes. Studies with formin gene knock out (KO) mice showed that the KO mice displayed the deficient meiosis of oocytes, suggesting the formin may control the actin filament reorganization and cell division. Physiological functions of formin-2 in the cells are not currently understood. it has not been determined if formin-2 is expressed in other tissues and how an actin regulator could control cell cycle progression in cells. In this study, we found that formin-2 mRNA and protein are expressed specifically in cultured human umbilical Wharton jelly cells based on the data obtained with a semi-quantitative RT-PCR and a western blotting analysis. These umbilical cells have been shown to have the self-renewal and differentiation capacity of mesenchymal stem cells. The expression level of forming-2 within the cultured cells is not varied by cell passages from P4 to P10. Genistein is known to be a specific inhibitor of tyrosine-specific protein kinase, ie., EGF receptor kinase, and Src. In this study, we found that this compound has growth arrested cultured human umbilical stem cells at the G0/G1 phase in cell cycle determined by the flow cytometry, as well as reduced the cell survival measured by the MTT assay and cell counting. To determine if forming-2 is involved in cell growth of the undifferentiated stem cells, we examined the effects of genistein on the cellular distributions of FMN-2, profilin, tropomyosin, caldesmon, F-actin within the cells by using a confocal fluorescence scanning microscope accompanied by applying a specific antibody against each protein. Results obtained indicated that genistein induced the translocation of actin nucleating proteins, forming-2 and profilin to the peri-nuclear areas. Caldesmon and tropomoysin become associated with the stress fiber. It seems likely that the actin filament remodeling in the peri-nuclear area and some binding proteins associated with the stress fiber might have the functions for inhibition of cell proliferation in undifferentiated stem cells. Future study might be invested to study further if changes in cell cytoskeleton remodeling are also associated with differentiation of mesenchymal stem cells.