Anna Stachurska, Hanna M. Kowalczy˝ska
- Zak│ad Biofizyki, Centrum Medycznego Kszta│cenia Podyplomowego w Warszawie
Introduction. Cancer progression is a complex and multistage process, in which the adhesive properties of cells are changed, namely, the cell-cell interactions and cell adhesion to components of extracellular matrix (ECM). Furthermore, in the define stage their migration ability increases. Apart from the adhesive qualities, the active movement of cancer cells play a very important role. Cancer cells are derived from the primary tumor and start migration going through the basement membrane. They penetrate the blood or lymphatic vessels, and migrate via the blood stream to colonize different places in the organism where the interaction between cancer cells, endothelium and the ECM proteins take place. In in vitro research on prostate cancer, experiments are conducted into the PC-3 – human prostate cancer cell line derived from bone metastases. The
- high degree of invasiveness characteristic of the human prostate cancer cell line causes it to be an interesting model of study. The results of the research associated with the interactions of such cells with the ECM proteins and the expression of receptors are ambiguous and are often even at variance with each other. The present work investigates, for the first time, the interaction, under in vitro conditions, of the extremely invasive human prostate cancer cell line PC-3 with fibronectin adsorbed on sulfonated surfaces of polystyrene.
Objectives. The examination of the mechanism of the extremely invasive prostate cancer cell line PC-3 interaction with fibronectin - the ECM protein.
Materials and methods. Human prostate cancer cell line: PC-3 derived from bone metastases. 1. The estimation of the integrin receptor expression with the use of a flow cytometry technique. 2. The measurements of the static adhesion as well as the dynamics of spreading of PC-3 cells with the use of a programme intended for morphometrical analysis of cells. 3. Staining of cytoskeleton proteins (F-actin, α-actinin, vinculin as well as tubulin) in the cells adhered to fibronectin and visualization of the FAK kinase; the stained cell structures were determined with the use of an image analysis method.
Results. The answer of PC-3 cells to the signal from fibronectin adsorbed on sulfonated polystyrene surfaces was defined by the measurements of cell adhesion, estimation of dynamics of cell spreading and the organization of their cytoskeleton as well as the ability of cell proliferation and migration. The results of the present study are as follows: 1. The human prostate cancer cell line PC-3 interacts with the native form of fibronectin adsorbed on sulfonated polystyrene surfaces. 2. The adhesion kinetics is very high and the dynamics of cell spreading intensive. 3. On surface of the majority of cells (80-100% of the whole population) the integrin expression α5β1 and αvβ3 was confirmed. 4. The cell integrin interaction with fibronectin proceeds mainly via the RGD sequence, but the participation of the synergistic peptide (PHSRN) is also essential. 5. As a result of the cell interaction with fibronectin induction of a signal cascade takes place, which leads to clear reorganization of cytoskeleton proteins (F-actin, α-actinin, vinculin, tubulin) as well as to the activation of FAK kinase. 6. More intensive dynamics of spreading after αv▀3 binding with Ig anti αv▀3 may testify for the existence of the stimulation of other receptors. 7. The human prostate cancer cell line PC-3 interacting with fibronectin has the ability of active movement and proliferation.
Conclusions. 1. The experimental arrangement employed can be considered to reflect the interaction of PC-3 cells with a native form of fibronectin. 2. The answer of cells to the signal from fibronectin is manifested by the adhesion and the reorganization of cytoskeleton proteins, which results in the change of the cell shape in the process of spreading, cell migration and cell proliferation. 3. Inhibition of both the cell adhesion as well as the dynamics of cell spreading by the use of Ig anti integrin α5β1 proves the significant role of this receptor and the interaction of cells with the fibronectin. 4. The high expression of integrins and their participation in the interaction of PC-3 cells with one of the essential ECM protein, i.e fibronectin, suggests an important role of this interaction in the prostate cancer metastasis.