Analysis Of Cell Surface Receptors On CD₈⁺T Cell And Effects Of The Bone Morphogenetic Protein On Human Breast Cancer Cell

This dissertation includes two parts:(1) the changes of the morphology of CD8 +T cells were detected by atomic force microscopy (AFM) after stimulation, and Visualization of the cell surface receptors recognition was also performed by AFM (2) we determined BMP-2 effects on the migration of the human breast cancer cells.In the first part, CD8+T cells were firstly isolated from human peripheral blood in vitro and activated by phytohaemagglutinin (PHA). Cell morphology was observed by atomic force microscopy (AFM) and CD8 antigen molecules distribution on CD8+T cells was studied by a functionalized AFM tip. Laser confocal scanning microscope (LCSM) experiment showed that the CD8 antigen molecules were distributed on the CD8+T cells surface, and the AFM experiment showed that comparing to resting CD8+T cells, the diameter and height of activated CD8+T cells became larger, the ultrastructure of the cells became more complex. The strength of the specific binding force of the CD8 antigen-antibody interaction was found to be approximately four times bigger than that of the unspecific force. The CD8 antigen molecules were not randomly distributed on the surface of a single activated CD8+T cell, which were formed into nanometer domain. The force of CD8 antigen-antibody interaction of CD8+T cell did not change significantly when CD8+T cells were activated, these results suggest that CD8 antigen-antibody interaction is highly selected and high affinity. Atomic force microscopy can provide a new tool to study the specificity T cell antigen recognition and activation, it makes us to better clarify the T cell antigen recognition and activation mechanism.In the second part, to determine the effects of BMP-2 on the migration of the human breast cancer cell MCF-7. the cell was first induced by BMP-2(30ng/mL) for 24h, atomic force microscopy(AFM) was used to observe the changes in cell morphology and the CLSM was used to detect the distribution of CD44 and E-cadherin molecules on MCF-7. Cell migration and invasion abilities were assayed by scratch healing and transwell experiments. The formation of lamellipodia together with increased cell length was observed in the cells of BMP-2-treated group, whereas lamellipodia of cells in the control group were not obvious and the majority of cells tended to be rounder with shorter cell diameter. Scratch healing and transwell experiments showed that the migration and invasion capacity of BMP-2-induced MCF-7 cells were markedly enhanced compared with the control group. AFM and CLSM experiments showed that the skeletons of the BMP-2-induced MCF-7 cells became ordered, and CD44 molecule distribution on the BMP-2-induced MCF-7 cells was not uniform, many CD44 molecules gathered together. The expression of CD44 molecules on the BMP-2-induced MCF-7 cells was found to be approximately four times bigger than that of control group by western blot experiment. We also found that the expression of E-cadherin molecule decreased.BMP-2-induced human breast cancer cell MCF-7 obtained migration phenotype and cell migration and invasion capacity were enhanced compared with the control group.