Research on EMT Phenomena Occurred in Human Colorectal Carcinoma Cells, Induced by Chemokine CCL20
|School||Kunming Medical College|
|Keywords||CCL20 EMT Transwell Caco-2 SW480 LoVo|
Objective To observe that whether the human colorectal cancer cells, Caco-2, SW480 and LoVo, will make the epithelial-mesenchymal transition (epithelial to mesenchymal transition, EMT)when stimulated by the different concentrations of chemokine CCL20 cell. And to observe the effect of different concentrations of CCL20 cell on the movement ability of the three cells by establishing chamber of Transwell and simulating internal environment in vitro.Method The three cells in logarithmic growth phase are observed after being starved for 12,24 hours under inverted microscope when stimulated by different concentrations of CCL20 (10ng/mL, 100ng/mL, in 1000ng/mL) to survey the morphological changes, immunohistochemistry(IHC), immunofluorescence(IMF) and western blot, and to detect the expression change of E-cadherin(E-cad)and Vimentin (Vim) before and after simulation. The migration assay and invasion assay are carried out through Transwell experiments by the simulation of CCL20.Result The three cells in the stimulation of CCL20 (10ng/mL,100ng/mL 1000ng/mL) have no obvious change in morphology, and the cells spacing becomes loose only. Detected by the IHC, IMF and western blot, there is some difference in the protein expression of E-cad and Vim before and after the stimulation, but not statistically significant(P>0.05). In the Transwell champer experiment, compared with other culture conditions, the migration and invasion abilities of the three cells are strengthened in the stimulation of CCL20, and the difference is statistically significant (P<0.01). The chemotactic ability of CCL20 of 100ng/mL is strengthened in contrast to that of 10ng/mL, and the difference is statistically significant (P<0.05). As for CCL20 of 1000ng/mL, compared with CCL20 of 100ng/mL, the difference is statistically significant (P<0.05).Conclusion 1. CCL20 cannot induce Caco-2, SW480 and LoVo to make EMT transition.2. CCL20 presents chemotaxis to the three cells, and the chemotaxis strengthens as the concentration increases in the range 10ng/mL to 1000ng/mL.