Novo modelo de interação célula epitelial e célula hematopoiética utilizando a linhagem HEK 293

Abstract

Cell adhesion is fundamental in cells interaction to form different tissues, cellular motility and immune response of living beings. The assembly of these tissues, and their organization into organs, are determined by molecular interactions at cellular level, due to the expression of a large number of adhesion molecules. Cell-cell adhesion models using cell culture, are very important to studying several diseases, such as ischemia, inflammatory diseases and cancer. There are few cellular models of cell adhesion and the most used add Jurkat cells to activated endothelial cells such as HUVEC, CACO-2 or mesenchymal cells. An alternative model for endothelial-hematopoietic cell interaction may be obtained using HEK 293 cells, an endothelial cell line widely used as a tool for recombinant proteins expression. Since increased expression of adhesion cells plays a critical role in the pathogenicity of several diseases, the development of cellular interaction new models with cells easily culturable and able for genetic manipulation become necessary. Thus, this work aimed to establish a new model of study for epithelial cell and hematopoietic cell interaction using the HEK 293 cell line. We tested different HEK 293 concentrations cultured for 24 and 48 hours, in order to obtain the best cell concentration in which the adherent cells did not release from the culture dish in the washes. We also carried out staining tests with different concentrations of rhodamine B and carboxyfluorescein, to establish the best labeling for both cell lines. The cell-cell interaction assay was performed and analyzed by fluorescence microscopy. Results obtained suggested that ideal concentration of HEK 293 cells was 5x104 cells / ml in 24 hours, demonstrating sufficient confluence for remain cells adherence during the well washings. The best labeling for the endothelial line was 40μM rhodamine B consonant with our tests, and for the lymphocytic cell line 500nM of carboxyfluorescein were sufficient. The interaction between cells was achieved at different concentrations, and the distinction between the lines was possible because the dyes had different and non-overlapping emission lengths. Therefore, the dyes choice and their respective concentrations were efficient for this interaction model; however, further studies using at HEK 293 cells will be necessary in order to increase the cell adhesion and to develop a quantitative method to evaluate the interaction in future experiments.