A new method of culturing and transferring IRIS pigment epithelium

Abstract

Purpose. To optimize the method of c ilturing and transferring of iris pigment epithelial (IPE) cells for cellular studie; in vitro or for transplantation in vivo. Methods. Porcine iris tissues were obtiined and IPE cells were isolated and cultured at high densities by plating them in a drop-form fashion. After seven to ten days, spherical-shaped structures conta rung high concentration of cells were obtained. Subcultures of these cells wei e performed by direct pipetting of the spheres into new culture dishes without employing enzymatic dissociation of the spheres from the plates. Immunohistochemical and light and electron microscopic analyses were carried out. Results: IPE cells, when cultured at high densities, tended to form elevated spherical structures containing viable colls. The cultured cells were pigmented and showed positive labeling with monoclonal cytokeratin antibody. They proliferated and migrated from the spheres to form nonolayers. The spheres could be easily aspirated and transferred from one culture dish to another. IPE cells originating from the transferred spheres continued to proliferate and migrate in a similar fashion to the originally cultivated cells to form monolayers after seven to ten days. Conclusions: This new method provices a simple means of cultunng a high quantity of IPE cells. Its high yield of lure IPE cells and the ease of cell transfer provide an ideal source for their deliveiy into the subretinal space in vivo or tor their study at the cellular level in vitro.