Eye diseases such as age-related macular degeneration or diabetes affect retinal pigment epithelium (RPE) function and lead to retinal degeneration, vision loss and blindness. To study RPE function, physiology, and pathology, many laboratories have attempted to culture RPE as a more accessible alternative to native tissue. This goal has been accomplished with varying degrees of success due to the functional and morphological complexity of the RPE and its neighboring cells in the retina and the choroid. We have developed techniques for culturing confluent monolayers of human fetal RPE cells that exhibit morphology, physiology, and patterns of protein expression similar to native human fetal RPE. One of the goals of this work was to identify a set of commercially available ingredients to create stable and reproducible RPE cell cultures. We been able to produce confluent pigmented RPE cell cultures with classic epithelial morphology, transepithelial potential of 1 - 3mV, and transepithelial resistance greater than 400 Ohms*cm2. In the present experiments we further characterized these cultures using electron-microscopy and immunohistochemistry to identify cell structures, and localize apical and basolateral membrane and intercellular junctional complex proteins. ELISAs were used to confirm the polarity of secretion of selected cytokines. Intracellular microelectrodes were used to characterize receptor-mediated second messenger pathways and their downstream electrophysiological properties at the apical and basolateral membranes. The capacitance probe technique was used to measure net transepithelial fluid transport. This model has been used to help identify some of the mitochondrial pathways that mediate oxidative stress and the protective effects of lipoic acid.
| Verma, Varun; Sauer, Theodor; Chan, Chi-Chao et al. (2008) Constancy of ERp29 expression in cultured retinal pigment epithelial cells in the Ccl2/Cx3cr1 deficient mouse model of age-related macular degeneration. Curr Eye Res 33:701-7 |
| Maminishkis, Arvydas; Chen, Shan; Jalickee, Stephen et al. (2006) Confluent monolayers of cultured human fetal retinal pigment epithelium exhibit morphology and physiology of native tissue. Invest Ophthalmol Vis Sci 47:3612-24 |
