Phenotypic Plasticity of Retinal Pigment Epithelial Cells from Adult Human Eye In Vitro

Abstract

Phenotypic plasticity of retinal pigment epithelial cells from adult human eye was studied by immunohistochemical methods under different culturing conditions. It was found that retinal pigment epithelium in adult human eye is a heterogeneous population of cells demonstrating different behavior in vitro. Some cells retain epithelial morphology for a long time in culture, while others are rapidly transformed into fibroblast-like cells and synthesize proteins typical of proneural, neural, glial, and photoreceptor cells. However, irrespective of initial morphological features differentiation of retinal pigment cells can be modulated by varying culturing conditions.

Cell Models to Study Regulation of Cell Transformation in Pathologies of Retinal Pigment Epithelium

“…The resulting cup-shaped segment with RPE on the inner surface is filled with a cell dissociation reagent and incubated at 37°C or room temperature for 8 min to 1 hour. Suitable dissociation reagents include solutions of pronase, papain, trypsin, hialuronidase/collagenase, or dispase [20–24] or of nonenzymatic substances such as EDTA [25, 26]. The solutions are usually prepared in calcium- and magnesium-free Hank's balanced salt solution (HBSS), and the incubation regime depends on the reagent used.…”

“…The effects of some media supplements on the improvement of barrier properties of the ARPE-19 cell monolayer are described in detail in the study by Mannermaa [31]. Typical supplements used in primary RPE cell cultures include basic fibroblast growth factor (bFGF) and optimized commercial mixtures such as N1 Supplement (containing transferrin, insulin, putrescine, progesterone, selenium, and biotin) or N2 Supplement (based on N1 Supplement without biotin) [25, 26, 28, 49]. Other supplements and different glucose concentrations have also been tested, but there still are no systematic data or any definitive conclusions on the role of media supplements in the properness of cultured RPE-derived cells [31].…”

“…All these events also take place in RPE cells grown in culture flasks. Thus, human RPE cells grown in vitro show distinct positive staining for vimentin [73] and synthesize various ECM molecules such as tissue inhibitor of metalloproteinase 3 (TIMP-3) [74]; collagen types I [26], IV, and V [75]; laminin [76]; fibronectin [26, 77, 78]; heparan sulfate proteoglycan; and hyaluronic acid [79, 80]. …”

“…Moreover, these cells are capable of subsequent transdifferentiation into neural cells, as indicated by the expression of Musashi 1, Pax6, and TUBB3 (Figure 1(a)) and positive staining with antibodies against protein markers of neuronal differentiation—nestin, TUBB3 (Figure 1(b)), tyrosine hydroxylase, neurofilaments 68 and 200 (Figure 1(c)), and nNOS—and glial differentiation (CNPase, GFAP) [25, 26, 87, 88]. RPE cells in vitro also show positive staining for vimentin, a marker of intermediate filaments [26, 73]. Simultaneous expression of vimentin and intermediate filament proteins of other classes, nestin and GFAP, observed in RPE cell culture [26] is also characteristic of human neural stem cells [89].…”

“…RPE cells in vitro also show positive staining for vimentin, a marker of intermediate filaments [26, 73]. Simultaneous expression of vimentin and intermediate filament proteins of other classes, nestin and GFAP, observed in RPE cell culture [26] is also characteristic of human neural stem cells [89]. This fact indicates that RPE cells in vitro are apparently multipotent.…”