New method of in vitro culturing of pigment retinal epithelium in the structure of the posterior eye sector of adult rat

Abstract

We propose a new method of organotypic roller 3D-culturing of the posterior sector of the eye. The method allows maintaining tissue viability in vitro for 14 days (which considerably surpasses the capacities of stationary culturing) and studying of the behavior, of pigment retinal epithelial cells and choriocapillary membrane. Using this method we demonstrated phenotypic transformation, migration, and proliferation of pigment retinal epithelial cells under conditions of roller organotypic culture. In the absence of the retina, these cells exhibit properties of scavenger cells (phagocytes) both within and outside the layer. Under conditions of roller culturing in vitro, cells of the pigment retinal epithelium undergo changes similar to those observed in various retinal pathologies in vivo, including age-associated changes. Here we discuss the possibility of using the proposed method for evaluation of the effect of various factors added to the culture medium on the pigment epithelium, for modeling of processes developing in damaged pigment epithelium or under conditions of various pathologies, and for the study of regeneration responses in cells of pigment retinal epithelium in adult vertebrates.

New antioxidant SkQ1 is an effective protector of rat eye retinal pigment epithelium and choroid under conditions of long-term organotypic cultivation

“…A number of in vitro and in vivo models have been employed to find the protective effects of various antioxidants upon the retina. In this work, we used an organotypic cultivation of the rat posterior eye cups, consisting of the RPE ↔ choroid ↔ sclera complex, in a roller [20], to examine the effect of the new antioxidant SkQ1 [8,9] upon RPE and choroid. The advantage of this method as compared with any of in vivo models is its relative simplicity and convenience.…”

“…Maintenance, anaesthesia and utilization of the animals were carried out in accordance with bioethics regulations of Russian Academy of Sciences. The procedures of eye cups obtaining, rotary cultivation, samples' fixation as well as the methods of the study of eye cup tissues are described in detail in [20]. Briefly, the eye anterior segment, containing the cornea, iris, and lens, was removed by means of microsurgery.…”

New antioxidant SkQ1 is an effective protector of rat neural retina under conditions of long-term organotypic cultivation

“…All animal experiments, including maintenance, narcotization and utilization, were carried out in accordance with bioethics regulations of Russian Academy of Sciences and approved by the Laboratory Animal Care and Use Committee of the Kol'tsov Institute of Developmental Biology, Russian Academy of Science. Procedures of eye tissues isolation, rotary culturing, sample fixation as well as the methods of following preparation of cultured eye cups are described in detail in [14]. Briefly, the eye anterior segment containing the cornea, iris and lens was removed surgically.…”

Cell Sources for Retinal Regeneration: Implication for Data Translation in Biomedicine of the Eye

“…According to our observations during experiments with organotypic 3D cultivation of the posterior sector of the rat eye, the RPE layer under conditions of partial death of its cells still continued to maintain the epithelial structure for a long time. This occurs through the increase in the size of cells and their stretching over the layer; however, such RPE cells cannot establish a normal functional connection with photoreceptors, simultaneously undergoing death [ 178 ]. The models of in vivo RPE damage in mice showed an increase in the size of RPE cells and also in the proportion of multinucleated cells [ 179 , 180 ].…”