Human Amniotic Fluid Promotes Retinal Pigmented Epithelial Cells'Trans-Differentiation into Rod Photoreceptors and Retinal Ganglion Cells

Abstract

To evaluate the effect of human amniotic fluid (HAF) on retinal pigmented epithelial cells growth and trans-differentiation into retinal neurons, retinal pigmented epithelium (RPE) cells were isolated from neonatal human cadaver eye globes and cultured in Dulbecco's modified Eagle's medium-F12 supplemented with 10% fetal bovine serum (FBS). Confluent monolayer cultures were trypsinized and passaged using FBS-containing or HAF-containing media. Amniotic fluid samples were received from pregnant women in the first trimester of gestation. Cell proliferation and death enzyme-linked immunosorbent assays were performed to assess the effect of HAF on RPE cell growth. Trans-differentiation into rod photoreceptors and retinal ganglion cells was also studied using immunocytochemistry and real-time polymerase chain reaction techniques. Primary cultures of RPE cells were successfully established under FBS-containing or HAF-containing media leading to rapid cell growth and proliferation. When RPE cells were moved to in vitro culture system, they began to lose their differentiation markers such as pigmentation and RPE65 marker and trans-differentiated neural-like cells followed by spheroid colonies pertaining to stem/progenitor cells were morphologically detected. Immunocytochemistry (ICC) analysis of HAF-treated cultures showed a considerable expression of Rhodopsin gene (30% Rhodopsin-positive cells) indicating trans-differentiation of RPE cells to rod photoreceptors. Real-time polymerase chain reaction revealed an HAF-dose-dependant expression of Thy-1 gene (RGC marker) and significant promoting effect of HAF on RGCs generation. The data presented here suggest that HAF possesses invaluable stimulatory effect on RPE cells growth and trans-differentiation into retinal neurons. It can be regarded as a newly introduced enriched supplement in serum-free kinds of media used in neuro-retinal regeneration studies.

Enhanced generation of retinal progenitor cells from human retinal pigment epithelial cells induced by amniotic fluid

“…These experiments confirmed that retinal progenitor cells are able to generate retinal terminally differentiated cells such as bipolar cells, amacrine cells, rod photoreceptors and retinal ganglion cells [39]. …”

“…We also examined cultures for their ability to differentiate into other cellular components of the retinal layer; specific retinal cell markers (PKC and CRABPI, (unpublished data) Rod and Thy1.1 [39]) were examined in the presence of AF using immunocytochemistry and real-time PCR. These experiments confirmed that retinal progenitor cells are able to generate retinal terminally differentiated cells such as bipolar cells, amacrine cells, rod photoreceptors and retinal ganglion cells [39].…”

Alginate as a Cell Culture Substrate for Growth and Differentiation of Human Retinal Pigment Epithelial Cells

“…Previous investigations have confirmed that growth factors possess an effective role in stimulating neural differentiation [29].…”

Targeting orphan G protein‐coupled receptors for the treatment of diabetes and its complications: C‐peptide and GPR146

“…In addition to its metabolic roles in the retina, the RPE possesses the unique ability to transdifferentiate into other retinal cell types, including photoreceptors and ganglion cells . The physiological significance of RPE transdifferentiation has not been fully elucidated, but this ability could be involved in retinal repair, with transdifferentiating RPE cells serving as a ‘stopgap’ when other retinal cell types have been damaged .…”