Retinal tissue engineering focuses on the development of biomaterials. It provides potent promises for regeneration of neuroretinal cells in degenerative eye diseases. Biomimetic three-dimensional (3D) microenvironments and specific growth factors motivate differentiation of human retinal pigment epithelial (hRPE) cells toward retinal neural lineage. In this study, we evaluated alginate/gelatin (A/G) as a substrate for cultures of hRPE cells.

RPE was isolated from neonatal human globes and cultured in DMEM/F12 supplement with 10% FBS. Alginate / gelatin substrate with 20:80 weight ratios were prepared. Passage 4 of RPE cells was cultivated on substrate and proliferation of cells was determined in different culture conditions of DMEM/F12; DMEM/F12 supplement with 10% FBS or DMEM/F12 supplemented with 30% HAF, by MTT and cell proliferation assay. Immunocytochemistry and real-time RT-PCR were performed to evaluate the effect of substrate on RPE cells differentiation.

Significant increase in cell proliferation rate was detected in hRPE cells cultivated on an A/G substrate. Continuous observations demonstrated that hRPE cells formed densely packed, suspended colonies in DMEM/F12 culture condition, and that there was a dominant trans-differentiation into amacrine, rod photoreceptors, bipolar cells, and retinal ganglion cells. Small adhered clusters of hRPE cells in HAF- and FBS-treated cultures represented de-differentiation toward retinal progenitor cells. These cultures generated amacrine, rod photoreceptors and bipolar cells when assessed by immustaining.

These findings indicated that A/G substrate induced neural retinal cell propagation in cultures and would therefore be promising for RPE-based tissue engineering studies.