Enriched Cultures of Retinal Cells From BJNhem20 XXXX Embryonic Stem Cell Line of Indian Origin.

@entity1 @entity11178 @entity25667 @entity8695 @entity16527 @entity14383 @entity13464 @entity31923 @entity538 @entity6286 @entity24739 @entity21107 @entity32846 @entity6484 @entity15470 @entity6678 @entity11442 @entity16906 @entity33444
Purpose: To test the retinal differentiation potential and to establish an optimized protocol for enriching retinal cells from an Indian origin, @entity1 embryonic stem cell (hESC) line, BJNhem20. Methods: The BJNhem20 cells were cultured and expanded under feeder-free culture conditions. Differentiation was initiated by embryoid body (EB) formation and were cultured on Matrigel in neural induction medium (NIM) for 1 week and further maintained in retinal differentiation medium (RDM). After 1 month, the neuro-retinal progenitor clusters located at the center of pigmented retinal patches were picked and cultured as suspended neurospheres in RDM for 3 days and subsequently on Matrigel in neuro-retinal medium. The mildly pigmented, immature retinal pigmented epithelial (RPE) cells were picked separately and cultured on Matrigel in RPE medium (RPEM). After 1 week, the confluent neuro-retinal and RPE cultures were maintained in RDM for 2 to 3 months and characterized by immunofluorescence and RT-PCR. Results: The BJNhem20 cells efficiently differentiated into both neuro-retinal and RPE cells. The early retinal progenitors expressed Nestin, @entity538 , @entity6484 , Rx, MitfA, @entity16527 , and @entity15470 . Neuro-retinal cells expressed the neural markers, @entity14383 , b-III tubulin, acetylated tubulin and photoreceptor-specific markers, @entity13464 , @entity8695 , @entity24739 , calbindin, PKC, @entity32846 , @entity21107 , @entity8695 kinase, @entity25667 , and @entity33444 . Mature RPE cells developed intense pigmentation within 3 months and showed @entity11178 and @entity6286 staining at cell-cell junctions and expressed @entity31923 , @entity11442 , @entity6678 , @entity16906 , and displayed phagocytic activity. Conclusions: This study confirms the retinal differentiation potential of BJNhem20 cells and describes an optimized protocol to generate enriched populations of neuro-retinal and RPE cells.
