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K. Dutt, R. Kumar, Y. Cao; Is Engineering Retina From Human Retinal Progenitors a Possibility? "Moscona Revisited". Invest. Ophthalmol. Vis. Sci. 2007;48(13):4086.
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© ARVO (1962-2015); The Authors (2016-present)
Tissue engineering is one strategy currently being explored for treatment of diseases including RP. We have previously shown that human retinal progenitor cells grown in a NASA developed bioreactor show a higher degree of differentiation confirmed by upregulation of neurotrophins (bFGF, CNTF, BDNF, etc.) and expression of retinal specific proteins. In this study, we investigated if the same could be achieved by growth in rotating vessel cultures, initially established by Moscona in 1950s.
Human retinal progenitors (cell line 205) were grown in rotating in conical flasks with and without RPE (D407). At designated times aggregates were analyzed by immunophenotyping, Western blot analysis and RT-PCR for expression of retinal specific proteins.
Cells grown in rotating cultures formed aggregates. Immunohistochemical Western blot analysis and RT-PCR confirmed the higher degree of differentiation by upregulation Aa-nat, a rate limiting enzymes in melatonin synthesis. Ganglion and bipolar cell differentiation was confirmed by upregulation of thy 1.1, neurofilament protein and PKCα upregulation, respectively. Photoreceptor differentiation in these aggregates was confirmed by expression of rhodopsin kinase, IRBP and transcription factor Nrl by RT-PCR.
The retinal progenitor cell line is faithful to its genetic program at the time of immortalization and is capable of generating several retinal cell types confirmed by photoreceptor specific protein expression (Aa-nat, rhodopsin kinase, IRBP, D2, D3, D4 receptors) ganglion cell (thy 1.1) and bipolar cell (PKCα) expression. Fidelity with which retinal progenitors (cell line) reproduce their genetic program holds promise for the other retinal progenitor cells for therapeutics.
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