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D. A. Carter, E. J. Mayer, A. D. Dick; CD133+ Progenitor Cell Populations Isolated From Adult Human Postmortem Retina Are Maintained in an Undifferentiated State in the Presence of Leukaemia Inhibitory Factor (LIF). Invest. Ophthalmol. Vis. Sci. 2007;48(13):4100.
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CD133 is used to identify haematopoietic stem and progenitor cells and subsequent to our original observations that neuospheres derived from adult human retina express CD133, our aim was to isolate a purified CD133+ population of progenitor cells which maintain an ability to generate neurospheres and differentiate into cells of retinal lineage. Given that LIF: maintain ES cells in an undifferentiated state and promote their renewal, we wished to determine if we could increase cell yield or maintain prolonged cultures of retinal progenitors with LIF
Retinal cell suspensions were derived from adult human post-mortem tissue with ethical approval and CD133+ cells were flow cytometrically or Automated Magnetic Cell (MACS) sorted. Recombinant LIF was added to cultures of CD133+ sorted cells. Rates of neurosphere generation and flow cytometric phenotype analysis were determined along with subsequent cell differentiation, when neurospheres were seeded in a 3D collagen gel matrix.
Successful isolation of viable CD133+ cells was achieved via both flow cytometry and MACS and represented less than 1% of retinal cells. Sorted CD133+ cells generated neurospheres and differentiated in 3D collagen gels. In the presence of both LIF and FGF, CD133+ cells survived but neither differentiated or generated neurospheres. Within 3 days of LIF removal, CD133+ cells rapidly generated neurospheres at a similar rate as FGF supplemented cultures.
The adult human retina possesses a population of CD133 progenitor cells capable of neurosphere formation and differentiation within collagen matrix gels. Sorted Cells were maintained in LIF and on withdrawal rapidly increased their neurosphere generation with subsequent retinal cell lineage differentiation in collagen gels.
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