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Memory in Induced Pluripotent Stem Cells: Reprogrammed Human Retinal Pigmented Epithelial Cells Show Tendency for Spontaneous Redifferentiation

From this month’s edition of Stem Cells

By Carla Mellough

Elucidation of the best methods for somatic cell reprogramming which fully harness the stem cell-like capacity of induced pluripotent stem cells (iPSCs) is key prior to the utilisation of iPSC for regenerative medicine purposes. The translation of iPSC therapy from the bench to the clinic has so far been slowed by limitations in the efficiency and safety of reprogramming protocols, alongside recent reports which indicate that although reprogrammed cells exhibit numerous properties of human embryonic stem cells (hESC), not all iPSC are fully reprogrammed and significant differences still remain1-4, with many studies now linking the profile of iPSC to their cell of origin. Adding to this body of evidence, results published in the November issue of Stem Cells by Hu et al.5 From the University of California Santa Barbara, assess the retention of epigenetic imprints in iPSC generated from retinal pigmented epithelial (RPE) cells and whether these iPSC have a propensity to redifferentiate back into RPE cells – their cell of origin.

RPE cells are important structural and support cells of the retina and a dysfunctional RPE is a major contributing factor in various forms of blindness. Using lentiviral delivery of the four Thomson factors (OCT4, Nanog, SOX2 and LIN28) into human fetal RPE, Hu et al. generated iPSC lines, three of which were focused on in the study due to their high hESC similarity. The cells displayed hESC markers, retained a normal karyotype and after 14 passages the authors report that transgene expression of the Thompson factors had been silenced and was thus unnecessary for further propagation of the iPSC lines. Analysis of DNA methylation showed, akin to hESCs, hypomethylation of the promoter region of both OCT4 and Nanog. The iPSC were able to generate cells from all three germ layers following embryoid body (EB) mediated differentiation in vitro and teratoma formation in vivo. Following spontaneous differentiation in vitro colonies of pigmented cells appeared which could be expanded in RPE-supportive medium to form sheets of pigmented, polygonal cells like native RPE. These expressed RPE genes and displayed phagocytic activity, a vital process for normal RPE function to rid of shed photoreceptor outer segments.

When the authors compared the number of colonies that generated pigmented cells from spontaneously differentiating iPSC derived from RPE with iPSC generated from lung fibroblasts, foreskin fibroblasts or hESC lines, they observed significant differences in two of the three RPE-derived lines. These lines displayed at least four-fold (60-70%) the number of colonies containing pigmented cells when compared with hESC, with even fewer pigmented colonies observed in lung- or foreskin-derived iPSC populations undergoing differentiation. QPCR analysis demonstrated elevation of RPE gene expression at early stages of spontaneous differentiation in RPE-derived iPSC compared to the other iPSC lines. However bisulphite sequencing analysis of key retinogenic transcription factor DNA methylation patterns demonstrated a similarity between RPE-derived iPSC and hESCs, indicating no retention of RPE patterning as far as the authors studied. Clearly further epigenetic characterisation is necessary to clarify the basis of these observations.

This study reports that in some, but not all, iPSC lines generated from RPE, colonies showed a higher propensity to differentiate once more down the RPE lineage. The number of colonies generating pigmented cells in this study was higher than reports which try to yield enriched populations of RPE from hESC using defined protocols designed to mimic in vivo RPE development6,7. Indeed, iPSC generated from differing somatic cell types and/or under different protocols do exhibit varying differentiation potential. It may then seem sensible to generate iPSCs from a cell type of origin that is related to the cell type desired for subsequent use. Although the field of iPSC remains a work in progress, tailoring iPSC generation for specific patient needs based on current knowledge may accelerate their use for regenerative medicine.

Read this article in the November edition of Stem Cells.



1. Chin MH et al. Induced pluripotent stem cells and embryonic stem cells are distinguished by gene expression signatures. Cell Stem Cell. 2009 Jul 2;5(1):111-23.

2. Marchetto MC et al. Transcriptional signature and memory retention of human-induced pluripotent stem cells. PLoS One. 2009 Sep 18;4(9):e7076.

3. Kim K et al. Epigenetic memory in induced pluripotent stem cells. Nature. 2010 Sep 16;467(7313):285-90.

4. Polo JM et al. Cell type of origin influences the molecular and functional properties of mouse induced pluripotent stem cells. Nat Biotechnol. 2010 Aug;28(8):848-55.

5. Hu Q, Friedrich AM, Johnson LV, Clegg DO. Memory in Induced Pluripotent Stem Cells: Reprogrammed Human Retinal-Pigmented Epithelial Cells Show Tendency for Spontaneous Redifferentiation. Stem Cells. 2010 Nov;28(11):1981-1991.

6. Osakada F et al. Toward the generation of rod and cone photoreceptors from mouse, monkey and human embryonic stem cells. Nat Biotechnol. 2008 Feb;26(2):215-24.

7. Osakada F et al. Stepwise differentiation of pluripotent stem cells into retinal cells. Nat Protoc. 2009;4(6):811-24.