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Transplantation of Photoreceptor Precursors Isolated via a Cell Surface Biomarker Panel From Embryonic Stem Cell-Derived Self-Forming Retina.

Lakowski J, Gonzalez-Cordero A, West EL, Han YT, Welby E, Naeem A, Blackford SJ, Bainbridge JW, Pearson RA, Ali RR, Sowden JC - Stem Cells (2015)

Bottom Line: Cell replacement therapy, using pluripotent stem cell-derived photoreceptor cells, may be a feasible future treatment.As genetically labelled cells are not desirable for therapy, here we developed a surface biomarker cell selection strategy for application to complex pluripotent stem cell differentiation cultures.Conversely, unsorted or negatively selected cells do not give rise to newly integrated rods after transplantation.

View Article: PubMed Central - PubMed

Affiliation: Stem Cells and Regenerative Medicine Section, UCL Institute of Child Health, University College London, London, United Kingdom.

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Characterization of photoreceptor precursor (PPr) surface biomarkers on three‐dimensional embryonic stem cell (ESC) retinal cultures. (A, B): Day 12 optic vesicle neuroepithelium showing Vsx2 (green, A) and Pax6 (red, B) positive retinal progenitor cells. (C, D): Day 28 retinal neuroepithelium regions containing Rhodopsin (green, C) and Recoverin (red, D) positive ESC‐derived photoreceptors. (E, F): Immunohistochemical analysis showing a small number of cells positive for Rod α‐transducin (E) and Peripherin 2 (F) at day 28 of culture. High magnification inserts highlight expression pattern of these markers. (G): Quantitative real‐time PCR analysis of ESC retinal cultures demonstrating the expression of PPr biomarker panel over time in culture. (H): Expression of the PPr biomarker panel on day 26 fluorescent‐activated cell‐sorted Rhop.GFP+ ESC‐derived rods. (I): Immunohistochemical analysis for CD73, CD24, CD133, and CD47 (red), and Recoverin and Rhodopsin (green) on cryosections of ESC‐retinal differentiations at day 12, 28, and 36 of culture. Scale bars: 25 µm. Abbreviation: ES, embryonic stem cell (undifferentiated); PCR, polymerase chain reaction.
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stem2051-fig-0002: Characterization of photoreceptor precursor (PPr) surface biomarkers on three‐dimensional embryonic stem cell (ESC) retinal cultures. (A, B): Day 12 optic vesicle neuroepithelium showing Vsx2 (green, A) and Pax6 (red, B) positive retinal progenitor cells. (C, D): Day 28 retinal neuroepithelium regions containing Rhodopsin (green, C) and Recoverin (red, D) positive ESC‐derived photoreceptors. (E, F): Immunohistochemical analysis showing a small number of cells positive for Rod α‐transducin (E) and Peripherin 2 (F) at day 28 of culture. High magnification inserts highlight expression pattern of these markers. (G): Quantitative real‐time PCR analysis of ESC retinal cultures demonstrating the expression of PPr biomarker panel over time in culture. (H): Expression of the PPr biomarker panel on day 26 fluorescent‐activated cell‐sorted Rhop.GFP+ ESC‐derived rods. (I): Immunohistochemical analysis for CD73, CD24, CD133, and CD47 (red), and Recoverin and Rhodopsin (green) on cryosections of ESC‐retinal differentiations at day 12, 28, and 36 of culture. Scale bars: 25 µm. Abbreviation: ES, embryonic stem cell (undifferentiated); PCR, polymerase chain reaction.

Mentions: PPrs can be generated efficiently using a previously described embryoid body‐based 3D mouse ESC (mESC) differentiation system 20, 22. In this culture system, continuous neuroepithelia are readily produced within 5 days of differentiation and optic vesicle‐like structures appear around days 7–9 (Fig. 2 shows optic vesicle neuroepithelium at day 12). Retinal cell genesis proceeds in a sequence similar to normal retinal development with all neural retinal cell types being present and correctly organized in layers by day 29 of the differentiation procedure, a stage that has been shown to correlate with P4–P8 during mouse retinal development 22.


Transplantation of Photoreceptor Precursors Isolated via a Cell Surface Biomarker Panel From Embryonic Stem Cell-Derived Self-Forming Retina.

Lakowski J, Gonzalez-Cordero A, West EL, Han YT, Welby E, Naeem A, Blackford SJ, Bainbridge JW, Pearson RA, Ali RR, Sowden JC - Stem Cells (2015)

Characterization of photoreceptor precursor (PPr) surface biomarkers on three‐dimensional embryonic stem cell (ESC) retinal cultures. (A, B): Day 12 optic vesicle neuroepithelium showing Vsx2 (green, A) and Pax6 (red, B) positive retinal progenitor cells. (C, D): Day 28 retinal neuroepithelium regions containing Rhodopsin (green, C) and Recoverin (red, D) positive ESC‐derived photoreceptors. (E, F): Immunohistochemical analysis showing a small number of cells positive for Rod α‐transducin (E) and Peripherin 2 (F) at day 28 of culture. High magnification inserts highlight expression pattern of these markers. (G): Quantitative real‐time PCR analysis of ESC retinal cultures demonstrating the expression of PPr biomarker panel over time in culture. (H): Expression of the PPr biomarker panel on day 26 fluorescent‐activated cell‐sorted Rhop.GFP+ ESC‐derived rods. (I): Immunohistochemical analysis for CD73, CD24, CD133, and CD47 (red), and Recoverin and Rhodopsin (green) on cryosections of ESC‐retinal differentiations at day 12, 28, and 36 of culture. Scale bars: 25 µm. Abbreviation: ES, embryonic stem cell (undifferentiated); PCR, polymerase chain reaction.
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stem2051-fig-0002: Characterization of photoreceptor precursor (PPr) surface biomarkers on three‐dimensional embryonic stem cell (ESC) retinal cultures. (A, B): Day 12 optic vesicle neuroepithelium showing Vsx2 (green, A) and Pax6 (red, B) positive retinal progenitor cells. (C, D): Day 28 retinal neuroepithelium regions containing Rhodopsin (green, C) and Recoverin (red, D) positive ESC‐derived photoreceptors. (E, F): Immunohistochemical analysis showing a small number of cells positive for Rod α‐transducin (E) and Peripherin 2 (F) at day 28 of culture. High magnification inserts highlight expression pattern of these markers. (G): Quantitative real‐time PCR analysis of ESC retinal cultures demonstrating the expression of PPr biomarker panel over time in culture. (H): Expression of the PPr biomarker panel on day 26 fluorescent‐activated cell‐sorted Rhop.GFP+ ESC‐derived rods. (I): Immunohistochemical analysis for CD73, CD24, CD133, and CD47 (red), and Recoverin and Rhodopsin (green) on cryosections of ESC‐retinal differentiations at day 12, 28, and 36 of culture. Scale bars: 25 µm. Abbreviation: ES, embryonic stem cell (undifferentiated); PCR, polymerase chain reaction.
Mentions: PPrs can be generated efficiently using a previously described embryoid body‐based 3D mouse ESC (mESC) differentiation system 20, 22. In this culture system, continuous neuroepithelia are readily produced within 5 days of differentiation and optic vesicle‐like structures appear around days 7–9 (Fig. 2 shows optic vesicle neuroepithelium at day 12). Retinal cell genesis proceeds in a sequence similar to normal retinal development with all neural retinal cell types being present and correctly organized in layers by day 29 of the differentiation procedure, a stage that has been shown to correlate with P4–P8 during mouse retinal development 22.

Bottom Line: Cell replacement therapy, using pluripotent stem cell-derived photoreceptor cells, may be a feasible future treatment.As genetically labelled cells are not desirable for therapy, here we developed a surface biomarker cell selection strategy for application to complex pluripotent stem cell differentiation cultures.Conversely, unsorted or negatively selected cells do not give rise to newly integrated rods after transplantation.

View Article: PubMed Central - PubMed

Affiliation: Stem Cells and Regenerative Medicine Section, UCL Institute of Child Health, University College London, London, United Kingdom.

Show MeSH
Related in: MedlinePlus