Journal
STEM CELLS
Volume 30, Issue 4, Pages 673-686Publisher
WILEY
DOI: 10.1002/stem.1037
Keywords
Embryonic stem cell; Photoreceptor; Retinal pigmented epithelium; Outer retinal degeneration
Categories
Funding
- Fight for Sight UK
- Newcastle Health Charity
- Sunderland Eye Infirmary
- Newcastle University
- UK NIHR Biomedical Research Centre for Ageing and Age-related disease
- Newcastle upon Tyne NHS Hospitals Trust
- Conselleria de Sanidad (Generalitat Valenciana)
- Instituto de Salud Carlos III (Ministry of Science and Innovation)
- BBSRC [BB/I02333X/1, BB/E012841/1] Funding Source: UKRI
- MRC [G0301182] Funding Source: UKRI
- Biotechnology and Biological Sciences Research Council [BB/E012841/1, BB/I02333X/1] Funding Source: researchfish
- Fight for Sight [1870] Funding Source: researchfish
- Medical Research Council [G0301182] Funding Source: researchfish
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Recent successes in the stem cell field have identified some of the key chemical and biological cues which drive photoreceptor derivation from human embryonic stem cells (hESC) and human induced pluripotent stem cells (hiPSC); however, the efficiency of this process is variable. We have designed a three-step photoreceptor differentiation protocol combining previously published methods that direct the differentiation of hESC and hiPSC toward a retinal lineage, which we further modified with additional supplements selected on the basis of reports from the eye field and retinal development. We report that hESC and hiPSC differentiating under our regimen over a 60 day period sequentially acquire markers associated with neural, retinal field, retinal pigmented epithelium and photoreceptor cells, including mature photoreceptor markers OPN1SW and RHODOPSIN with a higher efficiency than previously reported. In addition, we report the ability of hESC and hiPSC cultures to generate neural and retinal phenotypes under minimal culture conditions, which may be linked to their ability to endogenously upregulate the expression of a range of factors important for retinal cell type specification. However, cultures that were differentiated with full supplementation under our photoreceptor-induction regimen achieve this within a significantly shorter time frame and show a substantial increase in the expression of photoreceptor-specific markers in comparison to cultures differentiated under minimal conditions. Interestingly, cultures supplemented only with B27 and/or N2 displayed comparable differentiation efficiency to those under full supplementation, indicating a key role for B27 and N2 during the differentiation process. Furthermore, our data highlight an important role for Dkk1 and Noggin in enhancing the differentiation of hESC and hiPSC toward retinal progenitor cells and photoreceptor precursors during the early stages of differentiation, while suggesting that further maturation of these cells into photoreceptors may not require additional factors and can ensue under minimal culture conditions. STEM CELLS 2012; 30:673686
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