Differentiation of human embryonic stem cells using size-controlled embryoid bodies and negative cell selection in the production of photoreceptor precursor cells.
We proposed to optimize the retinal differentiation protocols for human embryonic stem cells (hESCs) by improving cell handling. To improve efficiency, we first focused on the production of just one retinal precursor cell type (photoreceptor precursor cells [PPCs]) rather than the production of a range of retinal cells. Combining information from a number of previous studies, in particular the use of a feeder-free culture medium and taurine plus triiodothyronine supplements, we then assessed the values of using size-controlled embryoid bodies (EBs) and negative cell selection (to remove residual embryonic antigen-4-positive hESCs). Using size-controlled 1000 cell EBs, significant improvements were made, in that 78% CRX+ve PPCs could be produced in just 17 days. This could be increased to 93% PPCs through the added step of negative cell selection. Improved efficiency of PPC production will help in efforts to undertake shorter and larger preclinical studies as a prelude to future clinical trials.
Department of Ophthalmology and Visual Sciences, Faculty of Medicine, University of British Columbia, Vancouver, British Columbia, Canada.
Anat
A
Yanai
Christopher R J
CR
Laver
Aaron W
AW
Joe
Ishaq A
IA
Viringipurampeer
Xia
X
Wang
Cheryl Y
CY
Gregory-Evans
Kevin
K
Gregory-Evans
D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation / Q000187:drug effects / Q000235:genetics; D002460:Cell Line; D019070:Cell Lineage / Q000187:drug effects / Q000235:genetics; D048429:Cell Size; D002871:Chromosome Banding; D003470:Culture Media / Q000494:pharmacology; D058732:Embryoid Bodies / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D020869:Gene Expression Profiling; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D018189:Immunomagnetic Separation; D007621:Karyotyping; D020419:Photoreceptor Cells, Vertebrate / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism
D003470:Culture Media; D018398:Homeodomain Proteins; D015534:Trans-Activators; C109288:cone rod homeobox protein