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Kyle Wallace, Amelia Gerner, Jessica Martin, Zhonggang Hou, David M. Gamm; Retinal Differentiation Of Human Es Cells Maintained In Chemically Defined, Xeno-free E8 Culture Medium. Invest. Ophthalmol. Vis. Sci. 2012;53(14):5913.
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Human pluripotent stem cells (hPSCs) can be differentiated along a retinal lineage using a variety of protocols and culture conditions. However, for translational applications, it is important to avoid media that is undefined and/or contains animal proteins. E8 medium, a highly simplified version of the commonly used mTeSR1, has the advantage of being both chemically defined and xeno-free. Moreover, E8 does not require human serum albumin, which is prohibitively expensive and prone to batch variability. In this study, we compared the capacity of human ES cells (hESCs) grown in E8, mTeSR1, or MEF-conditioned medium to undergo targeted retinal differentiation.
Pluripotent WA09 hESCs were maintained using each of the following culture conditions: ESC medium on MEF feeder layers, mTESR1 on Matrigel, or E8 on Matrigel. Targeted retinal differentiation was carried out using our previously described protocol, which yields 3-D optic vesicle-like structures (OVs) comprised of multipotent neuroretinal progenitor cells. Throughout the differentiation process, cultures were monitored by light microscopy, RT- and quantitative PCR, and ICC to compare morphological features and expression profiles between experimental groups.
hESCs maintained in E8 or mTeSR1 on Matrigel displayed less spontaneous differentiation and consistently produced more OVs (~2-3-fold) than hESCs maintained in ESC medium on MEF feeder layers. OVs produced from all three groups were indistinguishable and gave rise to multiple neuroretinal cell types, including photoreceptor-like cells. Similarly, RPE was observed in all three groups when the hESCs were allowed to differentiate in adherent cultures.
E8 and mTeSR1 were equally effective at maintaining hESCs in a pluripotent state and supporting their competency to generate retinal progeny, and both were superior to ESC medium on MEF feeder layers. Therefore, E8 medium is a valuable option for maintaining hPSCs for laboratory or clinical use.
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