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Experimental Study of the Biological Properties of Human Embryonic Stem Cell – Derived Retinal Progenitor Cells

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ABSTRACT

Retinal degenerative diseases are among the leading causes of blindness worldwide, and cell replacement is considered as a promising therapeutic. However, the resources of seed cells are scarce. To further explore this type of therapy, we adopted a culture system that could harvest a substantial quantity of retinal progenitor cells (RPCs) from human embryonic stem cells (hESCs) within a relatively short period of time. Furthermore, we transplanted these RPCs into the subretinal spaces of Royal College of Surgeons (RCS) rats. We quantified the thickness of the treated rats’ outer nuclear layers (ONLs) and explored the visual function via electroretinography (ERG). It was found that the differentiated cells expressed RPC markers and photoreceptor progenitor markers. The transplanted RPCs survived for at least 12 weeks, resulting in beneficial effects on the morphology of the host retina, and led to a significant improvement in the visual function of the treated animals. These therapeutic effects suggest that the hESCs-derived RPCs could delay degeneration of the retina and partially restore visual function.

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The expression level of proteins and genes of retinal progenitor cells during the differentiated development of hESCs analyzed by Western blot and real-time PCR.(A) Western blot analysis of retinal progenitor cell markers (Pax6, Sox2, Rax, Nestin, Otx2, Crx, and Recoverin) during their differentiated development. The gels in this experiment were run under the same experimental conditions. (B) Statistical analysis of the expression levels of proteins in retinal progenitor cell markers (Pax6, Sox2, Rax, Nestin, Otx2, Crx, and Recoverin). Data expressed as mean ± SD. (C) Statistical analysis of the expression of mRNA in retinal progenitor cell markers (Pax6, Sox2, Rax, Nestin, Otx2, Crx, and Recoverin). Data from at least three independent experiments are represented as the mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001, versus with data of previous time point.
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f4: The expression level of proteins and genes of retinal progenitor cells during the differentiated development of hESCs analyzed by Western blot and real-time PCR.(A) Western blot analysis of retinal progenitor cell markers (Pax6, Sox2, Rax, Nestin, Otx2, Crx, and Recoverin) during their differentiated development. The gels in this experiment were run under the same experimental conditions. (B) Statistical analysis of the expression levels of proteins in retinal progenitor cell markers (Pax6, Sox2, Rax, Nestin, Otx2, Crx, and Recoverin). Data expressed as mean ± SD. (C) Statistical analysis of the expression of mRNA in retinal progenitor cell markers (Pax6, Sox2, Rax, Nestin, Otx2, Crx, and Recoverin). Data from at least three independent experiments are represented as the mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001, versus with data of previous time point.

Mentions: We examined the protein expressions of Pax6, Sox2, Rax, Nestin, Otx2, Crx, and Recoverin on the 0, 10th, 20th, 30th, 40th days of differentiation by Western blot and real-time PCR. At the 10th day, the expressions of retinal progenitor cell markers (Rax and Nestin) reached crest, and then decreased over time. The expressions of retinal progenitor cell markers Pax6 and Rax increased over time, reached top at the 20th day, and then decreased over time. The expressions of photoreceptor precursor cell marker Otx2 reached its crest at 20th day. In contrast, Crx and Recoverin, which were also markers of photoreceptor cells progressively increased over time (Fig. 4).


Experimental Study of the Biological Properties of Human Embryonic Stem Cell – Derived Retinal Progenitor Cells
The expression level of proteins and genes of retinal progenitor cells during the differentiated development of hESCs analyzed by Western blot and real-time PCR.(A) Western blot analysis of retinal progenitor cell markers (Pax6, Sox2, Rax, Nestin, Otx2, Crx, and Recoverin) during their differentiated development. The gels in this experiment were run under the same experimental conditions. (B) Statistical analysis of the expression levels of proteins in retinal progenitor cell markers (Pax6, Sox2, Rax, Nestin, Otx2, Crx, and Recoverin). Data expressed as mean ± SD. (C) Statistical analysis of the expression of mRNA in retinal progenitor cell markers (Pax6, Sox2, Rax, Nestin, Otx2, Crx, and Recoverin). Data from at least three independent experiments are represented as the mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001, versus with data of previous time point.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
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getmorefigures.php?uid=PMC5304228&req=5

f4: The expression level of proteins and genes of retinal progenitor cells during the differentiated development of hESCs analyzed by Western blot and real-time PCR.(A) Western blot analysis of retinal progenitor cell markers (Pax6, Sox2, Rax, Nestin, Otx2, Crx, and Recoverin) during their differentiated development. The gels in this experiment were run under the same experimental conditions. (B) Statistical analysis of the expression levels of proteins in retinal progenitor cell markers (Pax6, Sox2, Rax, Nestin, Otx2, Crx, and Recoverin). Data expressed as mean ± SD. (C) Statistical analysis of the expression of mRNA in retinal progenitor cell markers (Pax6, Sox2, Rax, Nestin, Otx2, Crx, and Recoverin). Data from at least three independent experiments are represented as the mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001, versus with data of previous time point.
Mentions: We examined the protein expressions of Pax6, Sox2, Rax, Nestin, Otx2, Crx, and Recoverin on the 0, 10th, 20th, 30th, 40th days of differentiation by Western blot and real-time PCR. At the 10th day, the expressions of retinal progenitor cell markers (Rax and Nestin) reached crest, and then decreased over time. The expressions of retinal progenitor cell markers Pax6 and Rax increased over time, reached top at the 20th day, and then decreased over time. The expressions of photoreceptor precursor cell marker Otx2 reached its crest at 20th day. In contrast, Crx and Recoverin, which were also markers of photoreceptor cells progressively increased over time (Fig. 4).

View Article: PubMed Central - PubMed

ABSTRACT

Retinal degenerative diseases are among the leading causes of blindness worldwide, and cell replacement is considered as a promising therapeutic. However, the resources of seed cells are scarce. To further explore this type of therapy, we adopted a culture system that could harvest a substantial quantity of retinal progenitor cells (RPCs) from human embryonic stem cells (hESCs) within a relatively short period of time. Furthermore, we transplanted these RPCs into the subretinal spaces of Royal College of Surgeons (RCS) rats. We quantified the thickness of the treated rats&rsquo; outer nuclear layers (ONLs) and explored the visual function via electroretinography (ERG). It was found that the differentiated cells expressed RPC markers and photoreceptor progenitor markers. The transplanted RPCs survived for at least 12 weeks, resulting in beneficial effects on the morphology of the host retina, and led to a significant improvement in the visual function of the treated animals. These therapeutic effects suggest that the hESCs-derived RPCs could delay degeneration of the retina and partially restore visual function.

No MeSH data available.


Related in: MedlinePlus