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Induced pluripotent stem cells restore function in a human cell loss model of open-angle glaucoma.

Abu-Hassan DW, Li X, Ryan EI, Acott TS, Kelley MJ - Stem Cells (2015)

Bottom Line: A notable feature associated with glaucoma is outflow pathway cell loss.We then differentiated induced pluripotent stem cells (iPSCs) and used them to repopulate this cell depletion model.These differentiated cells (TM-like iPSCs) became similar to TM cells in both morphology and expression patterns.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology, Casey Eye Institute, Oregon Health & Science University, Portland, Oregon, USA; Department of Biochemistry & Molecular Biology, Oregon Health & Science University, Portland, Oregon, USA; Department of Biochemistry & Physiology, University of Jordan, Amman, Jordan.

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Differentiated TM-like iPSCs can perform phagocytosis, a typical TM cell property. (A): iPSCs, HTM, and differentiated TM-like iPSCs cultured on chamber slides were incubated with fluorescent-labeled zymosan particles for an hour. They were then washed, fixed, and immunostained with LAMP1, a lysosomal marker, to verify internalization. The colocalization (yellow; examples marked with white arrowheads) of zymosan particles (green) with LAMP1 (red) indicates the phagocytosis of the particles. Both HTM and TM-like iPSCs phagocytosed the particles but iPSCs did not. Nuclei were demarcated by DAPI staining (blue). Scale bar represents 100 µm. (B): The number of zymosan particles that colocalize with LAMP1 was counted and the total number was divided by total number of cells in the field. Significance is indicated by * where p < .05 and ns indicates not significant. Abbreviations: HTM, human trabecular meshwork; iPSCs, induced pluripotent stem cells; TM, trabecular meshwork.
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fig04: Differentiated TM-like iPSCs can perform phagocytosis, a typical TM cell property. (A): iPSCs, HTM, and differentiated TM-like iPSCs cultured on chamber slides were incubated with fluorescent-labeled zymosan particles for an hour. They were then washed, fixed, and immunostained with LAMP1, a lysosomal marker, to verify internalization. The colocalization (yellow; examples marked with white arrowheads) of zymosan particles (green) with LAMP1 (red) indicates the phagocytosis of the particles. Both HTM and TM-like iPSCs phagocytosed the particles but iPSCs did not. Nuclei were demarcated by DAPI staining (blue). Scale bar represents 100 µm. (B): The number of zymosan particles that colocalize with LAMP1 was counted and the total number was divided by total number of cells in the field. Significance is indicated by * where p < .05 and ns indicates not significant. Abbreviations: HTM, human trabecular meshwork; iPSCs, induced pluripotent stem cells; TM, trabecular meshwork.

Mentions: The outer portion of the TM acts as a filter to remove cells and debris from aqueous humor and is therefore very active in phagocytosis 42,43. However, iPSCs are not effective at phagocytosis. Hence, we examined whether or not the differentiated TM-like iPSCs had acquired phagocytic capability. We exposed iPS, TM, and differentiated TM-like iPSCs to fluorescent-labeled zymosan particles (red) and immunostained afterward for the lysosomal marker LAMP1 (green), to verify that particles were actually internalized and not just absorbed to the cell surface. The results showed that many of the zymosan particles colocalized with LAMP1, indicating internalization, for both human TM and TM-like iPSCs, but not for iPSCs, which exhibit essentially no lysosomes or zymosan uptake (Fig. 4).


Induced pluripotent stem cells restore function in a human cell loss model of open-angle glaucoma.

Abu-Hassan DW, Li X, Ryan EI, Acott TS, Kelley MJ - Stem Cells (2015)

Differentiated TM-like iPSCs can perform phagocytosis, a typical TM cell property. (A): iPSCs, HTM, and differentiated TM-like iPSCs cultured on chamber slides were incubated with fluorescent-labeled zymosan particles for an hour. They were then washed, fixed, and immunostained with LAMP1, a lysosomal marker, to verify internalization. The colocalization (yellow; examples marked with white arrowheads) of zymosan particles (green) with LAMP1 (red) indicates the phagocytosis of the particles. Both HTM and TM-like iPSCs phagocytosed the particles but iPSCs did not. Nuclei were demarcated by DAPI staining (blue). Scale bar represents 100 µm. (B): The number of zymosan particles that colocalize with LAMP1 was counted and the total number was divided by total number of cells in the field. Significance is indicated by * where p < .05 and ns indicates not significant. Abbreviations: HTM, human trabecular meshwork; iPSCs, induced pluripotent stem cells; TM, trabecular meshwork.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4359625&req=5

fig04: Differentiated TM-like iPSCs can perform phagocytosis, a typical TM cell property. (A): iPSCs, HTM, and differentiated TM-like iPSCs cultured on chamber slides were incubated with fluorescent-labeled zymosan particles for an hour. They were then washed, fixed, and immunostained with LAMP1, a lysosomal marker, to verify internalization. The colocalization (yellow; examples marked with white arrowheads) of zymosan particles (green) with LAMP1 (red) indicates the phagocytosis of the particles. Both HTM and TM-like iPSCs phagocytosed the particles but iPSCs did not. Nuclei were demarcated by DAPI staining (blue). Scale bar represents 100 µm. (B): The number of zymosan particles that colocalize with LAMP1 was counted and the total number was divided by total number of cells in the field. Significance is indicated by * where p < .05 and ns indicates not significant. Abbreviations: HTM, human trabecular meshwork; iPSCs, induced pluripotent stem cells; TM, trabecular meshwork.
Mentions: The outer portion of the TM acts as a filter to remove cells and debris from aqueous humor and is therefore very active in phagocytosis 42,43. However, iPSCs are not effective at phagocytosis. Hence, we examined whether or not the differentiated TM-like iPSCs had acquired phagocytic capability. We exposed iPS, TM, and differentiated TM-like iPSCs to fluorescent-labeled zymosan particles (red) and immunostained afterward for the lysosomal marker LAMP1 (green), to verify that particles were actually internalized and not just absorbed to the cell surface. The results showed that many of the zymosan particles colocalized with LAMP1, indicating internalization, for both human TM and TM-like iPSCs, but not for iPSCs, which exhibit essentially no lysosomes or zymosan uptake (Fig. 4).

Bottom Line: A notable feature associated with glaucoma is outflow pathway cell loss.We then differentiated induced pluripotent stem cells (iPSCs) and used them to repopulate this cell depletion model.These differentiated cells (TM-like iPSCs) became similar to TM cells in both morphology and expression patterns.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology, Casey Eye Institute, Oregon Health & Science University, Portland, Oregon, USA; Department of Biochemistry & Molecular Biology, Oregon Health & Science University, Portland, Oregon, USA; Department of Biochemistry & Physiology, University of Jordan, Amman, Jordan.

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Related in: MedlinePlus