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Soo Hyun Park, Yeoun Sook Chun, Jae Chan Kim; Human Mesenchymal Stem Cells can Differentiate into Keratocyte-like Cells by Amniotic Membrane Matrix and Conditioned Medium. Invest. Ophthalmol. Vis. Sci. 2011;52(14):2531.
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To determine if mesenchymal stem cells (MSCs) can be differentiated to keratocytes by exposure to keratocyte conditioned medium (KCM).
Isolated human keratocytes were seeded on the stromal side of amniotic membrane (AM) or plastic dish, and morphologic changes were evaluated. Human mesenchymal stem cells (hMSCs) were cultured on AM or plastic with minimum essential medium alpha (α-MEM) containing KCM. Reverse transcription polymerase chain reaction (RT-PCR) was used to confirm the gene expression patterns of specific keratocyte markers, including keratocan, lumican, and aldehyde dehydrogenase 1 family, member A1 (ALDH1A1), as well as alpha-smooth muscle actin (α-SMA), a mesenchymal marker in keratocytes and hMSCs. Western blot analysis was used to determine the expression levels of these marker proteins in cultured cells. Immunocytochemical staining was used to observe the expression patterns and cellular localization of the marker proteins.
Keratocytes on AM showed dendritic morphology with slow proliferation and constant expression of keratocan, lumican, and ALDH1A1. RT-PCR analysis showed that keratocytes expressed keratocan, lumican, and ALDH1A1 but not α-SMA. Culturing of hMSCs with KCM induced expression of keratocan and ALDH1A1 transcripts. Western blot analysis showed that hMSCs steadily expressed lumican and ALDH1A1 but ceased to express α-SMA when exposed to KCM. Immunocytochemistry results were consistent with western blotting findings.
Mesenchymal stem cells acquired characteristics similar to those of keratocytes when incubated with KCM. These results suggest that KCM can effect a change in the characteristics, leading to differentiation of hMSCs into keratocytes.
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