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George Inana, Christopher Murat, Xiang Yao, Ian R Harris, Jing Cao; Human umbilical tissue-derived cells rescue phagocytosis in cultured retinal pigment epithelial cells from Royal College of Surgeons rat through secretion of receptor tyrosine kinase ligands. Invest. Ophthalmol. Vis. Sci. 2014;55(13):1722.
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It is well established that retinal pigment epithelial (RPE) cells of Royal College of Surgeons (RCS) rat exhibit impaired rod outer segment (ROS) phagocytosis due to mutated Mertk gene. Lund et al. have demonstrated that subretinal injection of human umbilical tissue derived cells (hUTC) into the RCS rat eye improved visual acuity and ameliorated retinal degeneration. However, the mechanism of how hUTC improves vision in the RCS rat is unclear. Mertk is a member of the receptor tyrosine kinase (RTK) family and plays a crucial role in RPE phagocytosis. Basic fibroblast growth factor (bFGF), a ligand of the FGF receptor, a member of RTK, was shown to restore phagocytic function in cultured RPE cells from RCS rats. This leads to our hypothesis that other RTK ligands may compensate for the role of Mertk in RCS RPE cell, and hUTC may restore the phagocytic function of RCS RPE cell through release of RTK ligands. To test this hypothesis, we investigated the secretion of RTK ligands by hUTC and determined their effects on phagocytosis.
The effect of hUTC on RCS RPE phagocytosis was examined in cells pre-incubated with hUTC conditioned medium (CM) for 24 h and subjected to phagocytosis. RNA-Seq based transcriptome profiling was performed to identify gene expression in hUTC and RCS RPE cells. Levels of expressed RTK ligands, including brain-derived neurotrophic factor (BDNF), hepatocyte growth factor (HGF), and platelet-derived growth factor type D (PDGF-DD) were measured in hUTC CM by ELISA, and their effects on RCS RPE phagocytosis were examined by pre-incubating cells with the corresponding recombinant human proteins followed by phagocytosis assay in the presence of the respective ligand protein.
hUTC CM significantly rescued phagocytosis in RCS RPE cells, comparable to that of normal RPE cells. Transcriptome profile analysis demonstrated that RCS RPE cells express multiple RTK genes, while hUTC expresses genes for multiple RTK ligands. Selected RTK ligands, BDNF, HGF and PDGF-DD, could be detected in hUTC CM. Recombinant human BDNF, HGF, and PDGF-DD had a statistically significant rescue effect on phagocytosis in the RCS RPE cells.
These results suggest that hUTC could rescue RCS RPE cell phagocytosis in a Mertk-independent manner, possibly through secretion of RTK ligands.
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