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C. Lupien, C. Salesse; Comparison Between the Gene Expression Profile of Human Müller Cells and Two Spontaneous Müller Cell Lines . Invest. Ophthalmol. Vis. Sci. 2005;46(13):3152.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Müller cells are the principal glial cells in the retina. Alterations in Müller cell behavior are observed in retinal tissue from patients with various retinal disorders, including proliferative diabetic retinopathy. The purpose of this study was to compare gene and protein expression profile of normal human Müller cells (NHMC) with two spontaneously human Müller cell lines generated from types I (HMCL–I) and II (HMCL–II) diabetic donors using Serial Analysis Gene Expression (SAGE), RT–PCR. Methods: Recently, Müller glial cells have been postulated to act as progenitor cells of the retina. The expression of a marker of progenitor cells in addition to some growth and trophic factors known to be expressed by progenitor cells has thus been measured by RT–PCR using NHMC and both HMCL. The expression of VEGF, a growth factor known to be overexpressed during diabetic retinopathy, has also been measured. SAGE analysis of these three types of cells was then performed by isolating their mRNA and preparing the SAGE libraries. Sequences were analyzed by the SAGEana program, a modification of SAGEparser. We used the comparative count display (CCD) test to identify the transcripts that were differentially expressed significantly (P 0.05) between the groups with more than a twofold change. Results: All proteins studied (PAX–6, BDNF, CNTF, bFGF, EGF, IGF–1 and TGF–alpha) were shown to be expressed by NHMC and both HMCL. However, the expression of bFGF is stronger in HMCL than in NHMC whereas EGF and TGF–alpha are expressed less strongly in HMCL. The expression of PAX–6 is stronger in HMCL–II than in HMCL–I. Approximately 50 000 tags were sequenced for each of the three SAGE libraries. Tags corresponding to linker sequences were discarded, and duplicate concatemers were counted only once. Identification of the transcripts was obtained by matching the 15 bp (CATG+11 bp tags) with the UniGene and GenBank databases. Classification of the genes was based upon the updated information of the genome directory found at the TIGR website. Conclusions: SAGE allowed us to characterize the entire transcriptome of human Müller cells and to compare these data with those from Müller cells of type I and II diabetic donors.
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