Profound Re-Organization of Cell Surface Proteome in Equine Retinal Pigment Epithelial Cells in Response to In Vitro Culturing
Autor: | Stefanie M. Hauck, Marius Ueffing, Cornelia A. Deeg, Christoph M. Szober, Kerstin N. Euler, Kristina J. H. Fröhlich, Claudia S. Alge-Priglinger |
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Jazyk: | angličtina |
Rok vydání: | 2012 |
Předmět: |
Proteomics
Proteome Cell Retinal Pigment Epithelium Biology Catalysis Article Inorganic Chemistry lcsh:Chemistry chemistry.chemical_compound retinal pigment epithelium cells medicine Animals membrane protein Horses Physical and Theoretical Chemistry equine recurrent uveitis Molecular Biology lcsh:QH301-705.5 Spectroscopy Cells Cultured Retinal pigment epithelium membrane Organic Chemistry outer blood-retinal barrier Membrane Proteins Retinal Epithelial Cells General Medicine cell line eye diseases Computer Science Applications Cell biology medicine.anatomical_structure Membrane protein RPE65 chemistry lcsh:Biology (General) lcsh:QD1-999 Cell culture sense organs |
Zdroj: | International Journal of Molecular Sciences Int. J. Mol. Sci. 13, 14053-14072 (2012) International Journal of Molecular Sciences, Vol 13, Iss 11, Pp 14053-14072 (2012) Volume 13 Issue 11 Pages 14053-14072 |
ISSN: | 1422-0067 |
Popis: | The purpose of this study was to characterize the cell surface proteome of native compared to cultured equine retinal pigment epithelium (RPE) cells. The RPE plays an essential role in visual function and represents the outer blood-retinal barrier. We are investigating immunopathomechanisms of equine recurrent uveitis, an autoimmune inflammatory disease in horses leading to breakdown of the outer blood-retinal barrier and influx of autoreactive T-cells into affected horses’ vitrei. Cell surface proteins of native and cultured RPE cells from eye-healthy horses were captured by biotinylation, analyzed by high resolution mass spectrometry coupled to liquid chromatography (LC MS/MS), and the most interesting candidates were validated by PCR, immunoblotting and immunocytochemistry. A total of 112 proteins were identified, of which 84% were cell surface membrane proteins. Twenty-three of these proteins were concurrently expressed by both cell states, 28 proteins exclusively by native RPE cells. Among the latter were two RPE markers with highly specialized RPE functions: cellular retinaldehyde-binding protein (CRALBP) and retinal pigment epithelium-specific protein 65kDa (RPE65). Furthermore, 61 proteins were only expressed by cultured RPE cells and absent in native cells. As we believe that initiating events, leading to the breakdown of the outer blood-retinal barrier, take place at the cell surface of RPE cells as a particularly exposed barrier structure, this differential characterization of cell surface proteomes of native and cultured equine RPE cells is a prerequisite for future studies. |
Databáze: | OpenAIRE |
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