Proteomics characterization of cell membrane blebs in human retinal pigment epithelium cells.
Age-related macular degeneration (AMD) is the leading cause of legal blindness among the elderly population in the industrialized world, affecting about 14 million people in the United States alone. Smoking is a major environmental risk factor for AMD, and hydroquinone is a major component in cigarette smoke. Hydroquinone induces the formation of cell membrane blebs in human retinal pigment epithelium (RPE). Blebs may accumulate and eventually contribute first to sub-RPE deposits and then drusen formation, which is a prominent histopathologic feature in eyes with AMD. As an attempt to better understand the mechanisms involved in early AMD, we sought to investigate the proteomic profile of RPE blebs. Isolated blebs were subjected to SDS-PAGE fractionation, and in-gel trypsin-digested peptides were analyzed by LC-MS/MS that lead to the identification of a total of 314 proteins. Identified proteins were predominantly involved in oxidative phosphorylation, cell junction, focal adhesion, cytoskeleton regulation, and immunogenic processes. Importantly basigin and matrix metalloproteinase-14, key proteins involved in extracellular matrix remodeling, were identified in RPE blebs and shown to be more prevalent in AMD patients. Altogether our findings suggest, for the first time, the potential involvement of RPE blebs in eye disease and shed light on the implication of cell-derived microvesicles in human pathology.
Alcazar
Oscar
O
Bascom Palmer Eye Institute, University of Miami, Miami, Florida 33136, USA.
Hawkridge
Adam M
AM
Collier
Timothy S
TS
Cousins
Scott W
SW
Bhattacharya
Sanjoy K
SK
Muddiman
David C
DC
Marin-Castano
Maria E
ME
D000368:Aged; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D051926:Basigin / Q000032:analysis; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry* / Q000187:drug effects / Q000648:ultrastructure*; D022081:Cell Surface Extensions / Q000737:chemistry* / Q000187:drug effects / Q000648:ultrastructure*; D002853:Chromatography, Liquid; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D004847:Epithelial Cells / Q000737:chemistry / Q000166:cytology* / Q000187:drug effects; D005109:Extracellular Matrix / Q000737:chemistry / Q000378:metabolism; D005136:Eye Proteins / Q000032:analysis*; D006801:Humans; D006873:Hydroquinones / Q000494:pharmacology; D008268:Macular Degeneration / Q000473:pathology; D053511:Matrix Metalloproteinase 14 / Q000032:analysis; D020778:Matrix Metalloproteinase 2 / Q000032:analysis; D008969:Molecular Sequence Data; D040901:Proteomics / Q000379:methods; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D055213:Retinal Pigment Epithelium / Q000737:chemistry / Q000166:cytology*; D012906:Smoke; D012907:Smoking / Q000009:adverse effects; D053719:Tandem Mass Spectrometry; D014026:Nicotiana / Q000737:chemistry
D000975:Antioxidants; C497886:BSG protein, human; D005136:Eye Proteins; D006873:Hydroquinones; D011993:Recombinant Fusion Proteins; D012906:Smoke; D051926:Basigin; D020778:Matrix Metalloproteinase 2; D053511:Matrix Metalloproteinase 14; C031927:hydroquinone