Heat Shock Protein 90 Involvement in the Development of Idiopathic Epiretinal Membranes

Abstract

Purpose This work was aimed to further characterize cells of idiopathic epiretinal membranes (iERMs). We wanted to determine the contribution of 90-kDa heat shock protein (HSP90) to sustain the transforming growth factor-β (TGF-β)-mediated signal transduction pathway in iERM. Methods Immunofluorescence and confocal microscopy were carried out on deplasticized sections from 36 epiretinal membranes processed for electron microscopy and on frozen sections from five additional samples with antibodies against α-smooth muscle actin (αSMA), vimentin, glial fibrillary acidic protein (GFAP), SMAD2, HSP90α, type-II TGF-β1 receptor (TβRII), type-I collagen, and type-IV collagen. In addition, Müller MIO-M1 cells were transfected with HSP90 and challenged with TGF-β1. Results Double and triple labeling experiments showed that a variable number of TβRII + cells were present in 94.1% of tested iERMs and they were mostly GFAP − /αSMA + /vimentin + /HSP90α + . In almost half of the cases these cells contained type-I collagen, suggesting their involvement in matrix deposition. HSP90 overexpressing MIO-M1 cells challenged with TGF-β1 showed increased levels of TβRII, SMAD2, SMAD3, and phosphor-SMAD2. Nuclear SMAD2 staining could be observed in HSP90α + cells on frozen sections of iERMs. Conclusions Cells in iERMs that express TβRII are also HSP90α + and show the antigenic profile of myofibroblast-like cells as they are GFAP − /αSMA + /vimentin + . HSP90α-overexpressing MIO-M1 cells challenged with TGF-β1 showed an increased activation of the SMAD pathway implying that HSP90α might play a role in sustaining the TGF-β1-induced fibrotic response of iERM cells.

The extracellular matrix complexity of idiopathic epiretinal membranes and the bilaminar arrangement of the associated internal limiting membrane in the posterior retina

“…Idiopathic ERMs express collagens I, III, IV, and VI; fibronectin; and laminin as previously reported [17,22,23]. As far as collagen IV is concerned, the isoform detected in iERMs should be the [α1(IV)] 2 α2(IV).…”

“…However, the variations occurring in the relationships between iERMs and the ILM received little attention [ 20 ]. The unusual technique that we employ, immunofluorescence and confocal microscopy on semithin epon-embedded sections [ 17 , 23 ], produces high-quality images that allow the study of details that probably have gone unnoticed so far. In this way, we have observed that the relationships between iERMs and their associated ILMs vary greatly: from very thin iERMs, just covering the ILM, to iERMs that, slowly gliding over the ILM, induce the latter to form large loops [ 20 ].…”

“…ERMs are fibrocellular sheets of tissue that, when located in front of the macula and retract, generate a condition known as macular pucker which can strongly affect vision [9]. The cellular component of ERMs has been investigated in many occasions [10][11][12][13][14][15][16][17]. Though several studies have analysed a battery of molecular markers, the ultimate source of cells is still a matter of debate since results often appear inconclusive and conflicting.…”

“…Though several studies have analysed a battery of molecular markers, the ultimate source of cells is still a matter of debate since results often appear inconclusive and conflicting. Whichever the cell type of origin is, cells located in ERMs take on a myofibroblastic phenotype testified by α-smooth muscle actin (α-SMA) expression [10][11][12][13][14][15][16][17] and by their ability to deposit new ECM [16,17].…”

Epiretinal membrane: A review

“…TGF‐β is a cytokine with important homeostatic functions in proliferation, differentiation and apoptosis 68 . Activation of TGF‐β1 triggers signal transduction pathways promoting the transcription of TGF‐β target genes needed for fibrogenesis 69 . Interestingly, myofibroblasts themselves can release latent TGF‐β that bind to ECM proteins forming a reservoir of TGF‐β, and extracellular mechanical stress transmitted by integrins induces release of TGF‐β 70 .…”