Colocalization of increased transforming growth factor-β-induced protein (TGFBIp) and clusterin in Fuchs endothelial corneal dystrophy.

Fuchs endothelial corneal dystrophy (FECD) is the most common endogenous corneal endotheliopathy, leading to the progressive corneal edema, blindness and eventual need for corneal transplantation to restore vision. In early stages FECD is characterized by extracellular collagenous deposits that accumulate posterior to Descemet’s membrane, thicken it and cause the formation of mound-shaped aggregates, called “guttae.” The characteristic pattern on specular microscopy showing dark areas in the endothelial mosaic is due to the displacement of the endothelium posterior to the plane of focus by these excrescences. In the later stages, the coalescence of the guttae occurs, and a progressive disruption of the endothelial mosaic causes cell thinning, stretching, enlargement, and loss of hexagonal shape.1 Eventually, there is a compromise in the intact covering of the endothelial monolayer as endothelial cell loss progresses in an inversely proportional manner to guttae formation. The considerable strain on corneal endothelial cells situated at the apices of these excrescences causes the morphologic changes that lead to the loss of barrier function as well as cell apoptosis which has been widely implicated in FECD.2 Still, the exact composition of these outgrowths or guttae is currently not clear. Studies from this laboratory have described the results of 2-D gel analysis of proteins extracted from human corneal endothelial cell-Descement’s membrane (HCEC-DM) complexes dissected from the corneas of normal donors and FECD patients.3, 4 These studies showed that there is a marked overexpression of the pro-aggregative, chaperone-like protein, clusterin, in FECD.3 The staining of FECD corneas with CLU antibodies revealed a clustering of endothelial cells around guttae as well as CLU staining in the centers of guttae, suggesting the presence of cell remnants in these areas. Studies have shown that under stressed conditions CLU tends to cause cell aggregation and induce the formation of junctional contacts between cells.5 Results of the 2-D gel analysis also demonstrated a marked overexpression of a cell adhesion molecule, TGFBIp, in the FECD affected HCEC-DM complexes. TGFBIp was detected as a series of spots, migrating at ~38 kDa. The number and intensity of these spots was greater in FECD HCEC-DM as opposed to normal controls. The purpose of the current study is to further investigate the role of TGFBIp in the characteristic guttae formation and its relation to CLU. TGFBI (transforming growth factor-β [TGF- β]-induced) gene encodes the transforming growth factor beta induced protein (TGFBIp), an extracellular matrix protein that mediates cell adhesion by interacting with collagens, fibronectin, and integrins, mainly α3β1.6–10 This protein has been identified by TGFBI gene induction with TGF-β in a human adenocarcinoma line.11 Mutations in the TGFBI gene are responsible for several corneal stromal dystrophies, such as Lattice dystrophy type I, Reis-Bucklers dystrophy, Thiel-Behnke corneal dystrophy, granular corneal dystrophy II (Avellino corneal dystrophy), and granular corneal dystrophy type I.12–15 Depending on the dystrophy, the mutation in the TGFBI gene manifest as stromal amyloid or/and nonamyloid deposits in which TGFBI itself has been co-localized.16 Analysis of the pathological corneas taken from the stromal dystrophy patients has revealed that there are mutation-specific changes in the processing of the full-size protein as opposed to unaffected corneas.16 In the current study, we employed immunocytochemistry to localize the TGFBIp deposition and to elucidate further the role of TGFBIp in guttae formation. Also, using antibodies against two non-overlapping parts of the protein we investigated whether there are changes in the processing of TGFBIp between FECD and normal endothelial samples. To obtain greater insight into the role of TGFBIp in the HCEC-DM complex we also compared the TGFBIp expression patterns in the corneas of young and old normal donors.