Clusterin in Age-Related Ocular Exfoliation Syndrome

Abstract

 

Purpose:

 

Exfoliation syndrome (XFS) has been recognized as the most common identifiable cause of glaucoma. In addition to the complex mixture of extracellular matrix glycoproteins and proteoglycans heavily cross-linked and assembled in a supramolecular fibrillar structure that compose XFS deposits, our proteomic studies demonstrated that the extracellular protein clusterin was particularly overrepresented. The present study was conducted to assess the topographical distribution of clusterin in XFS deposits and evaluate its concentration levels in the anterior chamber in an effort to provide insight into the disease pathogenesis.

 

Methods:

 

Using anterior lens capsules and aqueous fluid collected at the time of surgery from glaucoma patients with and without XFS we conducted ultrastructural, immunohistochemical and biochemical studies using a combination of high resolution atomic force microscopy, confocal microscopy, capture ELISA and Western blot analysis.

 

Results:

 

XFS fibers were located only in the anterior surface of the lens whereas no fibrils were detected in the epithelial side in the samples tested. Individual fiber width measured an average of 40 nm (range: 30-50 nm) displaying a banding periodicity of 10 nm while the length of many individual fibers extended over several microns. Clusterin co-localized with the XFS fibrils and was absent in the epithelial side of the lenses. In non-XFS glaucoma specimens both sides of the lens were devoid of fibrils as well as of clusterin. Analysis of aqueous humor from 53 XFS glaucoma patients in comparison with 76 non-XFS glaucoma cases revealed over two-fold elevation of the clusterin levels (p<0.0002). No evidence of disrupted blood-aqueous barrier permeability was detected; Western blot analysis indicated the absence of plasma-specific proteins in aqueous fluid.

 

Conclusions:

 

The elevated levels of clusterin in the aqueous fluid of XFS-glaucoma patients together with the absence of specific plasma proteins strongly suggest a local response to cellular stress conditions. The co-localization with XFS fibrils highlights the ability of clusterin to bind to a wide variety of misfolded proteins, including many biochemically diverse amyloid fibrils. Consistent with finding in amyloid disorders, this extracellular chaperone fails to completely inhibit a large-scale misfolding process in spite of protein availability in the anterior chamber.

 

Keywords: aqueous • protein structure/function • microscopy: light/fluorescence/immunohistochemistry