Glycoproteins are potentially important biomarkers of disease and therapeutic targets. In particular, the N-linked glycoproteins are a focus of interest as they can be found in the extracellular environment and body fluids. In this study, we have sampled the tears, the extracellular fluid of the epithelial cells covering the surface of the eye, of patients with climatic droplet keratopathy (CDK) using tears of unaffected normal patients for comparison. Prefractionation of the tear sample used a hydrazide-resin capture method, and the previously N-glycosylated peptides were then subjected to two-dimensional nano-LC-nano-ESI-MS/MS analysis to obtain peptide fragmentation patterns for identification through protein database searches. We have identified a total of 43 unique N-glycoproteins, 19 of which have not previously been reported in tear fluid. In addition, we have quantitatively compared N-glycoprotein profiles in tear fluid of patients with CDK to tears of nondiseased controls using glycopeptide capture, iTRAQ labeling and 2D nano-LC-nano-ESI-MS/MS analysis. In tears of CDK patients, increased levels of four N-glycosylated proteins including haptoglobin (at sites N207, N211 and N241), polymeric immunoglobulin receptor (at sites N83, N90, N135, N186, N421, and N469), immunoglobulin J chain (at site N49) and an uncharacterized protein DKFZp686M08189 (at site N470), as well as a decrease in the N-glycosylation level of one N-glycosylated protein, lacritin (at site N119) were observed. However, the overall levels of these five proteins showed no appreciable changes between control and CDK samples. The findings could be clinically significant in terms of disease etiology and biomarkers.

BackgroundConjunctival amyloidosis is a very rare condition, generally unilateral, and presents mostly as an isolated condition without systemic compromise. Our purpose is to present a new case of systemic amyloidosis with a bilateral conjunctival involvement.Case PresentationA 66-years-old caucasian female complaining of conjunctival hemorrhage and chemosis in both eyes for the last five years had been discontinuously treated with topical antibiotics and corticosteroids without any evident improvement. She presented with a pink-yellow infiltration in the inferior conjunctiva of both eyes. Conjunctival biopsy under optical microscopy revealed amyloid deposit, confirmed by Congo red staining. Mucosal biopsy from esophagus and rectus confirmed amyloidosis by Congo red stain. Immunohistochemistry of bone marrow biopsy showed an increased number of plasma cells and an over-expression of light chain kappa subunit. She was treated with corticosteroids and lubrication with an improvement of symptoms. Ocular lesions remained stable after a follow-up of 3 years.ConclusionsConjunctival amyloidosis is a rare entity that may be overlooked, and should be differentiated from chronic conjunctivitis and conjunctival malignancies. Although it presents most frequently as a local process, a systemic involvement should always be ruled out.

This study provides a detailed in vivo and postfixed histological description of the Guinea pig's cornea and information about the physiological tests.

ABSTRACT.Climatic droplet keratopathy (CDK) is an acquired and potentially handicapping cornea degenerative disease that is highly prevalent in certain rural communities around the world. It predominantly affects males over their forties. It has many other names such as Bietti's band-shaped nodular dystrophy, Labrador keratopathy, spheroidal degeneration, chronic actinic keratopathy, oil droplet degeneration, elastoid degeneration and keratinoid corneal degeneration. CDK is characterized by the haziness and opalescence of the cornea's most anterior layers which go through three stages with increasing severity. Globular deposits of different sizes may be histopathologically observed under the corneal epithelium by means of light and electron microscopy. The coalescence and increased volume of these spherules may cause the disruption of Bowman's membrane and the elevation and thinning of the corneal epithelium. The exact aetiology and pathogenesis of CDK are unknown, but they are possibly multifactorial. The only treatment in CDK advanced cases is a corneal transplantation, which in different impoverished regions of the world is not an available option. Many years ago, the clinical and histological aspects of this disease were described in several articles. This review highlights new scientific evidence of the expanding knowledge on CDK's pathogenesis which will open the prospect for new therapeutic interventions.

ABSTRACT.Purpose: Climatic droplet keratopathy (CDK) is an acquired corneal disease characterized by progressive scarring of the cornea. In several corneal diseases, matrix metalloproteinases (MMPs) are upregulated during the degradation of epithelial and stromal tissues. We investigated the levels, degree of activation and molecular forms of MMP-2, MMP-9, MMP-8 and MMP-13 and their tissue inhibitors TIMP-1 and TIMP-2 in tear fluid of patients with CDK. Methods: Seventeen CDK patients and 10 controls living in Argentine Patagonia received a complete eye examination, and MMPs and TIMP-1 ⁄ 2 were determined by immunofluorometric assay (IFMA), gelatin zymography and quantitative Western immunoblot analysis in tear samples. Results: The MMPs were detected mostly in their latent forms. The levels of MMP-9 and MMP-2 were found to be significantly elevated in CDK patients, whereas latent and active MMP-8 levels were significantly enhanced in controls. There was no significant difference in the level of MMP-13. TIMPs were found as part of complexes, and the TIMP-1 levels were significantly lower in patients than controls. Conclusion: Elevated MMP-2 and MMP-9 levels have been implicated in the failure of corneal re-epithelialization, and enhanced MMP-2 and MMP-9 levels in CDK patients suggest that these MMPs may play a role in corneal scarring in CDK. Elevated levels of MMP-8 suggest a defensive role for this MMP in inflammatory reactions associated with recurring corneal traumas. Decreased expression of TIMP-1 in CDK patients suggest deficient antiproteolytic shield likely to render the corneas of CDK patients vulnerable to enhanced MMPs. Overall, these data suggest a mechanistic link between MMPs and TIMP-1 level in cornea and tears with corneal scarring in CDK.

A severe decrease of corneal sensitivity was observed in advanced stages of CDK. Some degree of dry eye was present in most patients, but severe disease was infrequent at any stage of CDK. Inferior iris depigmentation and atrophy was frequently observed among patients with CDK.

Approximately 105 proteins were identified in CDK specimens, and a subset of them was confirmed by immunohistochemistry. Several of these may play a role in fibril or deposit formation.

The authors demonstrate that the corneal epithelium could participate in CDK development as a source of cytokines and gelatinases. Additionally, in HCE cells, UV-B- modulated cytokine and subsequent MMP secretion. Local inhibition of cytokine secretion and gelatinases may prevent CDK progression.