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Expression of degradative enzymes and protease inhibitors in corneas with keratoconus
L Zhou1, S Sawaguchi, S S Twining
1Department of Ophthalmology and Visual Sciences, University of Illinois at Chicago College of Medicine, USA.
Purpose:
Keratoconus is characterized by thinning and scarring of the central region of the cornea. Previous research showed that, in corneas obtained from patients with keratoconus, lysosomal enzyme activities are elevated, whereas levels of protease inhibitors such as alpha1-proteinase inhibitor are reduced. This study was undertaken to examine further the expression of a spectrum of proteolytic enzymes and protease inhibitors.
Methods:
Corneal buttons were collected from patients with keratoconus, healthy subjects, and patients with other corneal diseases. Immunohistochemical staining was performed on paraffin sections. Enzymatic assays and western blot analysis were carried out for cathepsins B and G. In addition, an in situ zymography procedure was used to examine the gelatin- and casein-digesting activities in corneas with keratoconus.
Results:
An enhanced staining was found with antibodies to cathepsins B and G. Enzymatic assays and western blotting confirmed that the levels of these two enzymes were elevated in corneas with keratoconus. No alteration was noted with any of the matrix metalloproteinase (MMP) family members and other enzymes and inhibitors examined, although in situ zymography did indicate an increase in net gelatin- and casein-digesting activities in corneas with keratoconus. These activities were mostly abolished by inhibitors for serine and cysteine proteinases, but not by those for MMPs and aspartic proteinases.
Conclusions:
Levels of cathepsins B and G are increased in corneas with keratoconus. These enzymes may contribute to the heightened in situ gelatin- and casein-digesting activities, leading to abnormalities in keratoconus.
Insights
Levels of cathepsins B and G are elevated in keratoconus corneas, contributing to tissue damage. This study investigated proteolytic enzymes and inhibitors in keratoconus, revealing increased activity of specific enzymes.
Area of Science:
- Ophthalmology
- Biochemistry
- Proteomics
Background:
- Keratoconus involves corneal thinning and scarring.
- Elevated lysosomal enzyme activity and reduced protease inhibitors are noted in keratoconus.
- Further investigation into proteolytic enzymes and inhibitors in keratoconus is warranted.
Purpose of the Study:
- To examine the expression of various proteolytic enzymes and protease inhibitors in keratoconus.
- To compare enzyme and inhibitor levels in keratoconus corneas versus healthy and other corneal disease controls.
Main Methods:
- Corneal buttons from keratoconus patients and controls were analyzed.
- Immunohistochemical staining, enzymatic assays, and western blotting were used for cathepsins B and G.
- In situ zymography assessed gelatin- and casein-digesting activities.
Main Results:
- Cathepsins B and G levels were significantly elevated in keratoconus corneas.
- Increased gelatin- and casein-digesting activities were observed in keratoconus corneas.
- Matrix metalloproteinases (MMPs) and other examined enzymes/inhibitors showed no significant alterations.
Conclusions:
- Elevated levels of cathepsins B and G are confirmed in keratoconus.
- These enzymes likely contribute to increased proteolytic activity in keratoconus.
- The findings suggest cathepsins B and G play a role in keratoconus pathogenesis.