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Cleavage of human corneal type VI collagen alpha 3 chain by matrix metalloproteinase-2
E Myint1, D J Brown, A V Ljubimov
1Department of Surgery, Cedars-Sinai Research Institute, Cedars-Sinai Medical Center, University of California at Los Angeles Medical School Affiliate 90048, USA.
Abstract:
The human cornea contains significant amounts of type VI collagen and matrix metalloproteinase-2 (MMP-2), but there has been no established relation between these two components. The objective of this study was to determine whether corneal type VI collagen was susceptible to digestion by MMP-2. Human corneas were frozen and then pulverized in liquid nitrogen. The type VI collagen was isolated by sequential extractions with sodium chloride buffer, guanidinium chloride solution, and guanidinium chloride/dithiothreitol solution. Visualization of type VI collagen alpha 3(VI) chain was made by using Western blots with specific monoclonal antibodies. The extracts were incubated up to 24 h with isolated, activated MMP-2. Within 4 h of incubation, two lower molecular weight bands (approximately 190 and 170 kDa) appeared. These bands increased in intensity with time but were not further digested into smaller fragments. This cleavage activity was inhibited by ethylenediaminetetraacetic acid (EDTA). Because type VI collagen represents approximately 25% of the corneal dry weight, its degradation properties may be important for the integrity of the stroma in scarring episodes and corneal diseases, such as keratoconus.
Insights
Matrix metalloproteinase-2 (MMP-2) can digest type VI collagen, a major component of the human cornea. This collagen degradation may impact corneal integrity during disease and scarring.
Area of Science:
- Biochemistry
- Ophthalmology
- Extracellular Matrix Biology
Background:
- Type VI collagen is abundant in the human cornea.
- Matrix metalloproteinase-2 (MMP-2) is also present in the cornea.
- The relationship between corneal type VI collagen and MMP-2 was previously unestablished.
Purpose of the Study:
- To investigate the susceptibility of corneal type VI collagen to digestion by MMP-2.
- To understand the potential implications of this interaction on corneal structure.
Main Methods:
- Human corneas were processed to isolate type VI collagen.
- Isolated type VI collagen was incubated with activated MMP-2.
- Western blots were used to visualize collagen fragments.
- Ethylenediaminetetraacetic acid (EDTA) was used to assess MMP-2 activity.
Main Results:
- MMP-2 digestion of type VI collagen resulted in the appearance of lower molecular weight bands (approx. 190 and 170 kDa) within 4 hours.
- These fragments increased in intensity over time but were not further degraded.
- EDTA inhibited the observed cleavage activity, confirming MMP-2's role.
Conclusions:
- Corneal type VI collagen is susceptible to degradation by MMP-2.
- This MMP-2-mediated cleavage of type VI collagen may play a role in corneal stromal integrity, particularly in conditions like scarring and diseases such as keratoconus.