Matrix Metalloproteinases and their Inhibitors in Aqueous Humor

doi:10.1006/exer.1996.0058

Experimental Eye Research

Volume 62, Issue 5, May 1996, Pages 481-490

https://doi.org/10.1006/exer.1996.0058 Get rights and content

Abstract

Matrix metalloproteinase activity is the rate-limiting step in extracellular matrix degradation. One mechanism by which metalloproteinases are regulated is through the activity of their endogenous inhibitors, the tissue inhibitors of metalloproteinases. Since metalloproteinase activity is a key component of the angiogenic process and many anterior segment structures are largely avascular, we became interested in examining aqueous humor for the presence of metalloproteinases and their endogenous inhibitors. Using zymography, we have identified the presence of several metalloproteinases in normal aqueous humor. Treatment with 4-aminophenylmercuric acetate, an organomercurial which activates latent metalloproteinases, revealed that all metalloproteinases were in their active state. By Western blot analysis, normal aqueous humor was also found to contain at least two tissue inhibitors of metalloproteinases. Subsequent partial purification by two successive chromatographic steps revealed the presence of inhibitory activity against collagenase, endothelial cell DNA synthesis, and angiogenesis on the chick chorioallantoic membrane. The presence of metalloproteinases and their inhibitors in normal aqueous humor, a fluid which bathes avascular ocular structures, suggests that future studies should examine whether an imbalance in this protease/inhibitor family may contribute to the anterior chamber extracellular matrix alterations associated with diseases such as ocular neovascularization and glaucoma.

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Cited by (36)

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Using lens culture, gelatin zymography and immunofluorescence, we were not able to detect any significant differences in MMP-2 or MMP-9 between the WT lens and the transgenic lens epithelial cells overexpressing Spry1 or Spry2 (data not shown). Aqueous and vitreous humors also contain MMPs (Brown et al., 1994; De La Paz et al., 1998; Huang et al., 1996), with MMP-9 increasing dramatically in the humor (Huang et al., 1996; Brown et al., 1994; Kosano et al., 1999) of patients with proliferative diabetic retinopathy, or following cataract surgery. As noted earlier, given that Rac1 GTPase activity has been shown to play an essential role in lens ECM turnover (Maddala et al., 2011) and Spry2 can limit Rac1 activation (Poppleton et al., 2004), this may be the contributing mechanism to the lens capsule rupture.
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Secreted leukocyte protease inhibitor is present in aqueous humours from cataracts and other eye pathologies
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Previous studies identified serine, cysteine and metalloproteases in normal aqueous humours (AH) and suggested that a balance between proteases and their inhibitors may play a role in the modulation of the AH outflow. We aimed to determine whether secretory leukocyte protease inhibitor (SLPI), a serine protease inhibitor, is present in AH of patients with cataract and other eye pathologies. AH was collected from 117 cataract patients of which 55 were diagnosed with more when one eye disease: cataract only (n=62), pseudoexfoliation (PEX) (n=26), glaucoma (n=6), diabetes retinopathy (n=4), iritis–uveitis (n=4) and macular degeneration (n=28). The total protein in AH was determined by a Bradford assay and SLPI was analyzed by Western blot and ELISA methods. The average concentration of total protein and SLPI in AH samples was 160±15 μg/ml (n=117, ±SEM) and 500±94 pg/ml (n=105), respectively. The cataract patients with additional eye disease(s) showed higher protein levels (201±35 μg/ml) than cataract (controls) (128±31 μg/ml), P<0.01. It is noteworthy that no correlation was found between SLPI and the total protein concentrations in AH, but SLPI was positively correlated with age (r=0.2, P<0.05). No statistical difference in SLPI levels was found between controls (cataract) and other pathologies, while patients with iritis/uveitis had higher SLPI levels compared to those with diabetes (P<0.05). We show here for the first time that SLPI is present in AH and may play a role as well as serve as a marker in pathological states.
Matrix metalloproteinase gelatinase B (MMP-9) is associated with leaking glaucoma filtering blebs
2005, Experimental Eye Research
Citation Excerpt :
Although we cannot say whether gelB is the direct cause of bleb leaks, we observed that its presence is associated with a strong gelatinolytic activity that is not found under normal conditions. This is consistent with an earlier study where no gelB was found in normal conjunctiva (Huang et al., 1996). The fact that gelB is found in the uncleaved proenzyme form is not inconsistent with enzymatic activity.
The goal of glaucoma filtering surgery is to create a low resistance pathway for aqueous outflow. The result is a blister or ‘bleb’ on the conjunctiva, from which fluid drains into the vasculature. Filtering surgery results may be compromised if blebs develop leaks, a problem that surfaces more frequently when antimetabolites are used to control the wound healing response. We investigated the role of tissue remodelling enzymes of the Matrix metalloproteinase (MMP) family in the development of bleb leaks. Our design was a case series. We enrolled glaucoma patients with leaking blebs, glaucoma patients with overhanging blebs and normal eyes. Leaking bleb tissues (n=11) and bleb leak fluid were collected from patients undergoing bleb revision surgery. Overhanging bleb tissues (from non-leaking blebs, n=3), normal conjunctiva (n=8), and aqueous humour (n=4) were collected for comparison. Samples were analysed for MMP content and proteinase activity by the methods of zymography, western blotting, immunohistochemistry, and in situ zymography. Our main outcome measures were presence and activity of MMP in sample.
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MMP-g may be involved in the mechanism of formation of bleb leaks. Precise description of the cascade of events leading to bleb leakage may allow the design of therapeutic interventions to prevent, stabilize or reverse bleb leakage.
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The pathogenesis of many anterior segment disorders and ocular complications following surgery are secondary to the wound healing response. The extent of clinical damage observed is closely related to the amount of scarring and tissue contraction. Matrix metalloproteinases (MMPs) are a family of enzymes that play a vital role in all stages of the wound healing process. They degrade all extracellular matrix components and also have the ability to synthesize collagen and extracellular matrix members, and are therefore important in the remodeling of a wound. Overexpression of MMPs results in excessive extracellular matrix degradation, leading to tissue destruction and loss of organ function. In the case of the anterior segment, this may mean the loss of visual function. This review focuses on the role MMPs have in the development of various anterior segment disorders. The importance of MMPs in the wound healing response and its potential modulation to manipulate the scarring response is being recognized, and current developments will be described.
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2000, Experimental Eye Research
Matrix metalloproteinase 2 and 9 (MMP-2 and 9, also known as gelatinase A and B) have been implicated in a number of eye diseases, but their possible involvement in lens pathology is yet to be determined. In the present study, we therefore investigated a possible role of matrix metalloproteinases in cataract and posterior capsule opacification. Whole porcine lenses were removed from the eye and cultured in either Eagles Minimum Essential Medium (EMEM) or EMEM supplemented with 1 m $M$ hydrogen peroxide. The medium was sampled and changed every 2 days. On some occasions a sham cataract operation was performed on cultured lenses. The resulting capsular bag was secured to a Petri dish and cultured in EMEM. Culture media from all preparations were analysed for MMP-2 and 9 activity by gelatin zymography. Media samples from lenses which maintained clarity over the 6 day culture period did not display any detectable gelatinolytic activity. However, media from cataractous lenses demonstrated a gelatinolytic band, which had similar molecular weights to the pro-form of MMP-2. In addition to this band, bands with a similar molecular weight to pro-MMP-9 and its dimeric form were also detected in samples obtained from capsular bag preparations within 24 hr. The data presented indicate that normal lenses have undetectable gelatinase activity. However, there is an associated expression of gelatinases with pathological states of the lens, and therefore gelatinase expression could play an important role in cataractogenesis and posterior capsule opacification.
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^f1: For correspondence at: Massachusetts Eye and Ear Infirmary, 243 Charles Street, Boston, MA02114, U.S.A.

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Regular articleMatrix Metalloproteinases and their Inhibitors in Aqueous Humor

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Matrix Metalloproteinases and their Inhibitors in Aqueous Humor