Matrix metalloproteinase inhibitors suppress transforming growth factor-beta-induced subcapsular cataract formation.

Department of Pathology and Molecular Medicine, McMaster University, HSC 1R10, Hamilton, ON, Canada L8N3Z5.
American Journal Of Pathology (Impact Factor: 4.59). 02/2006; 168(1):69-79. DOI: 10.2353/ajpath.2006.041089
Source: PubMed


The pleotropic morphogen transforming growth factor-beta (TGFbeta) plays an important role in the development of fibrotic pathologies, including anterior subcapsular cataracts (ASCs). ASC formation involves increased proliferation and transition of lens epithelial cells into myofibroblasts, through epithelial-mesenchymal transformation that results in opaque plaques beneath the lens capsule. In this study, we used a previously established TGFbeta-induced rat cataract model to explore the role of matrix metalloproteinases (MMPs) in ASC formation. Treatment of excised rat lenses with TGFbeta resulted in enhanced secretion of MMP-2 and MMP-9. Importantly, co-treatment with two different MMP inhibitors (MMPIs), the broad spectrum inhibitor GM6001 and an MMP-2/9-specific inhibitor, suppressed TGFbeta-induced ASC changes, including the epithelial-mesenchymal transformation of lens epithelial cells. Using an anti-E-cadherin antibody, we revealed that conditioned media from lenses treated with TGFbeta contained a 72-kd E-cadherin fragment, indicative of E-cadherin shedding. This was accompanied by attenuated levels of E-cadherin mRNA. Conditioned media from lenses co-treated with TGFbeta and MMPIs exhibited attenuated levels of the E-cadherin fragment compared with those from TGFbeta-treated lenses. Together, these findings demonstrate that TGFbeta-induced E-cadherin shedding in the lens is mediated by MMPs and that suppression of this phenomenon might explain the mechanism by which MMPIs inhibit ASC plaque formation.

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    • "A number of signaling pathways, including TGFβ, are involved in promoting EMT during both development and disease (Kalluri and Weinberg, 2009; Thiery et al., 2009). The phenotypic changes found in EMT are associated with molecular changes including reduced expression of epithelial proteins such as the adherens junction protein, E-cadherin, and the tight junction protein, zonula occludens 1 (ZO-1), and upregulation in the expression of mesenchymal proteins (Kalluri and Weinberg, 2009; Thiery et al., 2009), and proteins involved in remodeling of the extracellular matrix such as certain matrix metalloproteinases (Dwivedi et al., 2006). While it is known that factors such as E-cadherin and ZO-1 are essential components of cell adhesion and polarity, and their loss promotes EMT, much less is known about the upstream factors that are required to establish and maintain epithelial structure in vertebrates. "
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    ABSTRACT: The integrity and function of epithelial tissues depends on the establishment and maintenance of defining characteristics of epithelial cells, cell-cell adhesion and cell polarity. Disruption of these characteristics can lead to the loss of epithelial identity through a process called epithelial to mesenchymal transition (EMT), which can contribute to pathological conditions such as tissue fibrosis and invasive cancer. In invertebrates, the epithelial polarity gene scrib plays a critical role in establishing and maintaining cell adhesion and polarity. In this study we asked if the mouse homolog, Scrib, is required for establishment and/or maintenance of epithelial identity in vivo. To do so, we conditionally deleted Scrib in the head ectoderm tissue that gives rise to both the ocular lens and the corneal epithelium. Deletion of Scrib in the lens resulted in a change in epithelial cell shape from cuboidal to flattened and elongated. Early in the process, the cell adhesion protein, E-cadherin, and apical polarity protein, ZO-1, were downregulated and the myofibroblast protein, αSMA, was upregulated, suggesting EMT was occurring in the Scrib deficient lenses. Correlating temporally with the upregulation of αSMA, Smad3 and Smad4, TGFβ signaling intermediates, accumulated in the nucleus and Snail, a TGFβ target and transcriptional repressor of the gene encoding E-cadherin, was upregulated. Pax6, a lens epithelial transcription factor required to maintain lens epithelial cell identity also was downregulated. Loss of Scrib in the corneal epithelium also led to molecular changes consistent with EMT, suggesting that the effect of Scrib deficiency was not unique to the lens. Together, these data indicate that mammalian Scrib is required to maintain epithelial identity and that loss of Scrib can culminate in EMT, mediated, at least in part, through TGFβ signaling.
    Developmental Biology 10/2013; 384(1). DOI:10.1016/j.ydbio.2013.09.027 · 3.55 Impact Factor
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    • "Previous studies from our laboratory have shown that MMP-2 and -9 are involved in TGFβ-induced EMT in the lens. For example, we have shown that in whole rat lenses, the MMP-2/9 inhibitor (MMPi) can suppress TGFβ-induced EMT in the lens (as determined by αSMA expression) and subsequent subcapsular cataract formation [38]. To determine the involvement of MMP-2 and -9 in TGFβ-induced MRTF-A translocation, rat lens explant cultures were treated with TGFβ or cotreated with TGFβ, and 25 µM of the MMP-2/9 inhibitor (MMPi) for 48 h and subsequently stained for MRTF-A and αSMA. "
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    ABSTRACT: Transforming growth factor beta (TGFβ) is a known inducer of epithelial to mesenchymal transition (EMT), and studies in other systems have shown that nuclear localization of the myocardin-related transcription factor (MRTF) is downstream of TGFβ. In the following study, we investigated whether nuclear translocation of MRTF-A or MRTF-B is involved in TGFβ-induced EMT of lens epithelial cells (LECs). We further investigated the relationship between matrix metalloproteinase-2 and -9 (MMP-2/9) and MRTF in the EMT of LECs. Rat lens explant cultures were used as the model system. Explants were treated with TGFβ, an MMP-2/9 inhibitor, or actin binding drugs and immunostained for alpha smooth muscle actin (αSMA), MRTF-A, and MRTF-B. Cytoplasmic and nuclear intensities of cells were measured using ImageJ. Production of αSMA was measured using western blot analysis and ImageJ. Untreated explant cells exhibited little αSMA expression, and MRTF-A and B were found to reside primarily in the cytosol. However, when stimulated with TGFβ, a significantly greater number of cells exhibited nuclear expression of MRTF-A, accompanied by an increase in αSMA expression. However, MRTF-B remained in the cytoplasm following TGFβ treatment. Cotreatment with an MMP-2/9 inhibitor and TGFβ resulted in reduced MRTF-A nuclear localization and αSMA expression compared to cells treated with TGFβ alone. Our results are the first to demonstrate the expression of MRTF-A in LECs and that its nuclear translocation can be stimulated by TGFβ. Our data further suggest that MMP-2 and -9 are involved in the translocation of MRTF-A in LECs during TGFβ-induced EMT.
    Molecular vision 05/2013; 19:1017-1028. · 1.99 Impact Factor
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    • "Our laboratory has shown that MMP inhibition can suppress TGFβ-stimulated fibrosis in the lens. For example, using a TGFβ-induced model of anterior subcapsular cataracts in rats, we demonstrated that cotreatment with the MMP-2/9 specific inhibitor effectively prevented anterior subcapsular cataract formation and the associated deposition of matrix [43]. In addition, MMP inhibition has also been shown to prevent matrix deposition in injury-induced tissue remodeling [44,45]. "
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    ABSTRACT: Extracellular matrix remodeling is thought to have profound effects on tissue architecture and associated function. We have shown previously that overexpression of transforming growth factor beta (TGFβ), which stimulates matrix accumulation, results in altered morphology, cataract, and ocular hypertension in rodents. We have further shown that TGFβ-induced cataracts can be mitigated through inhibition of the matrix metalloproteinases (MMP) MMP-2 and MMP-9. We therefore sought to determine whether loss of MMP expression also altered TGFβ-induced changes in intraocular pressure (IOP). To carry out this study, TGFβ1 transgenic mice were bred onto a MMP-9 null background. IOP measurements were made at 1- to 2-, 2- to 3-, and 3- to 4-month time points using a TonoLab rebound tonometer. Histological and immunofluorescence findings were obtained at the same time points. Our results demonstrate that lens-specific expression of TGFβ1 in mice results in altered morphology of the anterior segment and an accompanying significant increase in IOP. TGFβ1 transgenic mice bred onto the MMP-9 null background exhibited a further increase in IOP. Interestingly, the MMP-9-deficient animals (without the TGFβ transgene), which exhibited normal angle morphology, had increased IOP levels compared to their wild-type littermates. These results indicate that TGFβ and MMP-9 likely act independently in regulating IOP. Additionally, MMP-9 plays an important role in maintaining IOP, and further investigation into the mechanisms of MMP-9 activity in the anterior angle may give clues to how extracellular matrix remodeling participates in ocular hypertension and glaucoma.
    Molecular vision 03/2013; 19:684-95. · 1.99 Impact Factor
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