TGF-β induces sustained upregulation of SNAI1 and SNAI2 through Smad and non-Smad pathways, EMT-like morphologic changes, downregulation of E-cadherin, and upregulation of N-cadherin in HCECs. The authors' findings provide insight into the TGF-β signaling and the temporal expression patterns of EMT-inducible transcription factors in HCECs.

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It was found that Slug is upregulated during corneal wound healing in vivo. The overexpression of Slug mediated a change in the cellular phenotype affecting proliferation, migration, and expression levels of differentiation-related molecules. This is the first evidence that Slug is regulated during the process of corneal wound healing in the corneal epithelium in vivo, providing a novel insight into the EMT and Slug expression in corneal wound healing.

Citation: Sugioka K, KodamaTakahashi A, Yoshida K, et al. Extracellular collagen promotes interleukin1b-induced urokinase-type plasminogen activator production by human corneal fibroblasts. Invest Ophthalmol Vis Sci. 2017;58:148758: -149858: . DOI: 10.1167 PURPOSE. Keratocytes maintain homeostasis of the corneal stroma through synthesis, secretion, and degradation of collagen fibrils of the extracellular matrix. Given that these cells are essentially embedded in a collagen matrix, keratocyte-collagen interactions may play a key role in regulation of the expression or activation of enzymes responsible for matrix degradation including urokinase-type plasminogen activator (uPA), plasmin, and matrix metalloproteinases (MMPs). We examined the effect of extracellular collagen on the production of uPA by corneal fibroblasts (activated keratocytes) stimulated with the proinflammatory cytokine interleukin-1b (IL-1b).METHODS. Human corneal fibroblasts were cultured either on plastic or in a three-dimensional gel of type I collagen. Plasminogen activators were detected by fibrin zymography, whereas the IL-1 receptor (IL-1R) and MMPs were detected by immunoblot analysis. Collagen degradation by corneal fibroblasts was assessed by measurement of hydroxyproline in acid hydrolysates of culture supernatants.RESULTS. Collagen and IL-1b synergistically increased the synthesis and secretion of uPA in corneal fibroblasts. Collagen also upregulated IL-1R expression in the cells in a concentrationdependent manner. The conversion of extracellular plasminogen to plasmin, as well as the plasminogen-dependent activation of MMP-1 and MMP-3 and degradation of collagen apparent in three-dimensional cultures of corneal fibroblasts exposed to IL-1b, were all abolished by a selective uPA inhibitor.CONCLUSIONS. Collagen and IL-1b cooperate to upregulate uPA production by corneal fibroblasts. Furthermore, IL-1b-induced collagen degradation by these cells appears to be strictly dependent on uPA expression and mediated by a uPA-plasmin-MMP pathway.

PURPOSE. Staphylococcus aureus is a common cause of corneal ulceration, and staphylokinase (SAK) produced by this bacterium is a plasminogen activator. To investigate the pathogenesis of corneal ulceration induced by S. aureus, we examined the effects of bacterial culture broth and SAK on collagen degradation in a culture model in which human corneal fibroblasts are embedded in a collagen gel. METHODS. Corneal fibroblasts embedded in collagen were exposed to S. aureus culture broth or SAK. Collagen degradation was assessed by measurement of hydroxyproline in acid hydrolysates of culture supernatants. Expression of pro-matrix metalloproteinase-1 (pro-MMP-1) was detected by immunoblot analysis as well as reverse transcription and real-time polymerase chain reaction analysis. RESULTS. Both S. aureus culture broth and SAK markedly increased collagen degradation in the presence of corneal fibroblasts and plasminogen. This effect of the culture broth was dependent on cell number to a greater extent than was that of SAK. Whereas the culture broth also increased the expression of pro-MMP-1 in corneal fibroblasts at both mRNA and protein levels, SAK did not. CONCLUSIONS. Our results suggest that S. aureus may promote collagen degradation both by upregulating pro-MMP1 expression in corneal fibroblasts, with pro-MMP-1 then being converted to active MMP-1 by plasmin, and by directing plasmin activity toward collagen in a SAK-dependent manner.