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James L. Funderburgh, Richard P. Dannenberg, Mary M. Mann, Martha L. Funderburgh; Roles for Metalloproteinases in Remodeling of Corneal Stroma by Stem Cells. Invest. Ophthalmol. Vis. Sci. 2011;52(14):3946.
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Adult corneal stroma stem cells remodel stromal extracellular matrix (ECM) in vivo, restoring transparency to lumican knockout mice (Stem Cells 27:1635). This study tests the hypothesis that remodeling by stem cells requires action of specific matrix metalloproteinases (MMPs) by stem cells and that these differ from the MMPs active during wound healing.
Human corneal stromal stem cells (CSSC) isolated from donor corneas were cultured on collagen substrata in medium inducing differentiation to keratocytes. MMP expression was detected by quantitative RT-PCR and by digestion of fluorescent collagen substrata in the presence of MMP inhibitors. Keratan sulfate expression was detected using immunoblotting and mRNA expression of keratocyte markers examined by qPCR. Cell migration through collagen-coated transwell filters was examined in presence of MMP inhibitors.
Culture of CSSC on collagen gels or injection into mouse cornea induced expression of mRNA for MMPs 1,2,3,11 and 14. Corneal fibroblasts representing a model of wound healing cells, expressed much higher levels of MMP2 and MMP16 and less MMP11 than the CSSC. Induction of differentiation of CSSC resulted in a rapid degradation of fluorescent collagen substrata. This phenomenon was blocked by MMP inhibitor GM6001 and by antibodies or siRNA blocking MMP14 (MT1-MMP). Expression of keratan sulfate and the mRNA for keratan sulfate biosynthetic enzymes (B3GNT7, CHST6) was reduced by inhibition of MMP action with GM6001 as was the migration of keratocytes in a transwell assay.
To remodel disorganized stroma, restoring transparency, CSSC need to first remove existing collagenous ECM. These results indicate that in response to collagenous environment, a spectrum of MMPs is induced that is distinct from that of wound-healing fibroblasts. Digestion of the ECM is dependent on expression of the cell-associated MMP14. MMP action also appears to be necessary for CSSC to migrate through a collagen matrix and to synthesize the characteristic corneal glycosaminoglycan, keratan sulfate.
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