Roles of MMP/TIMP in regulating matrix swelling and cell migration during chick corneal development

Abstract

Tissue remodeling is central to embryonic development. Here, we used immunohistochemistry, Western blotting, and RT-PCR analysis to investigate the roles of matrix metalloproteinases (MMPs) and the related "a disintegrin and metalloproteinase" (ADAM) family proteinases in chick corneal development. While MMP-13 was expressed in developing chick corneas from embryonic day (ED) 5 to ED 10, its inhibitor, tissue inhibitors of metalloproteinase-1 (TIMP-1), was expressed from ED 18 to 2 days post-hatching (P2). Early MMP-13 activity may be associated with degradation of type IX collagen from the primary stroma, which loosens the collagen fibrils and facilitates neural crest (NC) cell migration. The membrane-bound and secreted forms of ADAM10 were both detected throughout corneal development, and active ADAM10 formed a cleavage complex with CD44v6, a CD44 splice variant that is a major cell surface adhesion molecule for hyaluronic acid (HA) and has been implicated in cell migration. Both CD44v6 and its ectodomain cleavage products were detected from ED 5 to ED 14, and a broad-spectrum MMP inhibitor blocked ectodomain cleavage in cultured stromal cells. These findings suggest that ADAM10 mediates CD44v6 cleavage in the developing cornea, facilitating NC cell-derived mesenchymal cell migration. Finally, we identified high levels of active membrane-type 3-MMP (MT3-MMP) in developing corneas at ED 7, ED 14, and ED 18. MT3-MMP takes part in MMP-2 activation and possibly also CD44v6 shedding, suggesting that this pathway may be involved in cell migration. These findings collectively show for the first time that multiple MMPs, ADAMs, and TIMPs appear to functionally interact during corneal development.