The matrix metalloproteinase (MMP) 72-Kda type IV collagenase(72T4Cl0, which is also known as gelatinase A, is thought to be involved in both normal connective tissue remodeling and invasive pathologic processes. 72T4Cl is secreted by fibroblast
monolayers as an inactive proenzyme. However, when fibroblasts are cultured in a collagen lattice, a situation thought to better approximate in vivo conditions, they secreted 72T4Cl in in not only pro- but also active (62-Kda) forms, and
contracted
the
lattice. The lattice contraction from collagen feorganization is mediated by α2βl integrin. We were interested in whether contact between the cell and its collagen matrix is the impetus for 72T4Cl activation, and 72T4Cl activation is related to
lattice contraction. We investigated whether processes resulting in 72T4Cl activation occur intracellarly or extracellularly.
Monoclonal antibidy to integrin β1 with/without α2 inhibited the activation of 72T4Cl and blocked lattice contraction, but showed different degree of inhibition between the two. Both cytochalasin D and α-methly-mannoside completely blocked
lattice
contraction. but not proenzyme activation. /active 72t4Cl was identified in cell extract earlier than in conditioned media using zymogram. Western blot and immunoprecipitaion. Not only cell fractions containing organelles but also cytosol
fraction
showed active 72T4cl. Normal fibroblasts seeded in collagen lattice and collagen gel itself did hardly uptake exogenous 72T4Cl to be provided. Above results suggest that processes of 72T4Cl activation are mediated by α2β1 integrin and occur
intracellularly. And are different from those of lattice contaction.
|