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Membrane-type matrix metalloproteinase-1 and -3 activity in primate smooth muscle cells
K I Shofuda1, D Hasenstab, R D Kenagy
1Division of Vascular Surgery, Department of Surgery, University of Washington School of Medicine, 1959 NE Pacific St., Seattle, WA 98195-6410, USA.
Abstract:
Membrane-type matrix metalloproteinases-1 and -3 (MT1- and MT3-MMPs) are expressed by activated smooth muscle cells (SMCs) both in vitro and in vivo (19). To define their functions in SMCs, we transduced MT1- and MT3-MMP cDNAs into baboon SMCs by using adenoviral vectors. Overexpression of MT1-MMP increased the conversion of proMMP-2 to the intermediate and active forms. In contrast, in MT3-MMP-overexpressing cells, MMP-2 was activated partially. Immunoblot analyses revealed that MT1-MMP protein was present in the SMCs and accumulated in the presence of the synthetic MMP inhibitor, BB94, or tissue inhibitor of metalloproteinase-2 (TIMP-2). However, MT3-MMP protein was detectable only when BB94, but not TIMP-2, was present. Zymographic analyses showed that MT3-MMP had much stronger casein- and gelatin-degrading activities than did MT1-MMP. Furthermore, when MT3-MMP and MT1-MMP were coexpressed, MT1-MMP degradation was enhanced; this result supports the possibility that MT3-MMP can degrade MT1-MMP. SMCs overexpressing either MT1- or MT3-MMP exhibited altered morphology, without changing their proliferation. This alteration was prevented by BB94 addition. The cells, which underwent this change, showed reduced adhesion to both collagen and fibronectin and increased migration in a Boyden chamber. The present study demonstrates that MT1- and MT3-MMPs have different enzymatic activities but may nevertheless affect SMC function in the same way.
Insights
Membrane-type matrix metalloproteinases-1 and -3 (MT1- and MT3-MMPs) affect smooth muscle cell (SMC) function differently but similarly. Overexpression alters SMC morphology, adhesion, and migration, impacting vascular biology.
Area of Science:
- Biochemistry
- Cell Biology
- Vascular Biology
Background:
- Activated smooth muscle cells (SMCs) express membrane-type matrix metalloproteinases-1 and -3 (MT1- and MT3-MMPs).
- Understanding the specific functions of MT1- and MT3-MMPs in SMCs is crucial for vascular research.
Purpose of the Study:
- To investigate the distinct and overlapping functions of MT1- and MT3-MMPs in smooth muscle cells.
- To elucidate the enzymatic activities and cellular effects of MT1- and MT3-MMP overexpression.
Main Methods:
- Adenoviral vectors were used to overexpress MT1- and MT3-MMP cDNAs in baboon SMCs.
- Immunoblot, zymographic analyses, and cell-based assays (morphology, proliferation, adhesion, migration) were employed.
- The effects of MMP inhibitors (BB94) and TIMP-2 were assessed.
Main Results:
- MT1-MMP overexpression enhanced MMP-2 activation, while MT3-MMP showed partial activation.
- MT3-MMP exhibited stronger casein and gelatin degradation activities than MT1-MMP.
- Coexpression suggested MT3-MMP can degrade MT1-MMP; both MMPs altered SMC morphology, reduced adhesion, and increased migration, effects blocked by BB94.
Conclusions:
- MT1- and MT3-MMPs possess distinct enzymatic properties but converge to influence SMC behavior.
- These MMPs play significant roles in modulating SMC adhesion, migration, and morphology, relevant to vascular remodeling.