Low Tissue Inhibitor of Metalloproteinases 3 and High Matrix Metalloproteinase 14 Levels Defines a Subpopulation of Highly Invasive Foam-Cell Macrophages

Abstract

Objective-An excess of metalloproteinases (MMPs) over tissue inhibitors of metalloproteinases (TIMPs) may favor atherosclerotic plaque rupture. We compared TIMP levels in nonfoamy and foam-cell macrophages (FCM) generated in vivo. Methods and Results-In vivo generated rabbit FCM exhibited 84% reduced TIMP-3 protein compared to nonfoamy macrophages, and immunocytochemistry revealed a TIMP-3 negative subset (28%). Strikingly, only TIMP-3 negative FCM invaded a synthetic basement membrane, and invasion was inhibited by exogenous TIMP-3. TIMP-3 negative FCM also had increased proliferation and apoptosis rates compared to TIMP-3 positive cells, which were retarded by exogenous TIMP-3; this also reduced gelatinolytic activity. TIMP-3 negative FCM were found at the base of advanced rabbit plaques and in the rupture-prone shoulders of human plaques. To explain the actions of low TIMP-3 we observed a 26-fold increase in MT1-MMP (MMP-14) protein in FCM. Adding an MT1-MMP neutralizing antibody reduced foam-cell invasion, apoptosis, and gelatinolytic activity. Furthermore, MT1-MMP overexpressing and TIMP-3 negative FCM were found at the same locations in atherosclerotic plaques. acrophages have been proposed to be involved in both atherosclerotic plaque fibrous cap formation and disruption. [1][2][3][4] Matrix metalloproteinases (MMPs) are one group of proteases produced by macrophages that also appear to have a dual role in plaque cap building and destruction. Consistent with this, MMPs-2 and -9 in particular are implicated in intima formation after vascular injury. On the other hand, are upregulated in human plaques in macrophage-rich areas that show a high propensity for plaque rupture. 5 Numerous MMPs are also upregulated in the plaques of cholesterol-fed rabbits 6,7 and ApoE null mice. 8 Studies with ApoE/MMP compound knockout mice and other transgenic models show clear effects of individual MMPs on both fibrous cap formation and rupture. 5 What seems to determine the outcome is the level and spectrum of MMPs produced and activated. There is therefore significant interest in the factors regulating MMP activity in macrophages. Conversion of macrophages to foam-cells is one important factor. For example, in vivo generated rabbit foam-cells have increased levels of MMPs-1 to 3 and -12. 6,9,10 Tissue inhibitors of MMPs (TIMPs) are a family of specific protein inhibitors of MMPs, 4 of which have been demonstrated in vascular cells. Smooth muscle cells (SMCs) in human atherosclerotic plaques harbor abundant TIMP-1 and -2 protein expression. 11 TIMP-1 and -2 expression is increased at the base of atherosclerotic plaques in cholesterol-fed rabbit aortas, and this correlates with areas of low MMP activity measured by in situ zymography. 12 TIMP-3 has been detected more selectively in plaque macrophages at the shoulder regions and between the fibrous cap and necrotic core. 13 Subsequently it was suggested that TIMP-3 may serve as a protective mechanism against plaque rupture by dampening local proteolysis. 13 Conclusions-These...

Vulnerable atherosclerotic plaque metalloproteinases and foam cell phenotypes

“…For example, previous work has suggested a selective role for M2 macrophages in aneurysm formation, in part through the socalled matrix metalloelastase, MMP-12 (9). Our data showing that TIMP-3-negative, MMP-14-positive foam cells are more invasive and destructive of matrigel, localise to the deep areas of plaques, and are more likely to undergo apoptosis, suggests a potential role for this subpopulation in lipid core formation (33). These proposals have, however, not yet been tested formally in any experimental model.…”

“…More recently we found that TIMP-3 is down-regulated in FCM compared to NFM (33). Furthermore, a distinct TIMP-3 negative subpopulation was identified, which was highly invasive through matrigel, proliferative and prone to undergo apoptosis.…”

“…Consistent with this, we found that the TIMP-3 negative subpopulation selectively over-expresses MMP-14. Moreover inhibitory antibodies against MMP-14 mimic the effects of adding exogenous TIMP-3 (33). This FCM sub-population may have M1-like properties, since IFN-γ down-regulates TIMP-3 in macrophages (unpublished observations).…”

“…We study foam cells formed in vivo and relate this to MMP regulation and functions including migration, invasion, proliferation and apoptosis measured ex vivo (32)(33)(34). Foam cells are obtained from polyurethane sponges implanted under the dorsal skin of rabbits.…”

Diabetes mellitus and apoptosis: inflammatory cells

“…Cytosolic cytochrome c then promotes caspase activation through apoptosome formation. Activation of these classical pathways has been shown to occur in macrophages with experimental evidence implicating the following: Bim [92], Bax [93], FasL [94], caspases [95], p53 [96], Poly (ADP-ribose) polymerase (PARP)-1 [97], peroxisome proliferator-activated receptor (PPAR)-a/c [98], migration inhibitory factor (MIF) [99], matrix metallo-proteinases (MMPs) [100], Cathepsin L/K [101,102], lipoprotein-associated phospholipase A2 (Lp-PLA2) [103], ox-LDL [104] and oxysterols [105].…”