Altered balance between extracellular proteolysis and antiproteolysis is associated with adaptive coronary arteriogenesis.

To study the role of extracellular proteolysis and antiproteolysis during adaptive arteriogenesis (collateral vessel growth) we took 58 collaterals at various developmental stages from 14 dogs with chronic occlusion of the left circumflex coronary artery (LCx) by ameroid constrictor. Immunofluorescence and quantitative immunofluorescence with antibodies against alpha-smooth muscle actin, desmin, matrix metalloproteinases 2 (MMP-2), MMP-9, tissue inhibitor of metalloproteinases 1 (TIMP-1) and 2 (TIMP-2), urokinase-type plasminogen activator (u-PA) and its inhibitor-1 (PAI-1) were studied with confocal microscopy. Additionally, SDS-PAGE zymography was employed. We found that in normal coronary arteries, MMP-2, MMP-9 and PAI-1 were present in all layers of the wall in small amounts. TIMP-1 was found only in smooth muscle cells. In contrast, in growing collaterals, MMP-2 and MMP-9 were 3.4-fold and 4.1-fold higher in the neointima than in the media respectively. TIMP-1 was 4.4-fold higher in the media over the growing neointima. Zymography showed MMP-2 and MMP-9 activated. PAI-1 was increased, especially in the growing neointima where it was 1.4-fold higher. In mature collaterals, MMP-2 and MMP-9 were downregulated in the neointima, 1.4-fold and 1.3-fold higher over the media. TIMP-1 was 1.4-fold increased in the neointima but PAI-1 was downregulated. Desmin and alpha-smooth muscle actin were significantly increased in the neointima compared to growing vessels. U-PA was moderately increased in growing vessels. TIMP-2 was not detectable in collaterals. We conclude that expression of MMP-2 and 9, TIMP-1 and PAI-1 showed a spatial and temporal pattern which is closely associated with the development of collateral vessels. The shift of the balance between proteolysis and antiproteolysis is regulated not only by MMPs and TIMP-1, but also by the PA-PAI system.

Vascular remodeling and altered protein expression during growth of coronary collateral arteries.

The cellular mechanism of growth of coronary collateral vessels (adaptive arteriogenesis) is still poorly understood. To define a possible role of an altered expression pattern of cellular and matrix proteins in this process we implanted a constricting device around the left circumflex artery in 25 canine hearts and sacrificed the animals at the time of initiation (3 weeks), high activity (6 weeks) and discontinuation (8 weeks) of vessel growth. Methods were electron microscopy, labeling with Ki-67, the TUNEL method and immunofluorescence with confocal laser microscopy. As described earlier, the collateral vessels increased in wall thickness by the formation of a neointima without luminal narrowing. We report here for the first time that extensive vascular remodeling including migration, proliferation and apoptosis in all cell types takes place during the growth phase but not in more mature vessels. The most obvious difference with normal vessels is the reiteration of an embryonal expression pattern in smooth muscle cells of the neointima which includes a significant reduction of desmin and alpha-smooth muscle actin, calponin and vinculin. Fibronectin as a promoter of migration and adhesion was abundant, its antagonist tenascin and chondroitin sulfate showed patchy localization. A completely new finding in arteriogenesis is the involvement of mast cells releasing histamine and serotonin and probably cytokines. Vascular protein expression returned to almost normal at 8 weeks indicating cessation of remodeling. We conclude that in collateral vessel development an altered cellular and matrix protein expression is involved in a drastic case of positive vascular remodeling finally resulting in mature vessels 20-fold increased in size which are capable of maintaining the functional and structural integrity of the myocardium at risk.