In situ localization and quantification of seventy-two-kilodalton type IV collagenase in aneurysmal, occlusive, and normal aorta.
OBJECTIVE
Seventy-two-kilodalton type IV collagenase (MMP-2), a potent collagenase and elastase, is present in inflammatory disease states and may be important in the pathogenesis of aortic aneurysms. Alteration in expression of MMP-2 or its inhibitor, the tissue inhibitor of metalloproteinases type two (TIMP-2), could increase extracellular matrix degradation and lead to aneurysm formation. The purpose of this study is (1) to measure relative tissue levels of MMP-2 and TIMP-2 mRNA in aneurysmal, occlusive, and normal human infrarenal aorta; (2) to test for expression by cultured aneurysmal and normal vascular smooth muscle cells (VSMCs); and (3) to identify, in situ, the cells responsible for mRNA production within aneurysmal, occlusive, and normal aortic wall.


METHODS
Total RNA extracted from aneurysmal (n = 8), occlusive (n = 9), and normal (n = 7) tissue was subjected to Northern analysis. Signals for MMP-2 and TIMP-2 were normalized to alpha-tubulin. Mean values +/- SE were compared by use of analysis of variance. Aneurysmal and normal VSMCs were cultured, passaged, and grown to confluence before RNA extraction and Northern analysis. In situ hybridization with digoxigenin RNA probes localized cells responsible for MMP-2 and TIMP-2 mRNA production in histologic sections of aneurysmal (n = 7), occlusive (n = 4), and normal (n = 3) aorta.


RESULTS
Tissue MMP-2 mRNA levels were significantly greater in aneurysmal aorta (1.032 +/- 0.164, n = 5) than in either occlusive (0.553 +/- 0.027, n = 4, p < 0.02) or normal aorta (0.230 +/- 0.038, n = 3, p < 0.002). Differences in TIMP-2 mRNA levels were not significant (aneurysmal aorta 0.207 +/- 0.042, n = 3; occlusive aorta 0.413 +/- 0.164, n = 3; normal aorta 0.260 +/- 0.079, n = 4; p = 0.34), although numbers were small. Cultured aneurysmal and normal VSMCs constitutively expressed both MMP-2 and TIMP-2. In situ studies colocalized tissue MMP-2 and TIMP-2 expression to VSMCs and macrophages surrounding inflammation in aneurysmal adventita, but to atherosclerotic plaque in occlusive aorta.


CONCLUSIONS
MMP-2 and TIMP-2 are expressed in aneurysmal, occlusive, and normal aorta. MMP-2 expression is significantly greater in aneurysmal than in either occlusive or normal aorta. Cultured aneurysmal VSMCs constitutively express both MMP-2 and TIMP-2. Differential patterns of expression seen in situ and elevated tissue MMP-2 mRNA levels in aneurysmal versus occlusive aorta suggest that MMP-2 may be responsible for localized plaque remodeling in occlusive disease and for diffuse adventitial collagen and elastin destruction in aneurysms.
McMillan
W D
WD
Department of Surgery, Northwestern University Medical School, Chicago, IL 60611, USA.
Patterson
B K
BK
Keen
R R
RR
Pearce
W H
WH
D000368:Aged; D001012:Aorta, Abdominal / Q000201:enzymology*; D001014:Aortic Aneurysm / Q000201:enzymology*; D001018:Aortic Diseases / Q000201:enzymology; D001157:Arterial Occlusive Diseases / Q000201:enzymology; D015152:Blotting, Northern; D002478:Cells, Cultured; D005260:Female; D018093:Gelatinases / Q000032:analysis*; D006801:Humans; D017403:In Situ Hybridization; D008297:Male; D020778:Matrix Metalloproteinase 2; D008666:Metalloendopeptidases / Q000032:analysis*; D009131:Muscle, Smooth, Vascular / Q000201:enzymology; D011480:Protease Inhibitors / Q000032:analysis; D011506:Proteins / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis; D019716:Tissue Inhibitor of Metalloproteinase-2
D011480:Protease Inhibitors; D011506:Proteins; D012333:RNA, Messenger; D019716:Tissue Inhibitor of Metalloproteinase-2; D018093:Gelatinases; D008666:Metalloendopeptidases; D020778:Matrix Metalloproteinase 2