McMillan W D, Patterson B K, Keen R R, Pearce W H
Department of Surgery, Northwestern University Medical School, Chicago, IL 60611, USA.
J Vasc Surg. 1995 Sep;22(3):295-305. doi: 10.1016/s0741-5214(95)70144-3.
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.
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.
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.
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.
72千道尔顿IV型胶原酶（基质金属蛋白酶-2，MMP-2）是一种强效的胶原酶和弹性蛋白酶，存在于炎症疾病状态中，可能在主动脉瘤的发病机制中起重要作用。MMP-2或其抑制剂金属蛋白酶组织抑制剂2型（TIMP-2）表达的改变可能会增加细胞外基质降解并导致动脉瘤形成。本研究的目的是：（1）测量人肾下腹主动脉瘤、闭塞性病变和正常组织中MMP-2和TIMP-2 mRNA的相对组织水平；（2）检测培养的动脉瘤和正常血管平滑肌细胞（VSMC）的表达情况；（3）在原位鉴定动脉瘤、闭塞性病变和正常主动脉壁内产生mRNA的细胞。
从动脉瘤组织（n = 8）、闭塞性病变组织（n = 9）和正常组织（n = 7）中提取总RNA，进行Northern分析。将MMP-2和TIMP-2的信号与α-微管蛋白进行标准化。采用方差分析比较平均值±标准误。培养动脉瘤和正常VSMC，传代并生长至汇合后进行RNA提取和Northern分析。用洋地黄毒苷RNA探针进行原位杂交，在动脉瘤（n = 7）、闭塞性病变（n = 4）和正常（n = 3）主动脉的组织切片中定位产生MMP-2和TIMP-2 mRNA的细胞。
动脉瘤主动脉组织中MMP-2 mRNA水平（1.032±0.164，n = 5）显著高于闭塞性病变主动脉（0.553±0.027，n = 4，p < 0.02）或正常主动脉（0.230±0.038，n = 3，p < 0.002）。TIMP-2 mRNA水平差异不显著（动脉瘤主动脉0.207±0.042，n = 3；闭塞性病变主动脉0.413±0.164，n = 3；正常主动脉0.260±0.079，n = 4；p = 0.34），尽管样本数量较少。培养的动脉瘤和正常VSMC组成性表达MMP-2和TIMP-2。原位研究表明，动脉瘤外膜炎症周围的VSMC和巨噬细胞中组织MMP-2和TIMP-2表达共定位，但在闭塞性病变主动脉中则定位于动脉粥样硬化斑块。
MMP-2和TIMP-2在动脉瘤、闭塞性病变和正常主动脉中均有表达。动脉瘤中MMP-2的表达显著高于闭塞性病变或正常主动脉。培养的动脉瘤VSMC组成性表达MMP-2和TIMP-2。原位观察到的差异表达模式以及动脉瘤与闭塞性病变主动脉中组织MMP-2 mRNA水平升高表明，MMP-2可能与闭塞性疾病中的局部斑块重塑以及动脉瘤中外膜胶原和弹性蛋白的弥漫性破坏有关。
