A novel urokinase receptor-targeted inhibitor for plasmin and matrix metalloproteinases suppresses vein graft disease
A novel urokinase receptor-targeted inhibitor for plasmin and matrix metalloproteinases suppresses vein graft disease
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DOI：
10.1093/cvr/cvq203
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发表时间：
2010-11-01
影响因子：
10.8
通讯作者：
Quax, Paul H. A.
中科院分区：
文献类型：
--
作者：
Eefting, Daniel;Seghers, Leonard;Quax, Paul H. A.
基质金属蛋白酶（MMP）和纤溶酶原激活物(PA)/纤溶酶介导的蛋白水解，特别是在细胞表面，在基质变性和平滑肌细胞迁移中起重要作用，这在很大程度上导致了静脉移植失败。在这项研究中，设计了一种新的杂交蛋白来同时抑制这两种蛋白酶系统。该杂交蛋白由尿激酶型PA的受体结合氨基末端片段（ATF）组成，与金属蛋白酶组织抑制剂（TIMP-1）和牛胰腺胰蛋白酶抑制剂（BPTI）（一种有效的蛋白酶抑制剂）连接，可抑制细胞表面的MMP和纤溶酶活性。研究了该蛋白过表达对静脉移植疾病的影响。编码杂交蛋白TIMP-1.ATF的非病毒表达载体。构建并验证了BPTI。接下来，用该载体转染培养的人静脉片段。表达timp - 1. atf。静脉段的BPTI在4周后导致新内膜形成平均减少36 +/- 14%。timp -1 - atf对静脉移植疾病的体内抑制作用。BPTI在高胆固醇血症APOE*3Leiden小鼠颈动脉静脉介入中得到证实。4周后，用TIMP-1、ATF或BPTI处理的小鼠的静脉移植物增厚明显受到抑制（减少36-49%）。在TIMP-1.ATF。bpti处理小鼠，静脉移植物增厚减少了67 +/- 4%，与单个成分相比也明显增强。这些数据证明了混合蛋白TIMP-1.ATF对PA和MMP系统的细胞表面抑制作用。BPTI在电穿孔介导的非病毒基因转移后在远端组织中过表达，是预防静脉移植疾病的一种有效方法。
Matrix metalloproteinases (MMP) and plasminogen activator (PA)/plasmin-mediated proteolysis, especially at the cell surface, play important roles in matrix degeneration and smooth muscle cell migration, which largely contributes to vein graft failure. In this study, a novel hybrid protein was designed to inhibit both protease systems simultaneously. MMP and plasmin activity were inhibited at the cell surface by this hybrid protein, consisting of the receptor-binding amino-terminal fragment (ATF) of urokinase-type PA, linked to both the tissue inhibitor of metalloproteinases (TIMP-1) and bovine pancreas trypsin inhibitor (BPTI), a potent protease inhibitor. The effect of overexpression of this protein on vein graft disease was studied.A non-viral expression vector encoding the hybrid protein TIMP-1.ATF.BPTI was constructed and validated. Next, cultured segments of human veins were transfected with this vector. Expressing TIMP-1.ATF.BPTI in vein segments resulted in a mean 36 +/- 14% reduction in neointima formation after 4 weeks. In vivo inhibition of vein graft disease by TIMP-1.ATF.BPTI is demonstrated in venous interpositions placed into carotid arteries of hypercholesterolaemic APOE*3Leiden mice. After 4 weeks, vein graft thickening was significantly inhibited in mice treated with the domains TIMP-1, ATF, or BPTI (36-49% reduction). In the TIMP-1.ATF.BPTI-treated mice, vein graft thickening was reduced by 67 +/- 4%, which was also significantly stronger when compared with the individual components.These data provide evidence that cell surface-bound inhibition of the PA and MMP system by the hybrid protein TIMP-1.ATF.BPTI, overexpressed in distant tissues after electroporation-mediated non-viral gene transfer, is a powerful approach to prevent vein graft disease.