Enhanced anti-angiogenic effect of a deletion mutant plasminogen kringle 5 on of neovascularization
Enhanced anti-angiogenic effect of a deletion mutant plasminogen kringle 5 on of neovascularization
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DOI：
10.1002/jcb.20601
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发表时间：
2005-12-15
影响因子：
4
通讯作者：
Gao, GQ
中科院分区：
文献类型：
--
作者：
Cai, WB;Ma, JF;Gao, GQ
Kringle 5（K5）是纤溶酶原的蛋白水解片段，已被证明是血管生成抑制剂。在此之前，我们已经评估了K5对氧诱导视网膜病变大鼠模型中血管渗漏和新生血管形成的影响。在这项研究中，我们表达K5和K5的缺失突变体（K5突变体）在原核细胞表达系统，并通过亲和层析纯化。K5突变体通过从K5中删除11个氨基酸而保留三个二硫键来产生。在内皮细胞和视网膜新生血管大鼠模型中比较完整的K5和K5突变体的抗血管生成活性。K5突变体以浓度依赖性方式抑制原代人视网膜毛细血管内皮细胞（HRCEC）的增殖，其表观EC 50约为35 nmol/L，是完整K5的2倍。在甚至更高的浓度范围内，K5突变体不抑制来自相同来源的HRCEC的周细胞，这表明内皮细胞特异性抑制。K5突变体在高浓度范围内对正常肝细胞和Bel 7402肝癌细胞也无作用。如荧光素血管造影所示，在氧诱导的视网膜病变大鼠模型中玻璃体内注射K5和突变体均导致比注射PBS的对照组显著更少的新生血管簇和非灌注区域。此外，通过定量血管细胞，K5突变体表现出比完整的K5更强的抑制新生血管形成的作用。这些结果表明，这种K5缺失突变体是一个更有效的血管生成抑制剂比完整的K5，并可能在治疗这些疾病与新生血管，如实体瘤，糖尿病视网膜病变，年龄相关性黄斑变性，类风湿性关节炎，前列腺增生的治疗潜力。J.细胞。96：1254-1261，2005. (c)2005 Wiley-Liss，Inc.
Kringle 5 (K5), a proteolytic fragment of plasminogen, has been proved to be an angiogenic inhibitor. Previously, we have evaluated the effect of K5 on the vascular leakage and neovascularization in a rat model of oxygen-induced retinopathy. In this study, we expressed K5 and a deletion mutant of K5 (K5 mutant) in a prokaryocyte expression system and purified them by affinity chromatography. K5 mutant was generated by deleting 11 amino acids from K5 while retaining the three disulfide bonds. The anti-angiogenic activity of intact K5 and K5 mutant were compared in endothelial cells and retinal neovascularization rat model. K5 mutant inhibited the proliferation of primary human retinal capillary endothelial cells (HRCEC) in a concentration-dependent manner, with an apparent EC50 of approximate 35 nmol/L,which is twofold more potent than intact K5. In the even higher concentration range, K5 mutant did not inhibit pericytes from the same origin of HRCEC, which suggested an endothelial cell-specific inhibition. K5 mutant had no effect on normal liver cells and Bel7402 hepatoma cells even at high concentration range either. Intravitreal injection of the K5 and mutant in the oxygen-induced retinopathy rat model both resulted in significantly fewer neovascular tufts and nonperfusion area than controls with PBS injection, as shown by fluorescein angiography. Furthermore, K5 mutant exhibited more strong inhibition effect on neovascularization than intact K5 by quantification of vascular cells. These results suggest that this K5 deletion mutant is a more potent angiogenic inhibitor than intact K5 and may have therapeutic potential in the treatment of those disorders with neovascularization, such as solid tumor, diabetic retinopathy, age-related macular degeneration, rheumatoid arthritis, and hyperplasia of prostate. J. Cell. Biochem. 96: 1254-1261, 2005. (c) 2005 Wiley-Liss, Inc.