Multiple forms of angiostatin induce apoptosis in endothelial cells.
Multiple forms of angiostatin induce apoptosis in endothelial cells.
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DOI：
10.1182/blood.v92.12.4730.424k20_4730_4741
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发表时间：
1998-12
期刊：
影响因子：
20.3
通讯作者：
R. Lucas;L. Holmgren;I. Garcia;B. Jiménez;S. J. Mandriota;F. Borlat;B. K. Sim;Zicheng Wu;G. Grau;Y. Shing;G. Soff;N. Bouck;M. Pepper
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文献类型：
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作者：
R. Lucas;L. Holmgren;I. Garcia;B. Jiménez;S. J. Mandriota;F. Borlat;B. K. Sim;Zicheng Wu;G. Grau;Y. Shing;G. Soff;N. Bouck;M. Pepper
血管生成抑制素是通过纤溶酶原的蛋白水解切割产生的血管生成的循环抑制剂。在本研究中，我们使用了重组人和鼠血管抑制素（kringles 1-4）以及天然人血管抑制素（通过纤溶酶原的弹性蛋白酶消化[kringles 1-3]或在游离巯基供体存在下纤溶酶自催化[kringles 1-4]制备）。我们报告，血管抑素减少内皮细胞数量在4天的增殖试验，而不影响细胞周期进展到S期（由溴脱氧尿苷标记确定）。这表明增殖试验中细胞数量的减少可能部分是由于细胞毒性。这一点通过观察到血管抑素以与肿瘤坏死因子-（TNF-）和转化生长因子-β 1（TGF-β 1）相似的方式增加了内皮细胞中的乙锭同二聚体掺入（质膜完整性的测量）得到证实，两者均诱导内皮细胞凋亡。与TNF-和TGF-β 1相反，血管抑素在人MRC-5成纤维细胞、大鼠平滑肌、犬MDCK上皮或鼠B16-F10黑色素瘤细胞系中未诱导细胞毒性。通过内皮细胞核吖啶橙子掺入以及膜联蛋白V和TUNEL染色证实血管抑素诱导的细胞凋亡。这些体外研究结果表明，内皮细胞凋亡是血管抑素体内抗血管生成作用的一种机制。
Angiostatin is a circulating inhibitor of angiogenesis generated by proteolytic cleavage of plasminogen. In this study we have used recombinant human and murine angiostatins (kringles 1-4) as well as native human angiostatin (prepared by elastase digestion of plasminogen [kringles 1-3] or by plasmin autocatalysis in the presence of a free sulfhydryl donor [kringles 1-4]). We report that angiostatin reduces endothelial cell number in a 4-day proliferation assay without affecting cell cycle progression into S-phase (as determined by bromodeoxyuridine labeling). This suggested that the reduction in cell number in the proliferation assay might in part be due to cytotoxicity. This was confirmed by the observation that ethidium homodimer incorporation (a measure of plasma membrane integrity) into endothelial cells was increased by angiostatin in a manner similar to that seen with tumor necrosis factor- (TNF-) and transforming growth factor-beta1 (TGF-beta1), both of which induce apoptosis in endothelial cells. In contrast to TNF- and TGF-beta1, angiostatin did not induce cytotoxicity in human MRC-5 fibroblast, rat smooth muscle, canine MDCK epithelial, or murine B16-F10 melanoma cell lines. Angiostatin-induced apoptosis was confirmed by endothelial cell nuclear acridine orange incorporation as well as by annexin V and TUNEL staining. These in vitro findings point to endothelial cell apoptosis as a mechanism for the antiangiogenic effect of angiostatin in vivo.