Hanford Holly Anne, Wong Christie A, Kassan Hallie, Cundiff Deborah L, Chandel Navdeep, Underwood Suzanne, Mitchell Christopher A, Soff Gerald A
Northwestern University, Feinberg School of Medicine, Department of Medicine, Division of Hematology/Oncology, Chicago, Illinois 60611, USA.
Cancer Res. 2003 Jul 15;63(14):4275-80.
Angiostatin, a proteolytic cleavage product of plasminogen, acts via a selective, yet poorly understood mechanism to potently inhibit angiogenesis (M. S. O'Reilly et al., Cell, 79: 315-328, 1994). Vascular endothelial cell proliferation assays revealed that angiostatin(4.5), a naturally occurring human isoform consisting of plasminogen kringle domains 1-4 and most of kringle domain 5 (G. A. Soff, Cancer Metastasis Rev., 19: 97-107, 2000), dose dependently reduces cell number despite the presence of a potent stimulus of proliferation. Flow cytometry using the vital dyes Hoechst 33342 and Pyronin Y revealed that approximately 40% of both control and angiostatin(4.5)-treated cells were in the proliferative phase, indicating that cell cycle progression is not impaired by exposure to angiostatin(4.5). Both bovine aortic endothelial cells and human umbilical endothelial cells were shown to undergo apoptosis in response to angiostatin(4.5). Caspases-3, -8, and -9 activation, specified by cleavage of fluorophore-conjugated specific peptide substrates, revealed a cascade of caspase activation that peaks at 36 h of angiostatin(4.5) treatment. Angiostatin(4.5) exposure induced release of cytochrome c from mitochondria in a caspase-dependent manner, but a pan-caspase inhibitor, zVAD-fmk, blocked cytochrome c release. Overall, these data indicate that human angiostatin(4.5) may function in vivo to block blood vessel formation by specifically inducing vascular endothelial cells to apoptose in a process likely involving both the intrinsic and extrinsic apoptosis pathways.
血管抑素是纤溶酶原的一种蛋白水解裂解产物，通过一种选择性但尚未完全了解的机制发挥作用，能有效抑制血管生成（M. S. 奥赖利等人，《细胞》，79: 315 - 328，1994年）。血管内皮细胞增殖试验表明，血管抑素（4.5）是一种天然存在的人类异构体，由纤溶酶原kringle结构域1 - 4和大部分kringle结构域5组成（G. A. 索夫，《癌症转移评论》，19: 97 - 107，2000年），尽管存在强大的增殖刺激，但它仍能剂量依赖性地减少细胞数量。使用活性染料Hoechst 33342和派洛宁Y进行的流式细胞术显示，对照细胞和经血管抑素（4.5）处理的细胞中约40%处于增殖期，这表明暴露于血管抑素（4.5）不会损害细胞周期进程。牛主动脉内皮细胞和人脐静脉内皮细胞均显示对血管抑素（4.5）有凋亡反应。通过荧光团偶联的特异性肽底物的裂解来确定的半胱天冬酶 - 3、 - 8和 - 9的激活，揭示了一个半胱天冬酶激活级联反应，并在血管抑素（4.5）处理36小时时达到峰值。血管抑素（4.5）的暴露以半胱天冬酶依赖性方式诱导线粒体释放细胞色素c，但一种泛半胱天冬酶抑制剂zVAD - fmk可阻断细胞色素c的释放。总体而言，这些数据表明，人类血管抑素（4.5）可能在体内发挥作用，通过特异性诱导血管内皮细胞凋亡来阻断血管形成，这一过程可能涉及内在和外在凋亡途径。
