Castle V P, Dixit V M, Polverini P J
Department of Pediatric Hematology/Oncology, University of Michigan School of Medicine, Ann Arbor 48109-0684, USA.
Lab Invest. 1997 Jul;77(1):51-61.
Thrombospondin-1 (TSP1) is a multifunctional matrix protein that influences the growth and function of a variety of normal and neoplastic epithelial and mesenchymal cell types. In vivo, TSP1 has shown potent antitumor activity in suppressing tumor neovascularization. Paradoxically, however, as we have reported, NIH 3T3 fibroblasts overexpressing TSP1 acquire the transformation-associated phenotypes of serum and anchorage independence in vitro but fail to form tumors in nude mice. To investigate these divergent results, and to determine the functional domains in TSP1 that confer serum and anchorage independence as well as antitumor and antiangiogenic activities, we transfected a series of deletion constructs of TSP1 into NIH 3T3 cells and into a v-src-transformed NIH 3T3 line. The antiangiogenic activity of TSP1-expressing, v-src-transformed NIH 3T3 cells was examined by assaying the conditioned media for inhibition of endothelial cell chemotaxis and suppression of basic fibroblast growth factor-mediated angiogenesis in the rat cornea. The link between TSP1 antitumor and antiangiogenic activities was assessed by measuring the rate of tumor growth and counting factor VIII-stained microvessels in the solid tumors developing in nude mice. Our results indicate that v-src NIH 3T3 cells transfected with a 449-amino acid N-terminal domain of TSP1 exhibit a dose-dependent suppression of tumor growth and neovascularization in nude mice. Truncated forms of TSP1 containing the type 1 properdin domain suppressed both endothelial cell chemotaxis and comeal neovascularization. Furthermore, when full-length TSP1 and deletion constructs containing the antiangiogenic type I properdin domain were transfected into highly tumorigenic v-src-transformed NIH 3T3 cells, they were able to confer transdominant suppression of tumorigenicity and angiogenesis of these cells in nude mice. These results confirm the role of TSP1 as a potent inhibitor of angiogenesis and provide support for the notion that alterations in the net balance between inducers and inhibitors of angiogenesis are largely responsible for the sustained growth of solid tumors in vivo.
血小板反应蛋白-1（TSP1）是一种多功能基质蛋白，可影响多种正常和肿瘤性上皮及间充质细胞类型的生长和功能。在体内，TSP1在抑制肿瘤新生血管形成方面显示出强大的抗肿瘤活性。然而，矛盾的是，正如我们所报道的，过表达TSP1的NIH 3T3成纤维细胞在体外获得了与转化相关的血清非依赖性和锚定非依赖性表型，但在裸鼠中未能形成肿瘤。为了研究这些不同的结果，并确定TSP1中赋予血清非依赖性和锚定非依赖性以及抗肿瘤和抗血管生成活性的功能域，我们将一系列TSP1缺失构建体转染到NIH 3T3细胞和v-src转化的NIH 3T3细胞系中。通过检测条件培养基对内皮细胞趋化性的抑制作用以及对大鼠角膜中碱性成纤维细胞生长因子介导的血管生成的抑制作用，来检测表达TSP1的v-src转化的NIH 3T3细胞的抗血管生成活性。通过测量裸鼠体内实体瘤的肿瘤生长速率并计数因子VIII染色的微血管，评估TSP1抗肿瘤和抗血管生成活性之间的联系。我们的结果表明，用TSP1的449个氨基酸的N端结构域转染的v-src NIH 3T3细胞在裸鼠中表现出剂量依赖性的肿瘤生长和新生血管形成抑制作用。含有1型备解素结构域的TSP1截短形式抑制了内皮细胞趋化性和角膜新生血管形成。此外，当将全长TSP1和含有抗血管生成I型备解素结构域的缺失构建体转染到高致瘤性v-src转化的NIH 3T3细胞中时，它们能够在裸鼠中赋予对这些细胞致瘤性和血管生成的反式显性抑制作用。这些结果证实了TSP1作为血管生成有效抑制剂的作用，并支持了以下观点：血管生成诱导剂和抑制剂之间净平衡的改变在很大程度上是体内实体瘤持续生长的原因。
