Rosenthal R A, Moses M A, Shintani Y, Megyesi J F, Langer R, Folkman J
Department of Surgery, Children's Hospital, Boston, Massachusetts 02115.
J Cell Biochem. 1994 Sep;56(1):97-105. doi: 10.1002/jcb.240560114.
We have previously shown that mouse sarcoma 180 cells produce vascular endothelial growth factor [VEGF; Rosenthal et al., 1990, Growth Factors, 4: 53-59], an endothelial mitogen that stimulates angiogenesis. Recent reports have implicated metalloproteinases and their inhibitors in the regulation of vascular morphogenesis, tumor invasion, and metastasis. We report here that mouse sarcoma 180 cells produce two collagenase inhibitors. These inhibitors were purified by heparin-Sepharose affinity chromatography, gel filtration, and C4 reverse phase h.p.l.c. Analytical gel electrophoresis of the purified inhibitors (MS-22 and MS-31) revealed molecular masses of 22,000 and 31,000 Da under reducing conditions, and 20,000 and 30,000 Da under nonreducing conditions, respectively. The NH2-terminal amino acid sequence of MS-22 was identical to that of tissue inhibitor of metalloproteinases type 2 (TIMP-2) produced by human melanoma cells [Stetler-Stevenson et al., 1989, J. Biol. Chem. 264: 17374-17378) over the first 30 amino acids. The NH2-terminal amino acid sequence of MS-31 was identical to that of murine TIMP-1 [Gewert et al., 1989, EMBO J 6:651-657]. Statistical analysis of the amino acid composition data of these two mouse sarcoma 180-derived collagenase inhibitors confirms the identification of MS-22 as TIMP-2 and MS-31 as TIMP-1.
我们先前已表明，小鼠肉瘤180细胞可产生血管内皮生长因子[VEGF；Rosenthal等人，1990年，《生长因子》，4：53 - 59]，一种刺激血管生成的内皮细胞有丝分裂原。最近的报告表明金属蛋白酶及其抑制剂参与血管形态发生、肿瘤侵袭和转移的调控。我们在此报告，小鼠肉瘤180细胞可产生两种胶原酶抑制剂。这些抑制剂通过肝素 - 琼脂糖亲和层析、凝胶过滤和C4反相高效液相色谱法进行纯化。纯化后的抑制剂（MS - 22和MS - 31）在还原条件下的分析凝胶电泳显示分子量分别为22,000和31,000 Da，在非还原条件下分别为20,000和30,000 Da。MS - 22的NH2末端氨基酸序列在前30个氨基酸上与人黑色素瘤细胞产生的金属蛋白酶组织抑制剂2（TIMP - 2）[Stetler - Stevenson等人，1989年，《生物化学杂志》264：17374 - 17378]相同。MS - 31的NH2末端氨基酸序列与鼠源TIMP - 1 [Gewert等人，1989年，《欧洲分子生物学组织杂志》6：651 - 657]相同。对这两种源自小鼠肉瘤180的胶原酶抑制剂的氨基酸组成数据进行统计分析，证实MS - 22为TIMP - 2，MS - 31为TIMP - 1。
