Taraboletti G, Sonzogni L, Vergani V, Hosseini G, Ceruti R, Ghilardi C, Bastone A, Toschi E, Borsotti P, Scanziani E, Giavazzi R, Pepper M S, Stetler-Stevenson W G, Bani M R
Department of Oncology, Mario Negri Institute for Pharmacological Research, Bergamo, 24125, Italy.
Exp Cell Res. 2000 Aug 1;258(2):384-94. doi: 10.1006/excr.2000.4936.
Matrix metalloproteinases (MMPs) play a critical role in the development of hemangioma-like vascular tumors in mice injected with murine eEnd.1 endothelioma cells. The current study was designed to (a) characterize the presence of MMPs in the vascular tumor, (b) define whether these MMPs originate from the transformed cells or from the recruited stromal cells and (c) study the stimulatory effect of eEnd.1 cells on the production of MMPs by endothelial cells. Several gelatinases were present in the eEnd.1 tumor extract, including latent and activated MMP-2 (72-kDa gelatinase A, EC 3.4.24. 24) and MMP-9 (92-kDa gelatinase B, EC 3.4.24.35). Immunohistochemical analysis of the tumor revealed focal reactivity for MMP-2. No gelatinase was produced by cultured eEnd.1 cells, or by six of nine related endothelioma cell lines, suggesting that stroma cells, particularly endothelial cells recruited by the tumor cells, rather than eEnd.1 cells themselves, are the source of the gelatinases observed in the tumors in vivo. The conditioned medium of eEnd.1 cells stimulated the release of MMP-2 and MMP-1 (interstitial collagenase, EC 3.4.24.7) by endothelial cells, but not of the inhibitor TIMP-2. The increased production of MMP-2 and MMP-1, observed at the protein level (zymogram and Western blot analysis), occurred through a posttranscriptional mechanism, since no increase in mRNA was observed and the stimulation was not prevented by inhibitors of protein synthesis. The inhibitory effects of monensin and brefeldin A, inhibitors of protein secretion, and the decrease in cell-associated MMP-2 in stimulated endothelial cells indicated that regulation occurred mostly at the level of protease secretion. MMPs are known to be regulated at different levels; this study indicates that, in endothelial cells, the stimulation of MMPs can also occur at the level of secretion, a mechanism that provides a rapid mobilization of these crucial enzymes in the early phases of angiogenesis.
基质金属蛋白酶（MMPs）在注射了鼠源eEnd.1内皮瘤细胞的小鼠中，对血管瘤样血管肿瘤的发展起着关键作用。当前的研究旨在：（a）鉴定血管肿瘤中MMPs的存在情况；（b）确定这些MMPs是源自转化细胞还是募集的基质细胞；（c）研究eEnd.1细胞对内皮细胞产生MMPs的刺激作用。eEnd.1肿瘤提取物中存在几种明胶酶，包括潜伏和活化的MMP-2（72 kDa明胶酶A，EC 3.4.24.24）和MMP-9（92 kDa明胶酶B，EC 3.4.24.35）。对肿瘤的免疫组织化学分析显示MMP-2呈局灶性反应。培养的eEnd.1细胞或九个相关内皮瘤细胞系中的六个均未产生明胶酶，这表明基质细胞，特别是肿瘤细胞募集的内皮细胞，而非eEnd.1细胞本身，是体内肿瘤中观察到的明胶酶的来源。eEnd.1细胞的条件培养基刺激内皮细胞释放MMP-2和MMP-1（间质胶原酶，EC 3.4.24.7），但不刺激抑制剂TIMP-2的释放。在蛋白质水平（酶谱和蛋白质印迹分析）观察到的MMP-2和MMP-1产量增加是通过转录后机制发生的，因为未观察到mRNA增加，且蛋白质合成抑制剂不能阻止这种刺激。莫能菌素和布雷菲德菌素A（蛋白质分泌抑制剂）的抑制作用以及受刺激的内皮细胞中细胞相关MMP-2的减少表明，调节主要发生在蛋白酶分泌水平。已知MMPs在不同水平受到调节；本研究表明，在内皮细胞中，MMPs也可在分泌水平受到刺激，这一机制可在血管生成早期快速调动这些关键酶。
