Zetser Anna, Bashenko Yulia, Edovitsky Evgeny, Levy-Adam Flonia, Vlodavsky Israel, Ilan Neta
Cancer and Vascular Biology Research Center, The Bruce Rappaport Faculty of Medicine, Technion, Haifa, Israel.
Cancer Res. 2006 Feb 1;66(3):1455-63. doi: 10.1158/0008-5472.CAN-05-1811.
Heparanase is an endo-beta-D-glucuronidase involved in cleavage of heparan sulfate moieties and hence participates in extracellular matrix (ECM) degradation and remodeling. Traditionally, heparanase activity was correlated with the metastatic potential of a variety of tumor-derived cell types. Cloning of the heparanase gene indicated that heparanase expression is up-regulated in a variety of primary human tumors. In some cases, heparanase up-regulation correlated with increased tumor vascularity, an angiogenic feature that could be recapitulated in a number of in vitro and in vivo models. The mechanism by which heparanase enhances angiogenic responses is not entirely clear but is thought to be mediated primarily by release of ECM-resident angiogenic growth factors such as basic fibroblast growth factor and vascular endothelial growth factor (VEGF). Here, we examined the possibility that heparanase directly participates in VEGF gene regulation. We provide evidence that heparanase overexpression in human embryonic kidney 293, MDA-MB-435 human breast carcinoma, and rat C6 glioma cells resulted in a 3- to 6-fold increase in VEGF protein and mRNA levels, which correlated with elevation of p38 phosphorylation. Moreover, heparanase down-regulation in B16 mouse melanoma cells by a specific siRNA vector was accompanied by a decrease in VEGF and p38 phosphorylation levels, suggesting that VEGF gene expression is regulated by endogenous heparanase. Interestingly, a specific p38 inhibitor did not attenuate VEGF up-regulation by heparanase whereas Src inhibitors completely abrogated this effect. These results indicate, for the first time, that heparanase is actively involved in the regulation of VEGF gene expression, mediated by activation of Src family members.
乙酰肝素酶是一种内切-β-D-葡糖醛酸酶，参与硫酸乙酰肝素部分的裂解，因此参与细胞外基质（ECM）的降解和重塑。传统上，乙酰肝素酶活性与多种肿瘤来源细胞类型的转移潜能相关。乙酰肝素酶基因的克隆表明，乙酰肝素酶在多种原发性人类肿瘤中表达上调。在某些情况下，乙酰肝素酶上调与肿瘤血管生成增加相关，这一血管生成特征可在许多体外和体内模型中重现。乙酰肝素酶增强血管生成反应的机制尚不完全清楚，但认为主要是由细胞外基质驻留的血管生成生长因子如碱性成纤维细胞生长因子和血管内皮生长因子（VEGF）的释放介导的。在此，我们研究了乙酰肝素酶直接参与VEGF基因调控的可能性。我们提供的证据表明，在人胚肾293细胞、MDA-MB-435人乳腺癌细胞和大鼠C6胶质瘤细胞中过表达乙酰肝素酶，导致VEGF蛋白和mRNA水平增加3至6倍，这与p38磷酸化的升高相关。此外，用特异性siRNA载体下调B16小鼠黑色素瘤细胞中的乙酰肝素酶，伴随着VEGF和p38磷酸化水平的降低，表明VEGF基因表达受内源性乙酰肝素酶调控。有趣的是，一种特异性p38抑制剂并未减弱乙酰肝素酶对VEGF的上调作用，而Src抑制剂则完全消除了这种作用。这些结果首次表明，乙酰肝素酶通过Src家族成员的激活，积极参与VEGF基因表达的调控。
