Pleiotrophic inhibition of pericellular urokinase-type plasminogen activator system by endogenous tumor suppressive maspin.
Pleiotrophic inhibition of pericellular urokinase-type plasminogen activator system by endogenous tumor suppressive maspin.
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发表时间：
2001-12
期刊：
影响因子：
11.2
通讯作者：
H. Biliran;S. Sheng
中科院分区：
文献类型：
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作者：
H. Biliran;S. Sheng
Maspin是一种新型的丝氨酸蛋白酶抑制剂，具有肿瘤抑制活性，抑制肿瘤侵袭和转移。到目前为止，maspin的潜在分子机制仍然不清楚。重组maspin被证明能特异性地抑制细胞表面相关的尿激酶型纤溶酶原激活物(UPA)和纤维蛋白原结合的组织型纤溶酶原激活物。然而，内源性maspin在纤溶酶原激活中的作用还完全不清楚。为了解决这个问题，我们使用前列腺癌细胞DU145作为亲本细胞系，建立了稳定表达maspin的转染体。我们在此报告内源性maspin对细胞周围的uPA系统具有多向抑制作用。Maspin的表达导致与细胞表面结合的uPA和uPA受体蛋白水平显著降低，而不改变各自mRNAs的稳定水平。低密度脂蛋白受体相关蛋白的特异性抑制剂受体相关蛋白(RAP)处理后，maspin转染组的uPA分泌量和细胞表面uPAR水平显著增加，而模型对照组细胞表面uPA和uPAR水平无明显变化。酶和分子分析的结合表明，maspin通过与细胞表面结合的uPA形成耐十二烷基硫酸钠的复合体来抑制细胞表面介导的纤溶酶原激活。此外，maspin的表达显著减少了条件培养液中活性uPA的释放，包括高分子量和低分子量。一致地，maspin转基因克隆的条件培养基使纤溶酶原转化为纤溶酶的活性显著降低。Maspin对细胞周围uPA的抑制作用与显著降低细胞侵袭能力和体外运动能力有关。Maspin中和抗体(Abs4A)以剂量依赖的方式逆转maspin转基因细胞的侵袭潜能。综上所述，本研究首次提供了内源性maspin是细胞周围uPA的有效抑制物的证据。此外，我们的结果支持了目前的假设，即maspin通过抑制细胞周围的局部蛋白分解来阻止肿瘤的侵袭和运动。
Maspin is a novel serine protease inhibitor with tumor suppressive activity, inhibiting tumor invasion and metastasis. To date, the underlying molecular mechanism of maspin remains elusive. Recombinant maspin has been shown to specifically inhibit cell surface-associated urokinase-type plasminogen activator (uPA) and fibrinogen-bound tissue-type plasminogen activator. However, the role of endogenous maspin in plasminogen activation is totally unknown. To address this issue, we generated stable maspin-expressing transfectants using prostate carcinoma cells DU145 as the parental cell line. We report here that endogenous maspin exerts pleiotropic inhibitory effects on the pericellular uPA system. Maspin expression led to a significantly reduced level of cell surface-bound uPA and uPA receptor proteins without altering the steady-state levels of the respective mRNAs. Treatment with receptor-associated protein (RAP), a specific inhibitor of low-density lipoprotein receptor-related protein, lead to a significantly increased level of secreted uPA and cell surface uPAR in maspin transfectants but not in the mock control cells. A combination of enzymatic and molecular analyses revealed that maspin inhibits the cell surface-mediated plasminogen activation by forming an SDS-resistant complex with cell surface-bound uPA. In addition, maspin expression led to a dramatic reduction in the release of active uPA, both high molecular weight and the low molecular weight, into the conditioned culture medium. Consistently, the conditioned medium of maspin transfectant clones had a significantly reduced activity in converting plasminogen to plasmin. The inhibitory effect of maspin on pericellular uPA correlates with significantly decreased cell invasion potential and motility in vitro. The maspin-neutralizing antibody (Abs4A) reversed the subdued invasive potential of maspin transfectant cells in a dose-dependent manner. In summary, this study provides the first evidence that endogenous maspin is a potent inhibitor of pericellular uPA. Furthermore, our results support a current hypothesis that maspin blocks tumor invasion and motility by inhibiting localized pericellular proteolysis.