Achbarou A, Kaiser S, Tremblay G, Ste-Marie L G, Brodt P, Goltzman D, Rabbani S A
Department of Medicine, McGill University, Montreal, Canada.
Cancer Res. 1994 May 1;54(9):2372-7.
We previously reported that urokinase (uPA) is produced by the human prostate cancer cell line, PC-3, and could function as a growth factor for cells of the osteoblast phenotype. To examine the role of uPA in metastasis to the skeleton and to extraskeletal sites, we have developed a homologous model of uPA overexpression in a rat prostate cancer cell line. Full length cDNA encoding rat (r) uPA was isolated and subcloned as a 1.4-kilobase XbaI-BspHI fragment in the sense and antisense orientation into the Moloney murine leukemia retroviral vector pYN. The control (pYN) and experimental (pYN-ruPA, pYN-ruPA-AS) plasmids were transfected into Dunning R 3227, Mat LyLu rat prostate carcinoma cells. Experimental clones expressing at least 5-fold higher (pYN-ruPA) or 3-fold lower (pYN-ruPA-AS) than controls were selected, and control and experimental cells were inoculated into the left ventricles of inbred male Copenhagen rats. Animals were sacrificed at timed intervals to examine the evolution of metastatic lesions. Control animals developed metastases to the lumbar vertebrae resulting in spinal cord compression and hind limb paralysis at 20-21 days postinoculation. Animals inoculated with cells overexpressing uPA developed hind limb paralysis significantly earlier (by day 14-15 postinoculation). Additionally, more widespread skeletal (ribs, scapula, and femora) metastases were seen. Serum from experimental animals showed a progressive elevation in alkaline phosphatase levels, and histological examination of lumbar metastases revealed markedly increased osteoblastic activity over that observed in control animals. In contrast to this, animals inoculated with cells underexpressing uPA developed hind limb paralysis significantly later (days 25-29 postinoculation) and displayed decreased tumor metastasis. These studies support a role for the catalytic domain of uPA in enhancing both skeletal and nonskeletal prostate cancer invasiveness and are consistent with a role for the growth factor domain of uPA in mediating an osteoblastic skeletal response.
我们之前报道过，尿激酶（uPA）由人前列腺癌细胞系PC-3产生，并且可作为成骨细胞表型细胞的生长因子发挥作用。为了研究uPA在骨骼及骨骼外部位转移中的作用，我们构建了大鼠前列腺癌细胞系中uPA过表达的同源模型。编码大鼠（r）uPA的全长cDNA被分离出来，并作为一个1.4千碱基的XbaI - BspHI片段，以正义和反义方向亚克隆到莫洛尼鼠白血病逆转录病毒载体pYN中。将对照质粒（pYN）和实验质粒（pYN - ruPA、pYN - ruPA - AS）转染到Dunning R 3227、Mat LyLu大鼠前列腺癌细胞中。挑选出表达水平比对照高至少5倍（pYN - ruPA）或低3倍（pYN - ruPA - AS）的实验克隆，将对照细胞和实验细胞接种到近交系雄性哥本哈根大鼠的左心室。在不同时间点处死动物，以检查转移灶的发展情况。对照动物在接种后20 - 21天出现腰椎转移，导致脊髓受压和后肢麻痹。接种过表达uPA细胞的动物后肢麻痹出现得明显更早（接种后第14 - 15天）。此外，还观察到更广泛的骨骼转移（肋骨、肩胛骨和股骨）。实验动物的血清碱性磷酸酶水平呈进行性升高，对腰椎转移灶的组织学检查显示，与对照动物相比，成骨细胞活性明显增强。与此相反，接种uPA低表达细胞的动物后肢麻痹出现得明显更晚（接种后第25 - 29天），并且肿瘤转移减少。这些研究支持uPA的催化结构域在增强前列腺癌骨骼及非骨骼侵袭性方面发挥作用，并且与uPA的生长因子结构域在介导成骨细胞骨骼反应方面的作用一致。
