Koutsilieris M, Frenette G, Lazure C, Lehoux J G, Govindan M V, Polychronakos C
Hormonal Bioregulation Laboratory, Hospital Center Laval University (C.H.U.L.), Ste. Foy, Quebec, Canada.
Anticancer Res. 1993 Mar-Apr;13(2):481-6.
The transplantation of PA-III rat prostate cancer cells onto rat skeleton produces osteoblastic metastases. Therefore w e studied the paracrine interactions between the PA-III cells and osteoblast-derived osteosarcoma cells (UMR 106 cells). A serine protease secreted by PA-III cells hydrolyzed IGF-binding protein-1 and IGF-binding protein-2 (IGFBP-1 and IGFBP-2) detected in the cell culture media (CM) of OMR 106 cells by western ligand blotting. The serine protease of PA-III cell CM was purified using a benzamidine affinity column. This protease was a protein of 45-50 kDa on polyacrylamide gel electrophoresis under non-reducing conditions but generated two protein bands under reducing conditions; a) one of 33-35 kDa possessing protease activity and b) another of 20-25 kDa which was proteinolytically inactive. Sequence analysis identified the amino acid sequence of the a-chain (20-25 kDa band) and of the b-chain (33-35 kDa band) of rat urokinase-type plasminogen activator molecule. Urokinase purified from PA-III cell CM hydrolyzed IGFBPs of UMR 106 cells and stimulated the proliferation of UMR 106 cells in serum-free cultures. Its protease activity was abolished by benzamidine and aprotinin. Its mitogenic activity for osteoblasts was inhibited by anti-IGF-I monoclonal antibody. Northern blot analysis documented the expression of the urokinase-type plasminogen activator gene in the mRNA extracted from PA-III cells. Urokinase expression was inhibited by dexamethasone. Therefore, we conclude that urokinase-type plasminogen activator stimulates osteoblasts via an IGF-I dependent mechanism. Hydrolysis of the IGFBOPs at the sites of PA-III cell-induced bone tumors account for an increased bioavailability of IGFs. This may facilitate the development and the growth of PA-III cell-induced bone tumor and can also mediate the subsequent local osteoblastic reaction.
将PA - III大鼠前列腺癌细胞移植到大鼠骨骼上会产生成骨细胞转移。因此，我们研究了PA - III细胞与成骨细胞衍生的骨肉瘤细胞（UMR 106细胞）之间的旁分泌相互作用。PA - III细胞分泌的一种丝氨酸蛋白酶通过western配体印迹法水解了在UMR 106细胞的细胞培养基（CM）中检测到的胰岛素样生长因子结合蛋白-1和胰岛素样生长因子结合蛋白-2（IGFBP - 1和IGFBP - 2）。使用苯甲脒亲和柱纯化PA - III细胞CM中的丝氨酸蛋白酶。在非还原条件下，这种蛋白酶在聚丙烯酰胺凝胶电泳上是一种45 - 50 kDa的蛋白质，但在还原条件下产生两条蛋白带；a）一条33 - 35 kDa具有蛋白酶活性，b）另一条20 - 25 kDa没有蛋白水解活性。序列分析确定了大鼠尿激酶型纤溶酶原激活剂分子的a链（20 - 25 kDa条带）和b链（33 - 35 kDa条带）的氨基酸序列。从PA - III细胞CM中纯化的尿激酶水解UMR 106细胞的IGFBPs，并在无血清培养中刺激UMR 106细胞的增殖。其蛋白酶活性被苯甲脒和抑肽酶消除。其对成骨细胞的促有丝分裂活性被抗IGF - I单克隆抗体抑制。Northern印迹分析记录了从PA - III细胞提取的mRNA中尿激酶型纤溶酶原激活剂基因的表达。尿激酶表达被地塞米松抑制。因此，我们得出结论，尿激酶型纤溶酶原激活剂通过IGF - I依赖性机制刺激成骨细胞。在PA - III细胞诱导的骨肿瘤部位IGFBOPs的水解导致IGFs生物利用度增加。这可能促进PA - III细胞诱导的骨肿瘤的发展和生长，也可以介导随后的局部成骨细胞反应。
