REQUIREMENT FOR RECEPTOR-BOUND UROKINASE IN PLASMIN-DEPENDENT CELLULAR CONVERSION OF LATENT TGF-BETA TO TGF-BETA
REQUIREMENT FOR RECEPTOR-BOUND UROKINASE IN PLASMIN-DEPENDENT CELLULAR CONVERSION OF LATENT TGF-BETA TO TGF-BETA
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DOI：
10.1002/jcp.1041580303
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发表时间：
1994-03-01
影响因子：
5.6
通讯作者：
RIFKIN, DB
中科院分区：
文献类型：
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作者：
ODEKON, LE;BLASI, F;RIFKIN, DB
在转染人uPA受体cDNA （LhuPAR(+)）、人prouPA cDNA （LhuPA）或对照新霉素耐药cDNA （Lneo）的小鼠LB6细胞模型系统中，研究了受体结合尿激酶型纤溶酶原激活剂（uPA）在细胞活化潜伏转化生长因子β (ltgf - β)中的作用。当LhuPAR（+）细胞与LhuPA细胞共培养时，产生的纤溶蛋白依赖的纤溶活性比LhuPA细胞数量增加5倍的同型培养或含有LhuPA和Lneo细胞而不是LhuPAR（+）细胞的共培养所产生的纤溶蛋白活性要高。三种独立的生物测定证实，具有最大纤溶酶生成潜力的LhuPAR（+）和LhuPA细胞共培养对tcf - β具有优先激活作用。在第一次实验中，用条件培养基（CM）处理的BAE细胞（LhuPA和LhuPAR（+）细胞）的PA活性降低了48%，PA活性是衡量tgf - β产生活性的指标。含有tcf - β的中和抗体消除了CM对PA活性的抑制作用，表明抑制分子是tcf - β。在共培养中添加uPA （ATF）的氨基末端片段或遗漏纤溶酶原会阻断CM中纤维蛋白溶解活性和tgf - β活性的产生。在第二个实验中，来自LhuPA和LhuPAR（+）细胞共培养的CM在伤口实验中抑制了BAE细胞的迁移。对照抗tgf - β IgG表明抑制作用是由tgf - β引起的。在第三个实验中，与不含纤溶酶原的相同细胞培养的CM或LhuPA与LhuPAR（-）细胞共培养的CM相比，LhuPA与LhuPAR（+）细胞共培养产生的CM抑制了水貂肺上皮细胞的增殖。根据BAE迁移和PA分析，过量甘露糖-6-磷酸（M6P）阻断了tcf - β的生成，可能是因为它干扰了ltcf - β的细胞表面定位。此外，少量LhuPA和LhuPAR（+）细胞与BAE细胞共培养，通过tgf - β的旁分泌作用抑制BAE细胞PA活性。这些结果支持了LB6细胞的纤溶蛋白依赖性ltcf - β激活是由其受体uPA的表面定位促进的。(C) 1994 Wiley-Liss, Inc。
The role of receptor-bound urokinase-type plasminogen activator (uPA) in cellular activation of latent transforming growth factor-beta (LTGF-beta) was investigated in a model system of mouse LB6 cells transfected with either a human uPA receptor cDNA (LhuPAR(+)), a human prouPA cDNA (LhuPA), or a control neomycin-resistance cDNA (Lneo). When LhuPAR(+) cells were co-cultured with LhuPA cells, the plasmin-dependent fibrinolytic activity generated was more than that observed in either homotypic cultures with fivefold greater number of LhuPA cells or co-cultures containing LhuPA and Lneo cells instead of the LhuPAR(+) cells. The preferential activation of TCF-beta by co-cultures with the greatest plasmin-generation potential, LhuPAR(+) and LhuPA cells, was confirmed by three independent bioassays. In the first assay, a 48% decrease in PA activity, a measure of active TGF-beta production, was observed with BAE cells treated with conditioned medium (CM) from co-cultures of LhuPA and LhuPAR(+) cells. Inclusion of neutralizing antibodies to TCF-beta abrogated the inhibitory effect of CM on PA activity demonstrating that the inhibitory molecule was TCF-beta. Addition of the amino terminal fragment of uPA (ATF) or omission of plasminogen from co-cultures blocked both the fibrinolytic activity and the generation of TGF-beta activity in the CM. In the second assay, CM from co-cultures of LhuPA and LhuPAR(+) cells inhibited the migration of BAE cells in a wound assay. Controls with anti-TGF-beta IgG indicated that the inhibition was due to TGF-beta. In the third assay, proliferation of mink lung epithelial cells was inhibited by CM generated by co-cultures of LhuPA and LhuPAR(+) cells as compared to CM from the same cells cultured in the absence of plasminogen or to CM from a co-culture of LhuPA with LhuPAR(-) cells. Excess mannose-6-phosphate (M6P) blocked the generation of TCF-beta as assayed by both the BAE migration and PA assays, presumably because it interfered with cell-surface localization of LTCF-beta. Additionally, small numbers of LhuPA and LhuPAR(+) cells co-cultured with BAE cells inhibited the BAE cell PA activity via the paracrine action of TGF-beta. These results support the conclusion that plasmin-dependent activation LTCF-beta by LB6 cells is promoted by the surface localization of uPA by its receptor. (C) 1994 Wiley-Liss, Inc.