Pepper M S, Matsumoto K, Nakamura T, Orci L, Montesano R
Department of Morphology, University of Geneva Medical Center, Switzerland.
J Biol Chem. 1992 Oct 5;267(28):20493-6.
We have recently demonstrated that fibroblast-conditioned medium induces Madin-Darby canine kidney (MDCK) epithelial cells to form branching tubules when grown in three-dimensional collagen or fibrin gels (Montesano, R., Schaller, G., and Orci, L. (1991) Cell 66, 697-711), and that this morphogenetic effect is mediated by hepatocyte growth factor (HGF), also known as scatter factor (Montesano, R., Matsumoto, K., Nakamura, T., and Orci, L. (1991) Cell 67, 901-908). In fibrin gels, this effect is inhibited by addition of exogenous serine protease inhibitors, which suggests a role for plasminogen activators (PAs) in the matrix remodeling required for tubulogenesis. In the studies reported in this paper, we have investigated the effect of fibroblast-conditioned medium (CM) and HGF on the production of PAs by MDCK cells. We have found that urokinase-type PA (u-PA) activity and mRNA are increased 4.9-fold by CM from human MRC-5 fibroblasts, which has tubulogenic activity, but not by CM from human Detroit-550 fibroblasts, which lacks tubulogenic activity. The u-PA inductive property of MRC-5 CM was completely inhibited by preincubation with antibodies to recombinant human HGF (rhHGF). Exogenously added rhHGF also increased u-PA activity and mRNA 5.9-fold in MDCK cells, with an optimal effect at approximately 10 ng/ml. MRC-5 CM also increased u-PA receptor mRNA 34.9-fold in MDCK cells, an effect which was inhibited by 71% by preincubating the CM with antibodies to rhHGF, and which was mimicked by exogenously added rhHGF (31.3-fold increase). These results demonstrate that HGF, which induces tubulogenesis by MDCK cells in vitro, also increases u-PA and u-PA receptor expression in these cells. Taken together with our previous observations, this suggests that the resulting increase in extracellular proteolysis, appropriately localized to the cell surface, is required for epithelial morphogenesis.
我们最近证明，当在三维胶原蛋白或纤维蛋白凝胶中生长时，成纤维细胞条件培养基可诱导马-达二氏犬肾（MDCK）上皮细胞形成分支小管（蒙特萨诺，R.，沙勒，G.，和奥尔西，L.（1991年）《细胞》66卷，697 - 711页），并且这种形态发生效应是由肝细胞生长因子（HGF）介导的，HGF也被称为分散因子（蒙特萨诺，R.，松本，K.，中村，T.，和奥尔西，L.（1991年）《细胞》67卷，901 - 908页）。在纤维蛋白凝胶中，添加外源性丝氨酸蛋白酶抑制剂可抑制这种效应，这表明纤溶酶原激活剂（PAs）在小管形成所需的基质重塑中起作用。在本文报道的研究中，我们研究了成纤维细胞条件培养基（CM）和HGF对MDCK细胞产生PAs的影响。我们发现，具有小管形成活性的人MRC - 5成纤维细胞的CM可使尿激酶型PA（u - PA）活性和mRNA增加4.9倍，但缺乏小管形成活性的人底特律550成纤维细胞的CM则无此作用。用抗重组人HGF（rhHGF）抗体预孵育可完全抑制MRC - 5 CM的u - PA诱导特性。外源性添加的rhHGF也可使MDCK细胞中的u - PA活性和mRNA增加5.9倍，在约10 ng/ml时具有最佳效果。MRC - 5 CM还可使MDCK细胞中的u - PA受体mRNA增加34.9倍，用抗rhHGF抗体预孵育CM可使该效应降低71%，而外源性添加rhHGF可模拟该效应（增加31.3倍）。这些结果表明，在体外诱导MDCK细胞发生小管形成的HGF，也可增加这些细胞中u - PA和u - PA受体的表达。结合我们之前的观察结果，这表明细胞表面适当定位的细胞外蛋白水解作用增强是上皮形态发生所必需的。
