Pathological and biochemical significance of the complex between kininogens and thiol proteinases

针对交联的高分子量激肽原-钙蛋白酶I复合物（CL复合物）产生单克隆抗体。所获得的5株单克隆抗体分别命名为HCC- 1、2、3、4和5，它们与CL复合物反应，但与其新生组分高分子量激肽原和钙蛋白酶I均不反应。它们的免疫球蛋白类别和轻链类型分别为IgM和κ。只有HCC-3和HCC-4能识别非交联的高分子量激肽原-钙蛋白酶I复合物（N复合物），而其它HCC-3和HCC-4则不能。HCC-3比HCC-4更强地识别N复合物。HCC-3还识别其他N复合物，高分子量激肽原-钙蛋白酶II复合物，低分子量激肽原-钙蛋白酶I复合物和低分子量激肽原-钙蛋白酶II复合物。HCC-3识别的表位可能是高分子量激肽原与钙蛋白酶I相互作用形成的特异性结构，其结构与四种激肽原-钙蛋白酶复合物相同。采用HCC-3的夹心ELISA测定系统，可以估计CL复合物的复合物的量高达1 ng/ml，N复合物的复合物的量高达100 ng/ml。17例正常人血浆中N复合物含量均小于100 ng/ml。然而，我们可以在10个肝细胞癌患者的血浆样品中检测到4个复合物（Max. 426 ng/ml），3例肝硬化患者中有2例（最大311 ng/ml），以及1例肝母细胞瘤（290 ng/ml）。这是首次报道表明体内存在与抗激肽原和钙蛋白酶复合物的单克隆抗体的复合物。

Monoclonal antibodies were raised against cross-linked high molecular weight kininogen- calpain I complex (CL complex). Five monoclonal antibodies obtained were designated as HCC- 1, 2, 3, 4 and 5, and found to react with CL complex, but not with its nascent components, high molecular weight kininogen or calpain I either. Their immunoglobulin class and light chain type are IgM and kappa, respectively. Only HCC-3 and -4 recognized non cross-linked high molecular weight kininogen-calpain I complex (N complex), but the other did not. HCC-3 recognized the N complex stronger than HCC-4. HCC-3 also recognized other N complexes, high molecular weight kininogen-calpain II complex, low molecular weight kininogen-calpain I complex and low molecular weight kininogen-calpain II complex. The epitope recognized by HCC-3 would be the specific structure formed by the interaction between high molecular weight kininogen and calpain I, and its structure common to four kininogen-calpain complexes. Employing sandwich ELISA assay system with HCC-3, it was possible to estimate the amount of the complex up to 1 ng/ml for CL complex and 100 ng/ml for N complex. The level of N complex in normal plasma (n=17) was less than 100 ng/ml. However, we could detect the complex in 4 of 10 plasma samples from the patients with hepatocellular carcinoma (Max. 426 ng/ml), 2 of 3 with liver cirrhosis (max. 311 ng/ml), and 1 of 1 with hepatoblastoma (290 ng/ml).This is the first report which indicated the presence of the complex in vivo with the monoclonal antibody against a complex between kininogen and calpain.