Ziccardi R J
Department of Immunology, IMM-11, Research Institute of Scripps Clinic, La Jolla, CA 92037.
Mol Immunol. 1988 Dec;25(12):1339-46. doi: 10.1016/0161-5890(88)90049-1.
We had previously demonstrated that in normal human serum (NHS) nascent C3b inhibited C1 activation by immune complexes (IC). We have now investigated the mechanism of this feedback inhibition. For these studies, EA-IgG were added to solutions containing physiological concns of purified C1, C1-In, C2, C3 and C4. Mixtures were then incubated at 37 degrees C for 30 min. Western blot and autoradiographic analyses revealed that almost half of the IgG molecules had become covalently linked to C3b in a 1:1 complex with the C3 alpha' chain of C3b being bound to the heavy chain of IgG. IgG-C3b and free IgG were separated by ion exchange chromatography and immune complexes were formed with each. The consumption of complement in NHS by EA-IgG and EA-(IgG-C3b) were then compared. The results indicate that binding of C3b to IgG did not significantly inhibit the C1 activating potential of the IgG. Thus feedback inhibition is not due to the binding of C3b to IgG. An alternative mechanism was next explored. After incubation of EA-IgG with C1 through C3, EA were separated from supernatant fluid by centrifugation. It was determined that one-third to one-half of the IgG had been released from the erythrocytes. Release appears not to have been due to C3b binding to IgG, since the released IgG-C3b readily bind to fresh sheep erythrocyte (E), and since IgG that was free of C3b was also released from EA by complement, it is more likely that C3b binding to the E caused the dissociation of antibody. These results indicate that under physiological conditions, the C1 activating potential of an immune complex is greatly reduced as the result of the binding of nascent C3b to the antigen moiety of the IC, thereby causing the displacement of complement activating antibody. In addition to IgG, IgG-C3b is also released from the IC.
我们之前已经证明，在正常人血清（NHS）中，新生的C3b可抑制免疫复合物（IC）激活C1。我们现在研究了这种反馈抑制的机制。在这些研究中，将EA-IgG添加到含有生理浓度的纯化C1、C1-In、C2、C3和C4的溶液中。然后将混合物在37℃孵育30分钟。蛋白质印迹和放射自显影分析显示，几乎一半的IgG分子已与C3b共价连接形成1:1复合物，其中C3b的C3α'链与IgG的重链结合。通过离子交换色谱分离IgG-C3b和游离IgG，并分别形成免疫复合物。然后比较EA-IgG和EA-(IgG-C3b)对NHS中补体的消耗情况。结果表明，C3b与IgG的结合并未显著抑制IgG激活C1的潜力。因此，反馈抑制不是由于C3b与IgG的结合。接下来探索了另一种机制。将EA-IgG与C1至C3孵育后，通过离心从上层清液中分离出EA。确定有三分之一至二分之一的IgG已从红细胞中释放出来。释放似乎不是由于C3b与IgG的结合，因为释放的IgG-C3b很容易与新鲜绵羊红细胞（E）结合，并且由于不含C3b的IgG也通过补体从EA中释放出来，更有可能是C3b与E的结合导致了抗体的解离。这些结果表明，在生理条件下，由于新生的C3b与IC的抗原部分结合，免疫复合物激活C1的潜力大大降低，从而导致补体激活抗体的置换。除了IgG外，IgG-C3b也从IC中释放出来。
