Monoclonal antibody to the thrombin receptor stimulates DNA synthesis in combination with gamma-thrombin or phorbol myristate acetate.
Monoclonal antibody to the thrombin receptor stimulates DNA synthesis in combination with gamma-thrombin or phorbol myristate acetate.
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DOI：
10.1083/jcb.105.6.2551
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发表时间：
1987-12
期刊：
影响因子：
--
通讯作者：
Carney DH
中科院分区：
文献类型：
--
作者：
Frost GH;Thompson WC;Carney DH
对不同凝血酶衍生物的研究表明，凝血酶启动细胞增殖需要两种不同类型的信号：一种是凝血酶或DIP-凝血酶(通过二异丙基磷酸盐使B链第205位的α-凝血酶失活)与受体高亲和力相互作用产生的；另一种是凝血酶的酶活性产生的。为了进一步研究高亲和力凝血酶受体在启动过程中的作用，我们用完整的人成纤维细胞免疫小鼠，并选择了阻断125I-凝血酶与仓鼠成纤维细胞上高亲和力受体结合的抗体。其中一种抗体，tr-9，能抑制80%到100%的125I-凝血酶结合，表现出与凝血酶与这些细胞上的受体结合的免疫荧光模式，并选择性地结合溶解的凝血酶受体。Tr-9本身并不启动DNA合成，也不阻止凝血酶的启动，但tr-9在α-凝血酶、γ-凝血酶(0.5微克/毫升)或PMA存在的情况下加入细胞，可刺激胸腺嘧啶核苷的掺入，最高可达对照组的三倍。在所有的情况下，最大的刺激观察到的浓度为TR-9，从1到4 nM，对应于从30%到100%抑制125I-凝血酶结合所需的浓度。这些结果表明，单抗与α-凝血酶受体的结合可以模拟凝血酶与该受体的高亲和力相互作用在刺激细胞增殖方面的作用。
Studies with various thrombin derivatives have shown that initiation of cell proliferation by thrombin requires two separate types of signals: one, generated by high affinity interaction of thrombin or DIP-thrombin (alpha-thrombin inactivated at ser 205 of the B chain by diisopropylphosphofluoridate) with receptors and the other, by thrombin's enzymic activity. To further study the role of high affinity thrombin receptors in initiation, we immunized mice with whole human fibroblasts and selected antibodies that blocked the binding of 125I- thrombin to high affinity receptors on hamster fibroblasts. One of these antibodies, TR-9, inhibits from 80 to 100% of 125I-thrombin binding, exhibits an immunofluorescent pattern indistinguishable from that of thrombin bound to receptors on these cells, and selectively binds solubilized thrombin receptors. By itself, TR-9 did not initiate DNA synthesis nor did it block thrombin initiation, but TR-9 addition to cells in the presence of alpha-thrombin, gamma-thrombin (0.5 microgram/ml), or PMA stimulated thymidine incorporation up to threefold over controls. In all cases, maximal stimulation was observed at concentrations of TR-9, ranging from 1 to 4 nM corresponding to concentrations required to inhibit from 30 to 100% of 125I-thrombin binding. These results demonstrate that the binding of the monoclonal antibody to the alpha-thrombin receptor can mimic the effects of thrombin's high affinity interaction with this receptor in stimulating cell proliferation.