Cloning of new endothelin B receptor and its physiological significance

1.为了研究内皮素（ET）受体介导兔和人大隐静脉平滑肌收缩的亚型，考察了几种ET受体激动剂和拮抗剂的作用。我们的研究结果表明，ET_A和ET_B受体介导的收缩均具有药理异质性（ET_<A1>, ET_<A2>, ET_<B1>, ET_<B2>）。目前可获得的数据，以及本研究中获得的数据，清楚地表明内皮素反应的药理学异质性，但它们还不足以为内皮素受体命名提供确凿的证据。然而，使用ET_B缺陷小鼠进行的其他药理学数据清楚地表明，ET_B受体的这些药理学异质性反应完全消失，这表明ET_<B1>和ET_<B2>受体来源于单个ET_B基因2。先天性巨结肠病（Hirschsprung's disease， HSCR）的特点是末端肠缺乏自主神经节细胞，是新生儿肠梗阻的一个相对常见的原因。最近，ET_B基因已被证明是HSCR的易感基因，在该基因零突变的小鼠中产生神经节结肠，并在HSCR高发病率的大近亲（Mennonite家系）中证明了一个错义突变。然而，在其他临床病例中未发现进一步的突变。我们分析了31例分离的HSCR患者中ET_B基因的改变。在核苷酸311,325和1170处检测到三种新的突变：A到T， T到C和C到A的转变。这些突变导致Asn变为Ile， Cys变为Arg， Ser变为Arg，预计会产生无功能的ET_B受体。这些观察结果表明，ET_B受体功能障碍或功能丧失可能与一些孤立HSCR患者的病因有关。

1.Analysis of endothelin receptor subtypesTo study endothelin (ET) receptor subtypes which mediate smooth muscle contraction of the rabbit and human saphenous veins, effects of some ET receptor agonists and antagonists were examined. Our results indicated pharmacological heterogeneity (ET_<A1>, ET_<A2> ; ET_<B1>, ET_<B2>) of both ET_A and ET_B recetor-mediated contractions. The currently available data, as well as those obtained in the present study, clearly indicate pharmacological heterogeneity of responses to endothelins, but they are not yet sufficient to provide conclusive evidence for endothelin receptor nomenclature. However, additional pharmacological data using ET_B-deficient mice clearly indicated that these pharmacologically heterogenous responses of ET_B receptor are completely lost, suggesting that ET_<B1> and ET_<B2> receptors are derived from a single ET_B gene.2.Mutation analysis of endothelin B receptor in Hirschsprung diseaseHirschsprung's disease (HSCR) is characterized by the absence of autonomic ganglion cells in the terminal bowel and is a relatively common cause of intestinal obstruction in the newborn. Recently, ET_B gene has been shown as a susceptibility gene for HSCR by the production of aganglionic colon in mice with a null mutation of this gene and by demonstrating a missense mutation in a large inbred kindred with a high incidence of HSCR (Mennonite pedigree). However, no further mutations have been demonstrated in other clinical cases. We analyzed alterations of the ET_B gene in 31 isolated patients of HSCR.Three novel mutations were detected : A to T, T to C and C to A transitions at nucleotide 311,325 and 1170, respectively. These mutations resulted in Asn to Ile, Cys to Arg and Ser to Arg, predicted to produce a non-functional ET_B receptor. These observations indicate that dysfunction or loss of function of ET_B receptor may be involved in the aetiology of some isolated patients with HSCR.