WI38人肺成纤维细胞中缓激肽受体亚型的选择性标记

Selective labelling of bradykinin receptor subtypes in WI38 human lung fibroblasts.

Phagoo S B, Yaqoob M, Brown M C, Burgess G M

Sandoz Institute for Medical Research, London.

Br J Pharmacol. 1996 Nov;119(5):863-8. doi: 10.1111/j.1476-5381.1996.tb15752.x.

Abstract

Binding of the B1 bradykinin receptor radioligand, [3H]-des-Arg10-kallidin (-KD) and the B2 receptor radioligand [3H]-bradykinin (-BK) was investigated in membranes prepared from WI38 human foetal lung fibroblasts. 2. One-site analysis of the saturation data for [3H]-des-Arg10-KD gave an equilibrium dissociation constant (KD) value of 0.51 +/- 0.12 nM and a maximum receptor density (Bmax) of 260 +/- 49 fmol mg-1 of protein. [3H]-des-Arg10-KD binding was displaced by ligands in the order: des-Arg10-KD > KD > > des-Arg9[Leu8]-BK > des-Arg9-BK > Hoe 140 > > BK, implying that it was binding selectively to B1 receptors. 3. One-site analysis of the binding of [3H]-BK to W138 membranes indicated that it had a KD value of 0.25 +/- 0.06 nM and a Bmax of 753 +/- 98 fmol mg-1 of protein. The potencies for displacement of [3H]-BK binding were: Hoe 140 > > BK = KD > > > des-Arg10-KD = des-Arg9[Leu8]-BK = des-Arg9-BK, which was consistent with binding to B2 receptors. 4. This is the first characterization of [3H]-des-Arg10-KD binding to include both kinetic and equilibrium data, and demonstrates that [3H]-des-Arg10-KD has a high affinity for human B1 bradykinin receptors and is sufficiently selective to be used as a radioligand for B1 receptors in human cells or tissues expressing an excess of B2 BK receptors.

摘要

在WI38人胚肺成纤维细胞制备的膜中研究了B1缓激肽受体放射性配体[3H]-去-Arg10-胰激肽（-KD）和B2受体放射性配体[3H]-缓激肽（-BK）的结合情况。2. 对[3H]-去-Arg10-KD饱和数据的单点分析得出平衡解离常数（KD）值为0.51±0.12 nM，最大受体密度（Bmax）为260±49 fmol/mg蛋白质。[3H]-去-Arg10-KD的结合被配体以如下顺序取代：去-Arg10-KD＞KD＞＞去-Arg9[Leu8]-BK＞去-Arg9-BK＞Hoe 140＞＞BK，这表明它选择性地与B1受体结合。3. 对[3H]-BK与W138膜结合的单点分析表明，其KD值为0.25±0.06 nM，Bmax为753±98 fmol/mg蛋白质。[3H]-BK结合的取代效力顺序为：Hoe 140＞＞BK = KD＞＞＞去-Arg10-KD = 去-Arg9[Leu8]-BK = 去-Arg9-BK，这与与B2受体结合一致。4. 这是首次对[3H]-去-Arg10-KD结合进行的包括动力学和平衡数据的表征，并证明[3H]-去-Arg10-KD对人B1缓激肽受体具有高亲和力，且具有足够的选择性，可作为在表达过量B2 BK受体的人细胞或组织中B1受体的放射性配体。