Instituto de Investigaciones Bioquímicas and UNESCO Chair of Biophysics & Molecular Neurobiology, 8000 Bahía Blanca, Argentina.
J Cell Biochem. 2011 Nov;112(11):3276-88. doi: 10.1002/jcb.23254.
The neuronal-type α7 nicotinic acetylcholine receptor (α7AChR) is also found in various non-neural tissues, including vascular endothelium, where its peculiar ionotropic properties (high Ca(2+) permeability) and its supervening Ca(2+) -mediated intracellular cascades may play important roles in physiology (angiogenesis) and pathology (inflammation and atherogenesis). Changes in molecular (up-regulation, affinity, and conformational states) and cellular (distribution, association with membranes) properties of the α7AChR related to angiogenesis (wound-repair cell migration) and atherogenesis (alterations in cholesterol content) were studied in living endothelial cells, with the aim of determining whether such changes constitute early markers of inflammatory response. The combination of pharmacological, biochemical, and fluorescence microscopy tools showed that α7AChRs in rat arterial endothelial (RAEC) and human venous endothelial (HUVEC) cells occur at extremely low expression levels (∼50 fmol/mg protein) but undergo agonist-induced up-regulation at relatively high nicotine concentrations (∼300-fold with 50 µM ligand), increasing their cell-surface exposure. When analyzed in terms of cold Triton X-100 solubility and subcellular distribution, α7AChRs occur in the "non-raft" subcellular membrane fractions. Acute cholesterol depletion reduced not only cholesterol levels but also the number of cell-surface α7AChRs. Nicotine exposure markedly stimulated cell migration and accelerated wound repair, which drastically diminished in cells deprived of the sterol. The angiogenic effect of nicotine appears to be synergistic with cholesterol content. Finally, the apparent K(D) of α7AChRs for the open-channel blocker crystal violet was found to be ∼600-fold lower in receptor-enriched membranes obtained from up-regulated HUVEC.
神经元型 α7 烟碱型乙酰胆碱受体（α7AChR）也存在于各种非神经组织中，包括血管内皮细胞，其独特的离子型特性（高 Ca(2+)通透性）及其随之而来的 Ca(2+)介导的细胞内级联反应可能在生理学（血管生成）和病理学（炎症和动脉粥样硬化形成）中发挥重要作用。研究了与血管生成（伤口修复细胞迁移）和动脉粥样硬化形成（胆固醇含量改变）相关的α7AChR 的分子（上调、亲和力和构象状态）和细胞（分布、与膜的关联）特性的变化在活的内皮细胞中，目的是确定这些变化是否构成炎症反应的早期标志物。药理学、生化和荧光显微镜工具的结合表明，大鼠动脉内皮（RAEC）和人静脉内皮（HUVEC）细胞中的α7AChR 表达水平极低（约 50 fmol/mg 蛋白），但在相对较高的尼古丁浓度下（用 50 µM 配体诱导约 300 倍）诱导其上调，增加其细胞表面暴露。从冷 Triton X-100 溶解度和亚细胞分布的角度分析，α7AChR 存在于“非筏”亚细胞膜部分。急性胆固醇耗竭不仅降低了胆固醇水平，还降低了细胞表面α7AChR 的数量。尼古丁暴露明显刺激细胞迁移并加速伤口修复，而在缺乏固醇的细胞中则急剧减少。尼古丁的血管生成作用似乎与胆固醇含量具有协同作用。最后，发现从上调的 HUVEC 中获得的富含受体的膜中，α7AChR 对开放通道阻滞剂结晶紫的表观 K(D)降低了约 600 倍。
