Bosco Domenico, Rouiller Dominique G, Halban Philippe A
Surgical Research Unit, Department of Surgery, Cell Isolation and Transplantation Center, CMU, Geneva University Hospitals, 1, rue Michel-Servet, 1211 Geneva-4, Switzerland.
J Endocrinol. 2007 Jul;194(1):21-9. doi: 10.1677/JOE-06-0169.
The aim of this study was to assess whether the expression of E-cadherin at the surface of rat beta-cells is regulated by insulin secretagogues and correlates with insulin secretion. When cultured under standard conditions, virtually all beta-cells expressed E-cadherin observed by immunofluorescence, but heterogeneous staining was observed. Using fluorescence-activated cell sorting (FACS), two beta-cell sub-populations were sorted: one that was poorly labeled ('ECad-low') and another that was highly labeled ('ECad-high'). After 1-h stimulation with 16.7 mM glucose, insulin secretion (reverse hemolytic plaque assay) from individual ECad-high beta-cells was higher than that from ECad-low beta-cells. Ca2+-dependent beta-cell aggregation was increased at 16.7 mM glucose when compared with 2.8 mM glucose. E-cadherin at the surface of beta-cells was increased after 18 h at 11.1 and 22.2 mM glucose when compared with 2.8 mM glucose, with the greatest increase at 22.2 mM glucose + 0.5 mM isobutylmethylxanthine (IBMX). While no labeling was detected on freshly trypsinized cells, the proportion of stained cells increased in a time-dependent manner during culture for 1, 3, and 24 h. This recovery was faster when cells were incubated at 16.7 vs 2.8 mM glucose. Cycloheximide inhibited expression of E-cadherin at 2.8 mM glucose, but not at 16.7 mM, while depolymerization of actin by either cytochalasin B or latrunculin B increased surface E-cadherin at low glucose. In conclusion, these results show that expression of E-cadherin at the surface of islet beta-cells is controlled by secretagogues including glucose, correlates with insulin secretion, and can serve as a surface marker of beta-cell function.
本研究的目的是评估大鼠β细胞表面E-钙黏蛋白的表达是否受胰岛素促分泌剂调节，并与胰岛素分泌相关。在标准条件下培养时，通过免疫荧光观察到几乎所有β细胞均表达E-钙黏蛋白，但染色存在异质性。使用荧光激活细胞分选（FACS）技术，分选了两个β细胞亚群：一个标记较弱的亚群（“ECad-low”）和另一个标记较强的亚群（“ECad-high”）。用16.7 mM葡萄糖刺激1小时后，单个ECad-highβ细胞的胰岛素分泌（反向溶血空斑试验）高于ECad-lowβ细胞。与2.8 mM葡萄糖相比，16.7 mM葡萄糖时Ca2+依赖性β细胞聚集增加。与2.8 mM葡萄糖相比，在11.1和22.2 mM葡萄糖培养18小时后，β细胞表面的E-钙黏蛋白增加，在22.2 mM葡萄糖 + 0.5 mM异丁基甲基黄嘌呤（IBMX）时增加最为显著。虽然在新鲜胰蛋白酶消化的细胞上未检测到标记，但在培养1、3和24小时期间，染色细胞的比例呈时间依赖性增加。当细胞在16.7 mM葡萄糖而非2.8 mM葡萄糖中孵育时，这种恢复更快。放线菌酮在2.8 mM葡萄糖时抑制E-钙黏蛋白的表达，但在16.7 mM葡萄糖时不抑制，而细胞松弛素B或拉春库林B使肌动蛋白解聚可增加低葡萄糖时的表面E-钙黏蛋白。总之，这些结果表明，胰岛β细胞表面E-钙黏蛋白的表达受包括葡萄糖在内的促分泌剂控制，与胰岛素分泌相关，可作为β细胞功能的表面标志物。
