Establishment of 2-mercaptoethanol-dependent differentiated insulin-secreting cell lines.
Establishment of 2-mercaptoethanol-dependent differentiated insulin-secreting cell lines.
复制标题
DOI：
10.1210/endo.130.1.1370150
复制
发表时间：
1992
期刊：
影响因子：
4.8
通讯作者：
M. Asfari;D. Janjic;P. Meda;Guodong Li;P. Halban;C. Wollheim
中科院分区：
文献类型：
--
作者：
M. Asfari;D. Janjic;P. Meda;Guodong Li;P. Halban;C. Wollheim
新的胰岛素分泌细胞系（INS-1和INS-2）建立从细胞分离的X射线诱导的大鼠可移植胰岛素瘤。发现这些细胞的连续生长依赖于还原剂2-巯基乙醇。这种巯基化合物的去除导致总细胞谷胱甘肽水平下降15倍。这些细胞增殖缓慢（群体倍增时间约100小时），一般情况下，显示出典型的天然β细胞的形态特征。大多数细胞对胰岛素染色呈阳性，并且不与针对其他胰岛激素的抗体反应。免疫反应性胰岛素的含量约为8 μ g/10（6）个细胞，相当于天然β细胞含量的20%。这些细胞合成胰岛素原I和II，并显示两种前体激素的转化率与大鼠胰岛中观察到的相似。然而，葡萄糖未能刺激胰岛素原生物合成的速率。在静态孵育中，葡萄糖刺激从漂浮的细胞簇或从附着的细胞分泌胰岛素。在灌流条件下，10 mM但不是1 mM葡萄糖使分泌增加2.2倍。在毛喉素和3-异丁基-1-甲基黄嘌呤的存在下，葡萄糖浓度从2.8-20 mM的增加引起分泌速率的4倍增强。葡萄糖也使INS-1细胞去极化，并升高胞浆Ca ~（2+）浓度。这表明葡萄糖仍然能够在质膜上引发部分离子事件，从而导致胰岛素分泌。INS-1细胞的结构和功能特征在2年的时间内（约80代）保持不变。虽然INS-2细胞尚未完全表征，但它们的胰岛素含量与INS-1细胞的胰岛素含量相似，并且它们也对作为促分泌素的葡萄糖保持部分敏感。INS-1细胞保留β-细胞表面抗原，如通过与抗神经节苷脂单克隆抗体R2 D 6和A2 B5的反应性所揭示的。这些发现表明INS-1细胞保持稳定并保持高度分化，这应该使它们成为研究β细胞功能各个方面的合适模型。
New insulin-secreting cell lines (INS-1 and INS-2) were established from cells isolated from an x-ray-induced rat transplantable insulinoma. The continuous growth of these cells was found to be dependent on the reducing agent 2-mercaptoethanol. Removal of this thiol compound caused a 15-fold drop in total cellular glutathione levels. These cells proliferated slowly (population doubling time about 100 h) and, in general, showed morphological characteristics typical of native beta-cells. Most cells stained positive for insulin and did not react with antibodies against the other islet hormones. The content of immunoreactive insulin was about 8 micrograms/10(6) cells, corresponding to 20% of the native beta-cell content. These cells synthesized both proinsulin I and II and displayed conversion rates of the two precursor hormones similar to those observed in rat islets. However, glucose failed to stimulate the rate of proinsulin biosynthesis. In static incubations, glucose stimulated insulin secretion from floating cell clusters or from attached cells. Under perifusion conditions, 10 mM but not 1 mM glucose enhanced secretion 2.2-fold. In the presence of forskolin and 3-isobutyl-1-methylxanthine, increase of glucose concentration from 2.8-20 mM caused a 4-fold enhancement of the rate of secretion. Glucose also depolarized INS-1 cells and raised the concentration of cytosolic Ca2+. This suggests that glucose is still capable of eliciting part of the ionic events at the plasma membrane, which leads to insulin secretion. The structural and functional characteristics of INS-1 cells remained unchanged over a period of 2 yr (about 80 passages). Although INS-2 cells have not been fully characterized, their insulin content was similar to that of INS-1 cells and they also remain partially sensitive to glucose as a secretagogue. INS-1 cells retain beta-cell surface antigens, as revealed by reactivity with the antigangloside monoclonal antibodies R2D6 and A2B5. These findings indicate that INS-1 cells have remained stable and retain a high degree of differentiation which should make them a suitable model for studying various aspects of beta-cell function.