Chen L, Komiya I, Inman L, McCorkle K, Alam T, Unger R H
Center for Diabetes Research, University of Texas Southwestern Medical Center, Dallas 75235.
Proc Natl Acad Sci U S A. 1989 Feb;86(4):1367-71. doi: 10.1073/pnas.86.4.1367.
We have evaluated in situ hybridization histochemistry as a means of estimating simultaneously the level of prohormone mRNA and the dimensions of rat pancreatic islets. Localization of the 27-mer 32P-labeled oligonucleotide probes for rat proinsulin I, glucagon, and prosomatostatin I corresponded with localization of antibodies to the three hormones. In normal rats subjected to chronic hyperglycemic clamping, the density of the proinsulin mRNA signal increased 54%, islet size and number increased approximately 100%, while proglucagon mRNA signal was reduced 81%. Resection of 50% of the pancreas increased proinsulin mRNA 36% and proglucagon mRNA 500%; islet area doubled and islet number increased 50%. In 150-day-old diabetic ob/ob mice, there was an 18-fold expansion in islet area, a 4-fold increase in islet number, but no increase in insulin gene expression. In insulin-dependent streptozotocin-treated diabetic rats, islet area and number were profoundly reduced; insulin deprivation failed to raise proinsulin mRNA in surviving beta cells above control levels. Proglucagon mRNA was high despite the hyperglycemia but was reduced by insulin within 1 hr, suggesting that insulin regulates glucagon gene expression or is required for its regulation by glucose. In situ hybridization of rat islets provides a valid semiquantitative index of insulin and glucagon biosynthesis and of islet dimensions and reveals that normal but not diabetic islets meet increased insulin demand by increasing both number and biosynthetic activity of beta cells.
我们已评估原位杂交组织化学，将其作为一种同时估计前激素mRNA水平和大鼠胰岛大小的方法。针对大鼠胰岛素原I、胰高血糖素和前生长抑素I的27聚体32P标记寡核苷酸探针的定位与这三种激素抗体的定位一致。在接受慢性高血糖钳夹的正常大鼠中，胰岛素原mRNA信号密度增加54%，胰岛大小和数量增加约100%，而胰高血糖素原mRNA信号减少81%。切除50%的胰腺使胰岛素原mRNA增加36%，胰高血糖素原mRNA增加500%；胰岛面积加倍，胰岛数量增加50%。在150日龄的糖尿病ob/ob小鼠中，胰岛面积扩大了18倍，胰岛数量增加了4倍，但胰岛素基因表达没有增加。在胰岛素依赖型链脲佐菌素处理的糖尿病大鼠中，胰岛面积和数量显著减少；胰岛素缺乏未能使存活的β细胞中的胰岛素原mRNA升高至对照水平以上。尽管存在高血糖，但胰高血糖素原mRNA仍很高，但在1小时内被胰岛素降低，这表明胰岛素调节胰高血糖素基因表达或其受葡萄糖调节所必需。大鼠胰岛的原位杂交提供了胰岛素和胰高血糖素生物合成以及胰岛大小的有效半定量指标，并揭示正常而非糖尿病胰岛通过增加β细胞数量和生物合成活性来满足增加的胰岛素需求。
