Serrano J, Bevins C L, Young S W, de Pablo F
Receptors and Hormone Action Section, National Institute of Diabetes, Digestive and Kidney Diseases, Bethesda, Maryland 20892.
Dev Biol. 1989 Apr;132(2):410-8. doi: 10.1016/0012-1606(89)90237-6.
Insulin has metabolic, growth, and differentiation effects in chicken embryos in vivo and it is required for normal development. Whether the pancreas is the sole source of insulin in embryogenesis is controversial. In the present study we investigated (1) the developmental pattern of expression of the chicken insulin gene in the pancreas; (2) the expression of the insulin gene in three nonpancreatic tissues, liver, brain, and lower limb, during chicken development; and (3) the expression of the insulin gene at prepancreatic stages and during chicken embryo organogenesis. Hybridization of synthetic species-specific insulin oligonucleotides to pancreatic frozen section in situ and to Northern blots revealed a major increase in insulin messenger RNA (mRNA) levels during the third week of embryonic development. The hybridization histochemistry showed both an increase in the levels of insulin mRNA per pancreatic islet and, in addition, an increase in the number of insulin mRNA containing islets with development. By Northern analysis there was a major polyadenylated transcript of 0.6 kb, which increased in abundance approximately 30-fold during this interval. Under the same stringency conditions used for pancreatic RNA an insulin transcript was detected in liver RNA blots. The abundance of this hepatic insulin mRNA was about 100-fold less than the pancreatic insulin mRNA and, in contrast to the latter, did not increase in late development. Primer extension experiments demonstrated that the insulin transcripts of pancreas and liver had similar 5' ends. No insulin mRNA was detected by Northern analysis or primer extension either in whole brain or lower limb total RNA from several developmental stages. A very low abundance insulin mRNA was detected in whole embryo at Day 8 and body regions at Day 4 and Day 5 when organogenesis of the pancreas takes place. Interestingly, a polyadenylated insulin transcript was detected, as well, in whole Day 2 and Day 3 embryos (stages 10 to 20, with 20 to 40 somites) before differentiation of beta cells occurs. Thus, there is differential developmental regulation of the insulin gene in several chicken embryo tissues and the expression of insulin precedes pancreatic maturation. These findings support the proposed role of insulin in differentiation and development in vivo and suggest a paracrine type of action of the hormone in early embryos before blood circulation begins.
胰岛素在鸡胚体内具有代谢、生长和分化作用，是正常发育所必需的。胰腺是否是胚胎发生过程中胰岛素的唯一来源存在争议。在本研究中，我们调查了：（1）鸡胰岛素基因在胰腺中的表达发育模式；（2）鸡发育过程中胰岛素基因在肝脏、大脑和下肢这三种非胰腺组织中的表达；（3）胰腺前阶段及鸡胚器官发生过程中胰岛素基因的表达。合成的物种特异性胰岛素寡核苷酸与胰腺冰冻切片原位杂交以及与Northern印迹杂交显示，胚胎发育第三周期间胰岛素信使核糖核酸（mRNA）水平大幅增加。杂交组织化学显示，每个胰岛中胰岛素mRNA水平增加，此外，含胰岛素mRNA的胰岛数量也随发育增加。通过Northern分析，有一个0.6 kb的主要多聚腺苷酸化转录本，在此期间其丰度增加了约30倍。在用于胰腺RNA的相同严谨条件下，在肝脏RNA印迹中检测到胰岛素转录本。这种肝脏胰岛素mRNA的丰度比胰腺胰岛素mRNA少约100倍，与后者不同的是，在发育后期并未增加。引物延伸实验表明，胰腺和肝脏的胰岛素转录本具有相似的5'端。在几个发育阶段的全脑或下肢总RNA中，通过Northern分析或引物延伸均未检测到胰岛素mRNA。在胰腺发生器官形成的第8天全胚以及第4天和第5天的身体区域检测到极低丰度的胰岛素mRNA。有趣的是，在β细胞分化之前的第2天和第3天全胚（第10至20阶段，有20至40个体节）中也检测到了多聚腺苷酸化的胰岛素转录本。因此，鸡胚的几种组织中胰岛素基因存在差异发育调控，且胰岛素的表达先于胰腺成熟。这些发现支持了胰岛素在体内分化和发育中的作用，并表明该激素在血液循环开始前的早期胚胎中具有旁分泌作用类型。
