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目的:明确大鼠胰腺发育不同时期胰岛素合成及释放相关基因的表达差异,探讨胰岛素分泌功能完善的过程。方法:分离并提取SD大鼠胚胎发育12.5天(E12.5)、E15.5、E18.5、新生、出生后21天及成年大鼠胰腺组织,利用Affymetrix公司的高密度寡核苷酸芯片检测大鼠发育各阶段胰腺组织基因表达谱并分析胰岛素合成及释放相关基因的特异及差异表达情况。分别进行正常新生鼠、成年鼠血糖水平及正常新生第1、3、7、10、12天大鼠腹腔葡萄糖耐量试验(IPGTT)。采用ELISA法测定新生鼠和成年鼠血中胰岛素浓度。新生大鼠胰岛细胞原代培养观察新生鼠胰岛细胞的形态。结果:在胰岛素合成及释放相关基因中,胰岛素基因及其特异性转录因子如PDX-1在大鼠E12.5～E15.5时即开始有表达,以后胰岛素基因持续存在且表达增多;胰岛素胞吐过程中的关键基因Syntaxin-1a、Vamp-2、Rab-3a、Munc13-1等从E15.5开始出现,以后表达量亦增多;而葡萄糖刺激的胰岛素分泌(GSIS)相关基因中,葡萄糖转运体-2(Glut2)在胚胎早期开始表达,葡萄糖激酶(GCK)在胚胎中晚期才表达。自E18.5即检测到大鼠血中胰岛素的存在,随着胚胎的发育,血胰岛素浓度逐渐升高。新生大鼠对葡萄糖负荷不能耐受,出现IPGTT异常,出生1周以后IPGTT才逐渐趋于正常。新生鼠的胰岛形态与成年鼠相似,但体积相对较小,透亮度稍差。结论:大鼠胚胎胰腺发育早中期即已具备胰岛素合成及分泌的能力,但新生鼠刚出生时,其胰岛功能仍尚未完善,出生以后才逐渐对葡萄糖负荷有良好的反应性。
OBJECTIVE: To determine the expression of insulin-related genes in different stages of rat pancreatic development and to explore the process of improving insulin secretion. Methods: SD rat embryonic development was isolated and extracted for 12.5 days (E12.5), E15.5, E18.5, newborn, 21 days after birth and adult rat pancreatic tissue. Affymetrix high-density oligonucleotide microarray The gene expression profiles of pancreatic tissues at various stages of development were detected and the specific and differential expressions of insulin-related genes were analyzed. The levels of glucose in normal neonatal rats and adult rats and the IPGTT in neonatal rats on days 1, 3, 7, 10, and 12 were measured respectively. Serum insulin concentrations in neonatal and adult rats were measured by ELISA. Primary Culture of Islet Cells in Newborn Rats to Observe the Morphology of Islet Cells in Newborn Rats. Results: Insulin gene and its specific transcription factors such as PDX-1 began to express in E12.5-E15.5 rats during the process of insulin synthesis and release. Insulin gene persisted and increased after insulin release. Insulin cell Syntaxin-1a, Vamp-2, Rab-3a and Munc13-1, the key genes in vomiting, began to appear from E15.5, and their expression levels increased thereafter. In glucose-stimulated insulin secretion (GSIS) related genes, glucose transport Body-2 (Glut2) begins to express early in the embryo, and glucose kinase (GCK) is expressed late in the embryo. From E18.5 that is detected in the blood of rats the presence of insulin, with the embryonic development, blood insulin levels gradually increased. Neonatal rats were unable to tolerate glucose overload and had abnormal IPGTT. IPGTT tended to be normal after one week of birth. The morphology of islets in neonatal rats is similar to that in adult mice, but the volume is relatively small and the translucency is slightly worse. CONCLUSION: The ability of insulin synthesis and secretion in the early and mid-stage of pancreas development in rat embryo is already present. However, the function of pancreatic islets in newborn rats is still not perfect after birth, and it has good response to glucose load after birth.