论文摘要
【目的】1.探讨宫内发育迟缓（Intrauterine Growth Retardation, IUGR）大鼠在胚胎期和出生后Insulin/Fox01/Pdx1/MafA基因的表达。2.探讨宫内发育迟缓大鼠在胚胎期和出生后microRNA（miRNA）的表达。【方法】实验一：（1）妊娠全程食物限制法制备宫内发育迟缓大鼠模型：清洁级SD大鼠，雌雄鼠1:3合笼交配，孕鼠按受孕顺序随机分为两组（IUGR组和对照组）。IUGR组自妊娠第1天至分娩给予对照组饲料进食量的半量；对照组妊娠全程任意摄取饲料；（2）标本的采集：应用显微分离技术提取大鼠胚胎发育14.5天（E14.5），19.5天（E19.5）及新生大鼠胰腺组织；（3）胰腺组织形态学的观察：采用HE染色，光镜观察E14.5、E19.5及新生大鼠胰腺形态学变化；（4）大鼠胰腺发育相关基因表达的检测：使用RT-PCR技术检测胚胎发育E14.5、E19.5及新生大鼠胰腺Insulin/Fox01/Pdx1/MafA基因的表达情况；（5）不同时期大鼠胰腺Pdx1、MafA蛋白表达的检测：采用Western-blot方法检测胚胎发育E14.5、E19.5及新生大鼠胰腺Pdx1、MafA的蛋白表达情况。实验二：（1）妊娠全程食物限制法制备宫内发育迟缓大鼠模型：同实验一；（2）标本的采集：应用显微分离技术提取大鼠胚胎发育14.5天（E14.5），19.5天（E19.5）及新生大鼠胰腺组织并成功提取RNA；（3）miRNA的检测：采用丹麦Exiqon公司miRCURY. LNA miRNA表达谱芯片，对各样本RNA进行miRNA表达水平的检测；（4）芯片检测结果的验证：用实时定量PCR方法对选取的miRNA进行验证。【结果】实验一：（1）与对照组大鼠相比，IUGR组在胰腺发育E14.5、E19.5及新生时均有形态学上的改变，表现为组织松散，结构不清晰，胰岛数量及面积减少；（2）Insulin/Fox01/Pdx1/MafA基因全程均有表达；（3）Insulin、MafA基因随胎龄的增长表达量增多，与对照组相比，IUGR组Insulin基因表达无明显差异（P>0.05），而MafA基因表达较之减少（P<0.05）；（4）Fox01、Pdx1基因均在胚胎早期表达较多，随胎龄的增长表达量下降，且与对照组相比，IUGR组Pdx1基因表达无明显差异（P>0.05），Fox01表达较对照组减少（P<0.05）；（5）MafA蛋白随胎龄的增长表达量增多，且IUGR组表达明显低于对照组（P<0.05）；Pdx1蛋白随胎龄的增长表达量减少，IUGR组与对照组表达无明显差异（P>0.05）。实验二：（1）宫内发育迟缓大鼠在胚胎期和新生后miRNA的表达：对照组中，E19.5与E14.5相比，上调的miRNA有48个，下调的28个；新生鼠与E14.5相比，上调的64个，下调55个；新生鼠与E19.5相比，上调40个，下调52个。同时，与对照组相比，IUGR组中E14.5上调的miRNA有10个，下调的有26个；E19.5上调22个，下调15个；新生鼠上调9个，下调13个；（2）对选取的miR-124、let-7d*验证：对照组中，miR-124在新生鼠均比E14.5、E19.5显著上调（FoldChange>=1.5），而IUGR组较对照组表达无明显差异(0.67<FoldChange
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