Exo utero gene introduction to specific organs and tissues during fetal and neonatal period

子宫外发育系统和胚胎显微注射技术不断完善。为了提高外源基因注入小鼠胚胎器官的效率，已建立了基因载体和电穿孔的最佳条件，并已估计反义寡核苷酸的有用性。为了证明子宫外系统的潜力，将几种生物活性物质注射到特定的胎儿器官中，并分析它们的作用。建立了分析几种器官组织发生的子宫外实验系统.子宫外系统的改进：减少每一次操作时间，改进子宫壁切口，麻醉和复苏，显著提高了操作后的存活率和显微注射的效率.基因导入条件的优化：在几种脂质体中， 关于我们 测定不同条件下报告基因的掺入。评价了用BTX系统进行电穿孔的条件，并且平板型电极显示出优异的操作性。已经建立了注射溶液良好扩散到脑实质中的条件，对组织的损伤最小。反义寡核苷酸的导入：将Tag1和Mab21的反义寡核苷酸以单剂量或双剂量间隔注射到胎鼠脑室，蛋白表达水平显著降低.子宫外系统在器官组织发生调控分子功能分析中的应用：将LIF注入脑室，对大脑皮层表现出特异性作用，包括增加特定皮层层的神经母细胞和纤维。ACTH对胎儿肾上腺的影响也被已建立的子宫外实验系统所证实。建立了分析脑积水和无脑畸形发病机制及内耳和关节发育的子宫外实验系统。少

Exo utero development system and techniques of microinjection into fetuses have been improved. To increase efficiency of the uptake of exogenous genes injected into mouse embyonic organs, optimal conditions of gene carriers and electroporation have been established, and usefulness of antisense oligonucleotides has been estimated. To demonstrate the potential of exo utero system, several biologically active substances were injected into the specific fetal organs, and their effects were analyzed. The exo utero experimental systems to analyze the histogenesis of several organs have been established.1. Improvement of exo utero system : Reduction of manipulation time for each procedure, modification of incision of the uterine wall, anesthesia and resuscitation, all together have significantly improved the survival rate after the manipulation and efficiency of microinjection.2. Optimization of conditions for gene introduction : Among several liposomes, the ones with the highest efficiency of … More reporter gene incorporation under different conditions were determined. Conditions of electroporation with the BTX system were evaluated and the flat type electrode showed an excellent handling. Conditions have been established for good diffusion of injected solution into the cerebral parenchyme with minimum damage to the tissue.3. Introduction of antisense oligonucleotides : Antisense oligonucleotides for Tag1 and Mab21 was injected into the fetal cerebral ventricle at single-dose or double-dose with an interval, and the protein expression levels decreased significantly.4. Application of exo utero system into the functional analysis of regulatory molecules in the histogenesis of the organs : LIF was injected into the cerebral ventricle and showed specific effects on the cerebral cortex including increase in the neuroblasts and fibers in the specific cortical layrs. The effects of ACTH on the fetal adrenal were similarly evidenced by the established exo utero experimental system. The exo utero experimental systems for analysis of pathogenesis of hydrocephaly and excencephaly, and development of the inner ear and joints have also been established. Less