小鼠原始生殖细胞在同种正常肝脏中的定向迁移和分化

发布时间：2018-01-07 03:39

本文关键词：小鼠原始生殖细胞在同种正常肝脏中的定向迁移和分化　出处：《青岛大学》2015年硕士论文　论文类型：学位论文

【摘要】：目的:移植的干细胞在正常体内是如何迁移和分化的研究报道甚少。本研究动态观察来源于小鼠原始生殖腺嵴的胚胎干细胞在正常肝脏微环境中的迁移、分化、及对正常肝组织的影响。对胚胎干细胞移植的安全性进行评价。方法:无菌分离小鼠原始生殖腺嵴,提取胚胎干细胞,体外扩增后应用Brd U标记干细胞,然后经尾静脉注入正常小鼠体内。于注射后2周、4周、6周、8周取材肝脏,制备连续冰冻切片,应用苏木精-伊红(HE)染色、免疫荧光双重染色和过碘酸雪夫氏(PAS)染色方法检测移植的干细胞在受体肝组织内的分布、干细胞特异性标志物白蛋白表达和糖原合成;显微计数分析肝细胞的增殖状态。实验结果:干细胞移植2周时,可见受体小鼠肝组织内有散在分布的大而圆的Brd U和白蛋白双阳性细胞,PAS染色呈强阳性反应;体积小的Brd U阳性细胞,白蛋白表达呈阴性。HE染色可见肝样细胞体积和细胞核变大、染色质松散、核膜不清晰;单位面积内肝细胞的数量较正常肝组织减少。移植4周时小鼠肝组织单位面积内Brd U阳性细胞的数量较2周时减少,糖原颗粒PAS着色也较2周浅,肝样细胞体积和细胞核仍较大、染色质松散,单位面积内肝细胞的数量较移植2周时增多。移植6周、8周Brd U阳性细胞的数量和染色强度明显下降,肝细胞的体积和胞核的形态趋于正常。移植8周约4/7的小鼠肝组织内Brd U阳性肝样细胞呈阴性;其中1只小鼠肝组织内可见少量Brd U阳性细胞聚集。实验结论:静脉移植胚胎干细胞可以迁移至正常肝脏,分化为肝细胞,并刺激肝细胞增殖;移植8周时肝组织结构恢复正常。
[Abstract]:Objective: there are few reports on the migration and differentiation of transplanted stem cells in normal vivo. This study dynamically observed the migration and differentiation of embryonic stem cells derived from the primordial gonadal ridge of mice in normal liver microenvironment. Methods: embryonic stem cells were isolated from mouse primordial gonadal cristae and expanded in vitro with Brd U labeled stem cells. Then the normal mice were injected into the caudal vein. Liver samples were obtained from the liver at 4 weeks, 6 weeks and 8 weeks after injection, and continuous frozen sections were prepared and stained with hematoxylin-eosin (HEH). The distribution of transplanted stem cells in recipient liver tissues, the expression of albumin and glycogen synthesis were detected by immunofluorescence double staining and Shevger's periodinate PAS-staining. Results: after 2 weeks of stem cell transplantation, there were large and round Brd U and albumin double positive cells scattered in the liver tissue of recipient mice. PAS staining showed strong positive reaction. In small Brd U positive cells, the expression of albumin was negative. He staining showed that the size and nucleus of hepatocytes became larger, the chromatin was loose and the nuclear membrane was not clear. The number of Brd U positive cells in the unit area of liver tissue of mice at 4 weeks after transplantation was lower than that in normal liver tissue, and the PAS staining of glycogen granules was lighter than that in 2 weeks. The size and nucleus of hepatoid cells were still large, chromatin was loose, the number of hepatocytes per unit area was higher than that at 2 weeks after transplantation, and the number and staining intensity of Brd U positive cells decreased significantly at 6 weeks and 8 weeks after transplantation. The volume of hepatocytes and the morphology of nuclei tended to be normal. The Brd U-positive hepatocytes in the liver tissues of about 4/7 mice were negative at 8 weeks after transplantation. A small amount of Brd U positive cells could be found in the liver of one mouse. Conclusion: vein transplantation of embryonic stem cells can migrate to normal liver, differentiate into hepatocytes, and stimulate the proliferation of hepatocytes. The liver structure returned to normal at 8 weeks after transplantation.
【学位授予单位】：青岛大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R457.7

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