本文關(guān)鍵詞：小鼠原始生殖細(xì)胞在同種正常肝臟中的定向遷移和分化　出處：《青島大學(xué)》2015年碩士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 胚胎干細(xì)胞 細(xì)胞移植 肝臟 小鼠

【摘要】：目的:移植的干細(xì)胞在正常體內(nèi)是如何遷移和分化的研究報(bào)道甚少。本研究動(dòng)態(tài)觀察來(lái)源于小鼠原始生殖腺嵴的胚胎干細(xì)胞在正常肝臟微環(huán)境中的遷移、分化、及對(duì)正常肝組織的影響。對(duì)胚胎干細(xì)胞移植的安全性進(jìn)行評(píng)價(jià)。方法:無(wú)菌分離小鼠原始生殖腺嵴,提取胚胎干細(xì)胞,體外擴(kuò)增后應(yīng)用Brd U標(biāo)記干細(xì)胞,然后經(jīng)尾靜脈注入正常小鼠體內(nèi)。于注射后2周、4周、6周、8周取材肝臟,制備連續(xù)冰凍切片,應(yīng)用蘇木精-伊紅(HE)染色、免疫熒光雙重染色和過(guò)碘酸雪夫氏(PAS)染色方法檢測(cè)移植的干細(xì)胞在受體肝組織內(nèi)的分布、干細(xì)胞特異性標(biāo)志物白蛋白表達(dá)和糖原合成;顯微計(jì)數(shù)分析肝細(xì)胞的增殖狀態(tài)。實(shí)驗(yàn)結(jié)果:干細(xì)胞移植2周時(shí),可見(jiàn)受體小鼠肝組織內(nèi)有散在分布的大而圓的Brd U和白蛋白雙陽(yáng)性細(xì)胞,PAS染色呈強(qiáng)陽(yáng)性反應(yīng);體積小的Brd U陽(yáng)性細(xì)胞,白蛋白表達(dá)呈陰性。HE染色可見(jiàn)肝樣細(xì)胞體積和細(xì)胞核變大、染色質(zhì)松散、核膜不清晰;單位面積內(nèi)肝細(xì)胞的數(shù)量較正常肝組織減少。移植4周時(shí)小鼠肝組織單位面積內(nèi)Brd U陽(yáng)性細(xì)胞的數(shù)量較2周時(shí)減少,糖原顆粒PAS著色也較2周淺,肝樣細(xì)胞體積和細(xì)胞核仍較大、染色質(zhì)松散,單位面積內(nèi)肝細(xì)胞的數(shù)量較移植2周時(shí)增多。移植6周、8周Brd U陽(yáng)性細(xì)胞的數(shù)量和染色強(qiáng)度明顯下降,肝細(xì)胞的體積和胞核的形態(tài)趨于正常。移植8周約4/7的小鼠肝組織內(nèi)Brd U陽(yáng)性肝樣細(xì)胞呈陰性;其中1只小鼠肝組織內(nèi)可見(jiàn)少量Brd U陽(yáng)性細(xì)胞聚集。實(shí)驗(yàn)結(jié)論:靜脈移植胚胎干細(xì)胞可以遷移至正常肝臟,分化為肝細(xì)胞,并刺激肝細(xì)胞增殖;移植8周時(shí)肝組織結(jié)構(gòu)恢復(fù)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.
【學(xué)位授予單位】：青島大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：R457.7

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