本文選題：神經(jīng)干細(xì)胞　切入點：豬　出處：《東北農(nóng)業(yè)大學(xué)》2011年碩士論文

【摘要】：神經(jīng)干細(xì)胞用于神經(jīng)學(xué)臨床修復(fù)和基礎(chǔ)理論研究的前提是首先完成神經(jīng)干細(xì)胞的體外分離、培養(yǎng)、純化并大量擴(kuò)增。鼠、人中都已成功分離出神經(jīng)干細(xì)胞并已嘗試用于動物神經(jīng)系統(tǒng)損傷等疾病的治療,盡管在鼠和人上的研究很多,相對于鼠神經(jīng)干細(xì)胞在神經(jīng)學(xué)臨床應(yīng)用上的局限和人神經(jīng)干細(xì)胞在材料來源上的不便,豬作為神經(jīng)干細(xì)胞臨床應(yīng)用和基礎(chǔ)研究的模式動物有很大的潛力。成體神經(jīng)干細(xì)胞在細(xì)胞替代療法的研究和應(yīng)用中具有廣闊的前景。成體神經(jīng)干細(xì)胞移植不僅可以避免應(yīng)用胚胎神經(jīng)干細(xì)胞進(jìn)行細(xì)胞替代治療所帶來的倫理問題,而且還可解決由異體或異種移植而導(dǎo)致的免疫排斥問題,使臨床應(yīng)用成為可能。目前,豬成體神經(jīng)干細(xì)胞系尚未被建立起來。 本研究旨在探索如何從成豬腦組織中分離培養(yǎng)神經(jīng)干細(xì)胞,觀察神經(jīng)干細(xì)胞生長特性和體外增殖的特點。主要的實驗方法和實驗結(jié)果如下: 第一部分:選取適用于人或/和鼠的4種培養(yǎng)體系篩選適用于豬的成體神經(jīng)干細(xì)胞的培養(yǎng)條件。結(jié)果表明無血清的懸浮或貼壁培養(yǎng)條件不適用于原代分離豬成體神經(jīng)干細(xì)胞,只有完全培養(yǎng)液中含有血清的DMEM/F12+10% FBS+20ng/ml bFGF+20ng/ml hEGF+103U/ml LIF培養(yǎng)體系支持豬成體神經(jīng)干細(xì)胞的存活和增殖。 第二部分:選取3月齡成豬腦室下區(qū)(SVZ)、海馬齒狀回顆粒細(xì)胞下區(qū)(SGZ)、嘴側(cè)遷移通路(RMS)、海馬角(HH)4個腦區(qū)分離培養(yǎng)成體神經(jīng)干細(xì)胞,檢測可以分離出豬成體神經(jīng)干細(xì)胞的部位。結(jié)果表明4個區(qū)域均含有成體神經(jīng)干細(xì)胞,可進(jìn)行原代分離,但無法繼代培養(yǎng)。 第三部分:探討豬齡對神經(jīng)干細(xì)胞培養(yǎng)的影響。結(jié)果顯示3月齡豬SVZ來源的成體神經(jīng)干細(xì)胞在P3呈現(xiàn)的是完全分化的貼壁形態(tài),而1日齡新生豬SVZ來源的成體神經(jīng)干細(xì)胞在P3呈現(xiàn)的是懸浮生長的神經(jīng)球形態(tài),說明1日齡新生豬SVZ比3月齡成豬SVZ獲得P3神經(jīng)球的效率高。
[Abstract]:The premise of neural stem cells for clinical nerve repair and basic theoretical research is to isolate, culture, purify and amplify neural stem cells in vitro.Neural stem cells have been successfully isolated from both rats and humans and have been tried to treat diseases such as neurological damage in animals, despite much research on mice and humans.Compared with the limitation of clinical application of neural stem cells in rats and the inconvenience of human neural stem cells in material source, pigs have great potential as model animals for clinical application and basic research of neural stem cells.Adult neural stem cells have broad prospects in the research and application of cell replacement therapy.Adult neural stem cell transplantation can not only avoid the ethical problems caused by the application of embryonic neural stem cells in cell replacement therapy, but also solve the problem of immune rejection caused by allogeneic or xenotransplantation, which makes clinical application possible.At present, porcine adult neural stem cell lines have not been established.The purpose of this study was to investigate how to isolate and culture neural stem cells from adult porcine brain tissue and observe the characteristics of neural stem cell growth and proliferation in vitro.The main experimental methods and results are as follows:Part one: four culture systems suitable for human and / or mouse were selected to screen the culture conditions of adult neural stem cells suitable for pigs.The results showed that serum-free suspension or adherent culture was not suitable for primary isolation of porcine adult neural stem cells. Only DMEM/F12 10% FBS 20ng/ml bFGF 20ng/ml 20ng/ml 103U/ml LIF containing serum could support the survival and proliferation of porcine adult neural stem cells.The second part: the adult neural stem cells were isolated and cultured in the three month old adult pig subventricular area (SVZ), the subgranular cells of the dentate gyrus (SGZ), the lateral migration pathway (RMSA) and the hippocampal horn (HH), and the location of the porcine adult neural stem cells (NSCs) could be detected.The results showed that all the four regions contained adult neural stem cells, which could be isolated in primary culture, but could not be subcultured.Part three: to investigate the effect of pig age on neural stem cell culture.The results showed that 1 day old newborn pig SVZ was more efficient than 3 month old adult pig SVZ in obtaining P3 neurospheres.
【學(xué)位授予單位】：東北農(nóng)業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2011
【分類號】：R329

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