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研究生:羅典妮
研究生(外文):TIEN-NI LO
論文名稱:大白鼠寡樹突神經膠細胞CD200的特性
論文名稱(外文):Characterization of oligodendrocyte CD200 in rats
指導教授:吳慶祥
指導教授(外文):CHING-HSIANG WU
學位類別:碩士
校院名稱:國防醫學院
系所名稱:生物及解剖學研究所
學門:生命科學學門
學類:生物訊息學類
論文種類:學術論文
論文出版年:2007
畢業學年度:95
語文別:中文
論文頁數:78
中文關鍵詞:寡樹突神經膠細胞
外文關鍵詞:oligodendrocyte
相關次數:
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摘要

在中樞神經的神經膠細胞有星狀神經膠細胞、寡樹突神經膠細胞、小神經膠細胞及室管膜細胞。寡樹突神經膠細胞,可以藉由細長的細胞突起將軸突包覆形成髓鞘,形成髓鞘時需要免疫球蛋白超級家族的細胞黏著分子的參與。CD200也是免疫球蛋白超級家族的一員,具有二個免疫球蛋白功能區,可以與CD200R接合媒介細胞與細胞間的交互作用。有文獻指出CD200分佈在小腦的某些神經元上,包含其細胞本體及軸突,並且與軸突的生成及髓鞘的形成有關。這樣的證據引導我們假設CD200可表現在寡樹突神經膠細胞上,並且在軸突的生長及髓鞘化的過程中扮演重要的角色。因為要證實這樣的關連,所以我的實驗第一步是要證實CD200確實存在寡樹突神經膠細胞上。結果發現在視神經及初級神經膠細胞培養,都可以看在神經纖維上以及類似寡樹突神經膠細胞有CD200的免疫反應物,並且呈現斑塊狀分佈。為了更加確定寡樹突神經膠細胞確實表現CD200,我們使用雙重免疫螢光染色,標示CD200及RIP(寡樹突神經膠細胞表面上的特殊抗原)以共軛焦螢光顯微鏡做觀察。並以電子顯微鏡觀察培養中的寡樹突神經膠細胞,更加確認寡樹突神經膠細胞可以表現CD200。將含有寡樹突神經膠細胞以及具CD200R的小神經膠細胞之混合神經膠細胞培養中,加入抗CD200抗體、抗CD200R抗體,來了解CD200在寡樹突神經膠細胞上所扮演的角色。我們實驗結果發現加入抗CD200抗體會引發寡樹突神經膠細胞死亡,並且死亡情形有抗體濃度的差別;另外發現加入抗CD200R抗體,會使小神經膠細胞的形態改變,貼附較為平坦。神經球內含有神經前驅細胞並且大部份可以分化成寡樹突神經膠細胞,在加入抗CD200抗體處理之後,發現神經球的貼附及展開形態會被抑制;並且也會干擾寡樹突神經膠細胞從神經球內遷移的情形。實驗結果證實CD200黏著分子確實表現在寡樹突神經膠細胞的胞膜上並且加入抗CD200抗體之後會影響寡樹突神經膠細胞的存活,並且有抗體濃度的差別。另外發現在混合神經膠細胞培養中加入抗CD200R抗體,會使小神經膠細胞的貼附情況較好。神經球經抗CD200抗體處理之後,對於神經球的貼附及寡樹突神經膠細胞遷移有抑制的效果。
Abstract

Glial cells in the central nervous system include astrocytes, oligodendrocytes, microglia and ependymal cells. Oligodendrocytes which we focus on in the present study show few long and thin processes investing the axons and forming the myelin sheath through kinds of adhesion molecules, in particular the immunoglobulin superfamily. Also being a member of immunoglobulin superfamily, CD200 contains two immunoglobulin domains and mediates cell-cell interaction by binding with CD200 receptor. The previous researches pointed out that CD200 was distributed at some neurons, including neuronal cell bodies and axon processes in the cerebellum, and related to the axonogenesis and myelination. These evidences lead us to assume that CD200 may exist on oligodendrocytes and play the importance role in events relating to axonal growth and myelination. In this connection, the purposes of the present study are to examine CD200 expression on oligodendrocytes. Our results showed that CD200 immunoreactivity was distributed along the nerve fibers in a patchy pattern and at oligodendrocyte-like cells in the optic nerves and primary glial culture. The labeled cells were confirmed as an oligodendrocyte using the double labeling of CD200 and RIP (oligodendrocyte marker) and laser confocal microscope. Examinations with the electron microscope and enriched oligodendrocyte culture also verified the existence of oligodendrocyte CD200. To know the role of oligodendrocyte CD200, co-cultures of oligodendrocytes and microglia that possessing CD200 receptors (CD200R) and the treatment of anti-CD200 or anti-CD200R antibody in these mixed glial cultures were applied. We found that oligodendrocytes were died after treatment of anti-CD200 antibody in dose-dependent manner, while microglia spread out more widely when anti-CD200R antibody was added. In the culture model of the neurosphere that oligodendrocytes constituted most of the neural precursor cells progeny, we found that the spreading and adhesion of the neurospheres were inhibited with the treatment of anti-CD200 antibody that also interfered the migration of differentiating oligodendrocytes from neurospheres. Our results showed that CD200 immunoreactivity was distributed at cytoplasmic membrane of oligodendrocytes and the loops at paranodular region. Unexpectedly, oligodendrocytes were degenerated when incubated with anti-CD200 antibody in dose-dependent manner, but microglia spread out more widely as anti-CD200R antibody was added. Furthermore, in the culture model of the neurosphere that oligodendrocytes constituted most of the neural precursor cells progeny, we found that the spreading and adhesion of the neurospheres were inhibited with the treatment of anti-CD200 antibody that also interfered the migration of differentiating oligodendrocytes from neurospheres.
目錄
頁次
目錄 -------------------------------------------------I
圖目錄 -----------------------------------------------III
縮寫表 -----------------------------------------------V
中文摘要 ---------------------------------------------VI
英文摘要 ---------------------------------------------VIII
第一章 緒言 -----------------------------------------1
第一節 寡樹突神經膠細胞 -----------------------------1
第二節 細胞黏著分子 -----------------------------------6
第三節 CD200與CD200R ---------------------------------8
第四節 小神經膠細胞 -----------------------------------11
第五節 神經球 -----------------------------------------12
第二章 實驗目的 ---------------------------------------13
第三章 材料與方法 ---------------------------------------14
第一節 實驗材料 --------------------------------------14
第二節 實驗方法 --------------------------------------17
第四章 實驗結果 ----------------------------------------25
第一節 視神經寡樹突神經膠細胞CD200的表現 ------------------25
第二節 寡樹突神經膠細胞對小神經膠細胞形態之影響 -------------28
第三節 抗CD200抗體對寡樹突神經膠細胞形態及存活率之影響
---------------------------------------------------------30
第四節 抗CD200抗體對於神經球貼附及其內寡樹突神經膠細胞向外遷移之影響
--------------------------------------------------32
第五章 討論 ---------------------------------------------34
第六章 結論 ---------------------------------------------43
第七章 參考文獻 ------------------------------------------74
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