中 文 摘 要

骨髓來源的樹突狀細(xì)胞(dendritic cells DC) 是已知的體內(nèi)抗原提呈功能最強的抗原提呈細(xì)胞( antigenpresenting cell，APC ) ，能攝取各類抗原, 體內(nèi)、外均能激發(fā)T 細(xì)胞增殖, 誘導(dǎo)特異性細(xì)胞殺傷性T 淋巴細(xì)胞( cy to to xic T lymphocy tes, CT L) 生成, 產(chǎn)生抗腫瘤效應(yīng)。近些年來越來越多的人認(rèn)識到DC 在腫瘤免疫中的作用, DC 的研究也已經(jīng)進(jìn)入了體內(nèi)研究階段。但既往體內(nèi)試驗主要是以裸鼠為動物模型，由于裸鼠為細(xì)胞免疫缺陷性動物, 很難反應(yīng)出卵巢癌DC 疫苗的真正免疫調(diào)節(jié)作用，因此有必要對正常免疫狀態(tài)的實驗動物進(jìn)行DC 培養(yǎng)。大鼠是體內(nèi)實驗最常用的動物模型, 而目前國內(nèi)外對大鼠來源DC 培養(yǎng)方法有限。本實驗利用大鼠骨髓細(xì)胞進(jìn)行DC 的分離培養(yǎng), 進(jìn)一步探討了大鼠骨髓DC 的培養(yǎng)方法, 為DC 體內(nèi)研究奠定基礎(chǔ)。

目的：建立體外培養(yǎng)、誘導(dǎo)和擴(kuò)增大鼠骨髓來源的樹突狀細(xì)胞的方法。

方法：實驗通過運用大鼠淋巴細(xì)胞分離液和培養(yǎng)過程的手法篩選相結(jié)合,培養(yǎng)、誘導(dǎo)和擴(kuò)增大鼠骨髓來源的樹突狀細(xì)胞,通過形態(tài)學(xué)和免疫表型進(jìn)行鑒定。

結(jié)果： 成功建立了體外大量培養(yǎng)、誘導(dǎo)及擴(kuò)增大鼠骨髓來源樹突狀細(xì)胞的方法;經(jīng)倒置相差顯微鏡、透射電鏡及流式細(xì)胞儀鑒定,證實所培養(yǎng)細(xì)胞為樹突狀細(xì)胞。 

結(jié)論：通過運用大鼠淋巴細(xì)胞分離液和培養(yǎng)過程的手法篩選相結(jié)合能培養(yǎng)、誘導(dǎo)、擴(kuò)增大量的樹突狀細(xì)胞。

關(guān)鍵詞: 樹突狀細(xì)胞; 免疫耐受; 大鼠

Astract 

Bone marrow derived dendritic cells (dendritic cells DC) is known in vivo antigen presenting function is the strongest antigen-presenting cells(antigenpresenting, cell, APC), intake of various antigens, body, can stimulate the proliferation of T cells, induce specific cytotoxic T lymphocyte (CY to to Xic T Lymphocy tes, CTL) generation, producing antitumor effect. In recent years, more and more people realize that the role of DC in tumor immunity, DC research has entered a phase of the study in vivo. But previous in vivo test is mainly to nude mice as animal model, the nude mice for cell immunodeficiency animal, it is difficult to reflect the real immune regulation effect of ovarian cancer DC vaccine, it is necessary to experimental animal of normal immune status were cultured in DC system. The rat is the most commonly used experimental animal models, while at home and abroad on the rat DC culture co.. Isolation and culture of rat bone marrow cells in this experiment using DC, to further explore the cultivation of rat bone marrow DC, lay the foundation for DC in vivo.

Objective: to establish a method of induction and proliferation of rat bone marrow-derived dendritic cells, in vitro culture.

Methods: the experiment by using the separation of rat lymphocytes and screening combined training techniques, training,induction and proliferation of rat bone marrow-derived dendritic cells, were identified by morphology and immunophenotype.

Results: successfully established the method of in vitro, induction and proliferation of rat bone marrow-derived dendritic cells; by the inverted microscope, transmission electron microscope and flow cytometry, confirmed that the cultured cells into dendritic cells.

Conclusion: through the use of liquid separation of rat lymphocytes and training process is manual screening combining culture, induction, amplification of dendritic cells.

Keywords: dendritic cells; immune tolerance; rats

目  錄

中 文 摘 要 I

Astract II

目  錄 III

第一章  前  言 1

第二章  實驗部分 3

2.1  實驗動物 3

2.2  主要儀器和試劑 3

2.3試劑配制 4

2.4  實驗方法 5

第三章  實驗結(jié)果分析及討論 7

3.1 骨髓樹突狀細(xì)胞的形態(tài)觀察 7

3.2 討論 7

第四章  結(jié)論 10

附  圖 11

參考文獻(xiàn) 12

致  謝 13

外文文獻(xiàn) 14

文獻(xiàn)翻譯 17