本文選題：樹突狀細(xì)胞 + 體外培養(yǎng)　； 參考：《第一軍醫(yī)大學(xué)》2005年碩士論文

【摘要】：目的: 建立一種簡便實(shí)用的體外培養(yǎng)未成熟樹突狀細(xì)胞的方法,并鑒定和研究其形態(tài)、表型以及生物免疫學(xué)功能。觀察不同成熟狀態(tài)的樹突狀細(xì)胞對T淋巴細(xì)胞功能的影響,探討未成熟樹突狀細(xì)胞在誘導(dǎo)機(jī)體產(chǎn)生免疫耐受中的作用,為樹突狀細(xì)胞在降低移植免疫排斥反應(yīng)和治療某些自身免疫性疾病的臨床應(yīng)用提供一些理論和實(shí)驗(yàn)依據(jù)。 方法:1.小鼠骨髓源樹突狀細(xì)胞的培養(yǎng):取小鼠的骨髓細(xì)胞作為樹突狀細(xì)胞的前體細(xì)胞,利用細(xì)胞因子rmGM-CSF和IL-4聯(lián)合刺激,手法篩選樹突狀細(xì)胞集落,培養(yǎng)6天后收集疏松貼壁細(xì)胞,即為小鼠骨髓來源的樹突狀細(xì)胞。將培養(yǎng)的樹突狀細(xì)胞分成兩組,一組在培養(yǎng)結(jié)束前18h加入LPS刺激其成熟(以下稱成熟DC組);一組不加入LPS刺激,使其保留未成熟樹突狀從不特性(以下稱未成熟DC組)。 2.樹突狀細(xì)胞的形態(tài)學(xué)觀察:在光鏡、電鏡下觀察兩組細(xì)胞的形態(tài),比較成熟組樹突狀細(xì)胞與未成熟組樹突狀細(xì)胞形態(tài)學(xué)上的差別。 3.樹突狀細(xì)胞的表型測定:利用流式細(xì)胞術(shù)檢測兩組細(xì)胞表面標(biāo)志,檢測小鼠樹突狀細(xì)胞的特異性標(biāo)志CD11c評估樹突狀細(xì)胞純度;檢測MHC-Ⅱ類分子以及共刺激分子(CD40,CD86等)評價樹突狀細(xì)胞成熟度。 4.樹突狀細(xì)胞分泌IL-12的水平測定:收集兩組樹突狀細(xì)胞培養(yǎng)上清,用ELISA法檢測上清中IL-12的表達(dá)水平,了解樹突狀細(xì)胞分泌IL-12的水平與其成
[Abstract]:Objective:
To establish a simple and practical method for the cultivation of immature dendritic cells in vitro, and to identify and study its morphological, phenotypic and Bioimmunological functions. To observe the effect of dendritic cells in different mature states on the function of T lymphocytes, and to explore the role of immature dendritic cells in inducing immune tolerance in the body, which is a dendritic shape. The cells provide some theoretical and experimental evidence for reducing the transplant rejection and treating some autoimmune diseases.
Methods: 1. mouse bone marrow derived dendritic cells were cultured: the mouse bone marrow cells were taken as the precursor cells of the dendritic cells. The dendritic cells were selected by the combination of cytokine rmGM-CSF and IL-4 to screen the dendritic cells and collect the loose adherent cells for 6 days, that is, the dendritic cells from the bone marrow of the mice. Two groups were divided into two groups. One group was added to LPS to stimulate maturity before the end of the culture (hereinafter referred to as the mature DC group); a group did not add LPS stimulation to retain the unmature dendritic uncharacteristic (hereinafter referred to as the immature DC group).
2. morphological observation of dendritic cells: the morphology of two groups of cells was observed under light microscope and electron microscope, and the morphological difference between dendritic cells from mature group and immature group was compared.
3. the phenotypic determination of dendritic cells: detection of two groups of cell surface markers by flow cytometry, detection of specific markers of dendritic cells in mice by CD11c, to evaluate the purity of dendritic cells, and to evaluate the maturity of dendritic cells by detecting MHC- class II molecules and co stimulators (CD40, CD86, etc.).
4. the level of IL-12 secreted by dendritic cells: collecting two groups of dendritic cells culture supernatant, using ELISA to detect the expression level of IL-12 in the supernatant, to understand the level of IL-12 in the dendritic cells and its formation.
【學(xué)位授予單位】：第一軍醫(yī)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2005
【分類號】：R392

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本文編號：2101134