本文選題：多發(fā)性骨髓瘤 + 樹突細胞�。� 參考：《昆明醫(yī)科大學》2016年碩士論文

【摘要】：目的：通過體外殺傷實驗,研究轉染粘蛋白核心肽MUC1-2VNTR(兩串聯(lián))基因的樹突細胞(DC)誘導的細胞毒T細胞(CTL)對人的多發(fā)性骨髓瘤細胞株(U266)的誘導殺傷效應,為研制多發(fā)生性骨髓瘤基因疫苗奠定基礎。方法：1、根據靶基因設計,將MUC1-2VNTR的編碼基因作為目的基因,將人工合成的全基因克隆入載體pcDNA3.1/myc-hisA中。2、用去內毒素質粒小提試劑盒擴增、提取質粒、并保存。3、選取健康人的外周血單個核細胞,體外由重組人粒細胞-巨噬細胞集落刺激因子(rhGM-CSF)和重組人白細胞介素4(rhIL-4)誘導成熟的DC細胞。4、構建pcDNA3.1-2VNTR重組質粒后,采用Lipofectimine脂質體介導法轉染DC細胞(DC-2VNTR),以轉染空載體質粒pcDNA3.1 (DC-pcDNA3.1)及l(fā)ipofctamine處理的DC (DC-Lipo)作為對照組。5、Western印跡法檢測重組體在DC細胞內的表達。6、在體外刺激同源T細胞,細胞毒實驗檢測DC-2VNTR誘導的CTL對多發(fā)性骨髓瘤細胞株(U266)的殺傷作用。結果：1.成功構建了重組質粒載體pcDNA3.1-2VNTR/myc-hisA;2.人的外周血DC細胞的培養(yǎng)和鑒定；3. Western印跡法檢測到MUC1-2VNTR在DC細胞內的表達；4.DC-2VNTR誘導的CTL對多發(fā)性骨髓瘤細胞(U266)有顯著殺傷效應,且顯著高于DC-pcDNA3.1組及DC-Lipo組(P0.05)。結論：以MUC1-2VNTR基因轉染的DC可以誘導CTL反應,并能特異地殺傷U266多發(fā)性骨髓瘤細胞株。
[Abstract]:Objective: to study the cytotoxic T cells (CTL) induced by dendritic cells (DC) transfected with MUC1-2VNTR gene in vitro on human multiple myeloma cell line (U266). To lay a foundation for the development of multiple myeloma gene vaccine. Methods according to the target gene design, the encoding gene of MUC1-2VNTR was used as the target gene, and the synthetic gene was cloned into the vector pcDNA3.1 / myc-hisA. In vitro, recombinant human granulocyte-macrophage colony stimulating factor (rhGM-CSF) and recombinant human interleukin-4 (rhIL-4) were used to induce mature DC cells. The recombinant plasmid pcDNA3.1-2VNTR was constructed. DC cells were transfected with Lipofectimine liposome mediated transfection of DC cells (DC-2VNTR) and transfected with unloaded pcDNA3.1 (DC-pcDNA3.1) and lipofctamine treated DC (DC-Lipo) as control group. The expression of recombinant cells in DC cells was detected by Western blotting, and the homologous T cells were stimulated in vitro. Cytotoxicity assay was used to detect the cytotoxicity of CTL induced by DC-2 VNTR on multiple myeloma cell line (U266). The result is 1: 1. The recombinant plasmid pcDNA3.1-2VNTR / myc-hisA2 was successfully constructed. Culture and identification of human peripheral blood DC cells. The expression of MUC1-2VNTR in DC cells was detected by Western blot. CTL induced by DC-2VNTR had a significant killing effect on multiple myeloma cells (U266) and was significantly higher than that in DC-pcDNA3.1 group and DC-Lipo group (P0.05). Conclusion: DC transfected with MUC1-2VNTR gene can induce CTL response and specifically kill U266 multiple myeloma cell line.
【學位授予單位】：昆明醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2016
【分類號】：R733.3

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1 秋玉珍;MUC1-2VNTR基因修飾的DC對骨髓瘤細胞的體外殺傷效應的研究[D];昆明醫(yī)科大學;2016年

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