本文選題：胰腺癌 + HSP70多肽復合物�。� 參考：《昆明醫(yī)學院》2009年碩士論文

【摘要】： 目的:探討從胰腺癌腫瘤塊中分離純化熱休克蛋白70-多肽復合物(heatshock protein 70-polypeptide complexes,HSP70-PCs)的方法,研究使用HSP70多肽復合物構建樹突狀細胞(dendritic cell,DC)多肽疫苗,觀察其在體外對CTL的增殖和活化效果。并進行免疫治療荷瘤小鼠,觀察小鼠的腫瘤大小及存活期。 方法:采用低滲裂解、ConA-Sepharose親和層析及ADP-Agarose親和層析法,從小鼠胰腺癌(MPC83)腫瘤組織中純化HSP70-PCs,所得蛋白經聚丙烯酰胺凝膠電泳檢測,Bradford法測定蛋白濃度,計算獲得率。用提純的HSP70-PCs激活小鼠骨髓來源的樹突狀細胞(DC),制備樹突狀細胞HSP70多肽腫瘤疫苗,用MTT法檢測混合淋巴細胞反應(mixed lymphocyte response,MLR)中HSP70-PCs致敏DC對CTL的增殖及活化效果;并觀察DC誘導的腫瘤特異性細胞毒性T淋巴細胞(cytotoxic T lymphocyte,CTL)對腫瘤細胞的特異性殺傷活性,建立MPC83荷瘤小鼠模型,并將DC接種于荷瘤小鼠皮下,觀察樹突狀細胞HSP70多肽腫瘤疫苗對腫瘤生長的抑制作用及對荷瘤小鼠生存期的影響。 結果:純化蛋白經鑒定為分子量70KD的HSP70-PCs;平均1克濕重腫瘤組織中獲得79.95μg的HSP70-PCs;HSP70-PCs在10-12μg/ml范圍內可達到刺激樹突狀細胞最強效果,用HSP70-PCs負載的DC能增強T細胞的增殖和活化能力,其誘導的CTL對小鼠胰腺癌MPC83腫瘤細胞具有特異性殺傷作用;應用樹突狀細胞HSP70多肽腫瘤疫苗治療荷瘤小鼠能顯著抑制荷瘤小鼠腫瘤的生長,延長荷瘤小鼠存活期。 結論:用上述層析法可分離出較高純度、保留腫瘤抗原信息的HSP70-PCs;體外將其負載經聯合細胞因子誘導培養(yǎng)的DC可激活T淋巴細胞,產生對該腫瘤細胞的特異性殺傷效應。體內對胰腺癌荷瘤小鼠具有顯著的免疫治療效果。
[Abstract]:Objective: to investigate the method of isolation and purification of heat shock protein 70-polypeptide complex HSP70-PCsfrom pancreatic cancer tumor, to study the construction of dendritic cell dendritic cell dendritic cell DCS vaccine using HSP70 polypeptide complex, and to observe the effect of HSP70-PCS on the proliferation and activation of CTL in vitro.The tumor size and survival period were observed.Methods: HSP70-PCswere purified from mouse pancreatic carcinoma tissue by ConA-Sepharose affinity chromatography and ADP-Agarose affinity chromatography. The protein was detected by polyacrylamide gel electrophoresis to determine the protein concentration and the yield was calculated.Dendritic cells derived from mouse bone marrow were activated with purified HSP70-PCs to prepare HSP70 polypeptide tumor vaccine from dendritic cells. The proliferation and activation of HSP70-PCs sensitized DCs in mixed lymphocyte response to MLRs were detected by MTT method.The specific cytotoxic cytotoxic T lymphocytes (CTL) induced by DC on tumor cells were observed. The MPC83 tumor-bearing mice model was established, and the DC was inoculated subcutaneously into the tumor-bearing mice.To observe the inhibitory effect of dendritic cell HSP70 polypeptide tumor vaccine on tumor growth and the survival time of tumor-bearing mice.Results: the purified protein was identified as HSP70-PCs with molecular weight of 70KD, and 79.95 渭 g HSP70-PCS70-PCs were obtained from tumor tissues with an average of 1 g wet weight, which could stimulate dendritic cells in the range of 10-12 渭 g/ml. DC loaded with HSP70-PCs could enhance the proliferation and activation of T cells.The CTL induced by CTL has a specific killing effect on MPC83 tumor cells in mice, and dendritic cell HSP70 polypeptide tumor vaccine can significantly inhibit the tumor growth and prolong the survival period of tumor-bearing mice.Conclusion: HSP70-PCs with high purity and retained tumor antigen information can be isolated by the method mentioned above, and the DC induced by the combination of cytokines can activate T lymphocytes in vitro and produce specific killing effect on the tumor cells.In vivo, there is a significant immunotherapy effect on mice bearing pancreatic cancer.
【學位授予單位】：昆明醫(yī)學院
【學位級別】：碩士
【學位授予年份】：2009
【分類號】：R392

【參考文獻】

相關期刊論文 前7條

1 陳蘭英,趙楨,李冰冰,屈二軍,王曉濤,陳建南;HSP70的腫瘤免疫作用及其基因表達調節(jié)[J];中國腫瘤;2004年07期

2 尹燕明,秦慶亮,張偉,趙晶,張長麗,于建國;肝細胞癌熱休克蛋白70過表達與自發(fā)性癌細胞凋亡[J];中華肝臟病雜志;2001年02期

3 傅慶國,沈曉東,孟凡東,郭仁宣;熱休克蛋白70與IL-2抗腫瘤作用的對比研究[J];中華腫瘤雜志;2002年04期

4 黃波,馮作化,張桂梅,李東,王洪濤;Hsp70-H22腫瘤抗原肽復合物激活樹突狀細胞誘導抗腫瘤免疫的研究[J];中華腫瘤雜志;2002年05期

5 吳偉忠!200032,劉康達!200032,高艷琴,吳厚生,湯曉雷,湯釗猷!200032;熱休克蛋白70的純化及其抗小鼠肝癌作用的研究[J];中華腫瘤雜志;2000年02期

6 黃波,馮作化,張桂梅;Hsp70-腫瘤抗原肽復合物修飾的DC疫苗體內外特異性抗瘤作用[J];中國免疫學雜志;2002年09期

7 郭愛林,隋延仿,曲萍,張立紅,葉菁,王小平;體外構建的HSP70-肝癌抗原肽誘導抗原肽特異性免疫反應[J];中國免疫學雜志;2001年11期

，

本文編號：1752157