大肠杆菌麦芽糖结合蛋白非特异性活化T细胞及其对血液肿瘤细胞作用机制研究

发布时间：2018-11-28 10:20

【摘要】：在现代分子克隆中，人们普遍认为大肠杆菌麦芽糖结合蛋白（maltose bindingprotein, MBP）无生物学活性或具有较低的生物学活性，故将其作为分子生物学常用的标签蛋白。各种实验性病原菌和病毒的亚单位疫苗即利用了它的这一特性来进行疫苗研究。然而，最近有文献报道MBP通过TLR4促进DC细胞的成熟并分泌细胞因子。本课题组研究发现MBP能提高MUC1的免疫原性，与BCG协同显著增强细胞免疫应答，明显抑制小鼠皮下人乳腺癌移植瘤和小鼠Lewis肺癌移植瘤和转移瘤的生长。探讨肿瘤生长抑制机制的结果显示，MBP与BCG联合免疫组小鼠乳腺癌组织周围有大量淋巴细胞浸润，且CD4+T淋巴细胞较MBP或BCG单独免疫组显著增多。研究也发现MBP不仅能增强巨噬细胞吞噬活性，诱导M1型巨噬细胞活化，也可促进NK细胞活化。研究结果均提示MBP具有免疫增强作用，但其具体作用及其机制还不清楚。本研究通过探讨MBP对小鼠脾脏淋巴细胞的作用，明确MBP对T细胞的作用及其诱导的免疫应答类型。研究表明MBP非特异性刺激T淋巴细胞增殖，诱导Th1细胞活化，且MBP直接作用于淋巴细胞。本研究揭示MBP可直接刺激机体的免疫系统，诱导Th1方向的细胞免疫应答，为MBP作为一种新型免疫增强剂应用于肿瘤疫苗提供理论基础。目前在肿瘤疫苗的研究中，力求能寻找到诱导Th1和CTL细胞应答的佐剂，用于肿瘤疫苗中诱导细胞免疫应答。MBP可诱导Th1细胞活化，其有望成为肿瘤疫苗的佐剂。本研究室已经展开了这方面的工作，已对MBP作为肿瘤疫苗的佐剂刺激机体产生细胞免疫应答而发挥抗肿瘤作用进行了相关文献报道。本研究拟从另一角度即MBP对肿瘤细胞的直接作用这一角度展开研究。文献报道微生物佐剂可直接作用于肿瘤细胞发挥促凋亡或抗凋亡作用，尤其在血液肿瘤，故本研究将进一步探讨MBP是否可在体外直接作用于血液肿瘤细胞发挥生物学作用及其具体的机制，希望在探讨MBP抗肿瘤作用的同时寻找MBP的作用靶点。本研究表明MBP直接作用于U937细胞，通过TLR2活化细胞的NF-κB和MAPK信号通道，并调节细胞表面TLRs和MyD88的表达，促进U937细胞增殖和分化；MBP直接作用于Jurkat细胞，活化细胞的NF-κB和MAPK信号通道，并调节细胞表面TLRs和MyD88的表达，，促进Jurkat细胞增殖，但这种作用未通过TLR2和TLR4发挥，同时研究发现MBP提高了U937和Jurkat细胞对化疗药物紫杉醇的敏感性。提示MBP直接作用于U937和Jurkat细胞，使细胞处于增殖状态，与化疗药物联合使用可提高化疗药物的作用效果。本研究揭示MBP可直接增强机体的免疫应答，为MBP作为一种新型免疫增强剂的广泛推广提供理论基础，并为血液肿瘤的免疫治疗提供新的佐剂和研究思路。
[Abstract]:In modern molecular cloning, it is generally believed that E. coli maltose binding protein (maltose bindingprotein, MBP) has no biological activity or low biological activity, so it is regarded as a commonly used tag protein in molecular biology. The subunit vaccines of various experimental pathogens and viruses use this property for vaccine research. Recently, however, it has been reported that MBP promotes the maturation and secretion of cytokines in DC cells through TLR4. Our study found that MBP could enhance the immunogenicity of MUC1, enhance the cellular immune response in combination with BCG, and inhibit the growth of subcutaneous human breast cancer xenografts in mice and Lewis lung cancer xenografts and metastases in mice. The results of the study on the mechanism of tumor growth inhibition showed that a large number of lymphocytes infiltrated around the breast cancer tissues of mice immunized with MBP and BCG, and CD4 T lymphocytes increased significantly compared with those of MBP or BCG alone. It was also found that MBP not only enhanced the phagocytic activity of macrophages, induced the activation of M1 macrophages, but also promoted the activation of NK cells. All the results suggest that MBP has immune enhancement effect, but its specific role and mechanism are not clear. The purpose of this study was to investigate the effect of MBP on T cells and the type of immune response induced by MBP. The results showed that MBP stimulated the proliferation of T lymphocytes and induced the activation of Th1 cells, and MBP acted directly on lymphocytes. This study revealed that MBP can directly stimulate the immune system and induce cellular immune response in the direction of Th1, which provides a theoretical basis for the application of MBP as a new type of immune enhancer in tumor vaccines. At present, in the research of tumor vaccine, we try to find the adjuvant to induce Th1 and CTL cell response, which can be used to induce cellular immune response in tumor vaccine. MBP can induce the activation of Th1 cells, which is expected to be the adjuvant of tumor vaccine. This work has been carried out in our laboratory, and it has been reported that MBP, as an adjuvant of tumor vaccine, can stimulate the cellular immune response and play an antitumor role in the body. The purpose of this study is to study the direct effect of MBP on tumor cells. It has been reported that microbial adjuvants can directly promote apoptosis or inhibit apoptosis in tumor cells, especially in blood tumors. Therefore, this study will further explore whether MBP can directly act on blood tumor cells in vitro and its specific mechanism. We hope to explore the anti-tumor effect of MBP and seek the target of MBP at the same time. This study showed that MBP acts directly on U937 cells, activates NF- 魏 B and MAPK signaling channels through TLR2, and regulates the expression of TLRs and MyD88 on the surface of U937 cells, thus promoting the proliferation and differentiation of U937 cells. MBP acts directly on Jurkat cells, activates NF- 魏 B and MAPK signal channels, regulates the expression of TLRs and MyD88 on the cell surface, and promotes the proliferation of Jurkat cells, but this effect is not played by TLR2 and TLR4. At the same time, MBP increased the sensitivity of U937 and Jurkat cells to paclitaxel. The results suggest that MBP acts directly on U937 and Jurkat cells, which makes the cells proliferate. The combination of MBP and chemotherapeutic drugs can improve the effect of chemotherapeutic drugs. This study revealed that MBP can directly enhance the immune response of the body, provide a theoretical basis for the wide application of MBP as a new immune enhancer, and provide a new adjuvant and research idea for immunotherapy of blood tumors.
【学位授予单位】：吉林大学
【学位级别】：博士
【学位授予年份】：2012
【分类号】：R392.9

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