滤泡性辅助T细胞及滤泡性调节T细胞在结直肠癌患者外周血中的表达及其临床意义

发布时间：2018-06-23 16:14

本文选题：Tfh细胞 + Tfr细胞　；参考：《河北医科大学》2015年硕士论文

【摘要】：背景:结直肠癌是全世界发病率排名第三的恶性肿瘤,每年有超过100万患者确诊为结直肠癌。传统治疗手段(手术,化疗,放疗,靶向治疗)疗效不理想。除了肿瘤本身的机制外,肿瘤微环境特别是炎症反应、免疫反应与肿瘤的进展甚至放化疗、靶向药物抵抗有关。可以说,肿瘤免疫治疗的时代已经到来。事实上越来越多的证据表明,在预防肿瘤的发生、生长和转移扩散中免疫系统扮演重要的角色。对于CD4+T淋巴细胞亚群的研究是近年来免疫学研究的一个热点。Tfh作为CD4+T淋巴细胞亚群的一种,是近年来研究发现的一种特殊的辅助性T细胞,表达CXCR5的Tfh细胞在CXCL13的趋化下,被募集到淋巴滤泡,参与诱导B细胞的分化及生发中心的形成,通过产生IL-21等细胞因子以及表达ICOS等粘附分子来调节机体的免疫功能。而Tfr细胞作为Tfh细胞的调节对应物,兼有Tfh和Foxp3+Treg细胞的表型特征,抑制生发中心的形成及B细胞的分化,充当与Tfh相反的角色。近来研究表明失调的免疫系统在乳腺癌、非小细胞肺癌和肉瘤的发生中扮演重要的作用。而关于Tfh和Tfr在结直肠癌患者中表达的研究,国内外尚少有报道。本研究分别采用流式细胞术和酶联免疫吸附实验检测结直肠癌患者和健康人外周血Tfh、Tfr的表达和上清IL-21的含量,并分析其与临床病理参数的关系,探讨其在结直肠癌中表达的意义。目的:检测Tfh和Tfr在结直肠癌患者和健康对照组人外周血表达水平,同时测定外周血上清IL-21含量。分析Tfh和Tfr与临床病理参数的关系,探讨其在结直肠癌中表达的意义。方法:1通过流式细胞术检测实验组结直肠癌患者和健康对照组人外周血中CD4+CXCR5+PD-1+Foxp3-Tfh细胞和CD4+CXCR5+PD-1+Foxp3+Tfr细胞的表达。2用酶联免疫吸附试验(ELISA)方法,在分子水平检测实验组结直肠癌患者和健康对照组人外周血上清IL-21含量。结果:1研究对象的一般信息和临床特点实验组结直肠癌患者共62例,男性34例,女性28例,年龄范围23~83岁,平均年龄57.6岁,中位年龄58.5岁,病理均证实为腺癌。TNM分期:Ⅰ期7例,Ⅱ期21例,Ⅲ期25例,Ⅳ期9例(肝转移4例,肺转移3例,腹腔广泛转移1例,脑转移1例),其中Ⅰ、Ⅱ、Ⅲ及部分Ⅳ期(4例肝转移)患者均行手术治疗。有淋巴结转移的32例,无淋巴结转移的30例。健康对照组共30例,男性6例,女性24例,年龄范围26~78岁,平均年龄53.2岁,中位年龄54岁。实验组与健康对照组在年龄、性别、吸烟状态等方面相比没有统计学差异。2实验组和对照组外周血中Tfh细胞和Tfr细胞的表达以CD4+CXCR5+PD-1+Foxp3-作为Tfh细胞的标记,CD4+CXCR5+PD-1+Foxp3+作为Tfr细胞的标记。通过流式细胞术检测实验组结直肠癌患者和对照组健康人外周血中Tfh细胞和Tfr细胞的表达。P1门为外周血淋巴细胞,设CD4+T细胞为2门,Q3和Q4分别为外周血中Tfh细胞和Tfr细胞的表达情况。3实验组和对照组人外周血Tfh细胞和Tfr细胞的表达实验组CRC患者外周血Tfh细胞占CD4+T细胞的百分比高于健康对照组(2.93±2.48vs2.09±1.34,P=0.038);而实验组CRC患者外周血Tfr细胞低于健康对照组(0.16±0.09vs0.2±0.11,P=0.047)。4结直肠癌患者外周血Tfh细胞与临床病理参数的关系有淋巴结转移者Tfh细胞占CD4+T细胞百分比较无淋巴结转移者高(2.85±1.5 vs2.20±1.2)。各TNM分期间Tfh细胞占CD4+T细胞百分比相比较,Ⅰ期(1.39±0.64)和Ⅳ期(1.97±0.82)相比较,P=0.04,有统计学差异;而Ⅰ期和Ⅱ期、Ⅲ期相比较及Ⅱ期、Ⅲ期、Ⅳ期间相互比较P值均大于0.05,没有统计学差异。比较CRC患者CEA5ng/ml和CEA5ng/ml两组间Tfh细胞表达(3.06±2.79vs2.19±2.16,P=0.173),无统计学差异。5酶联免疫吸附实验检测实验组结直肠癌患者和健康对照组外周血上清IL-21水平,结果显示:实验组结直肠癌患者血清IL-21水平高于健康对照组(196.48±69.85vs149.85±91.49,P0.05)。结论:1结直肠癌患者外周血中Tfh细胞表达量升高,同时伴有Tfr表达量下降,且外周血上清IL-21水平升高。2 Tfh、Tfr细胞可能参与了结直肠癌进展。
[Abstract]:Background: colorectal cancer is the third most malignant tumor in the world. More than 1 million patients are diagnosed with colorectal cancer every year. Traditional treatments (surgery, chemotherapy, radiotherapy, targeted therapy) are not effective. In addition to the mechanism of the tumor itself, the tumor microenvironment especially the inflammatory reaction, the immune response and the progression of tumor and even radiotherapy and chemotherapy. The age of tumor immunotherapy has come. In fact, more and more evidence suggests that the immune system plays an important role in the prevention of tumor occurrence, growth and metastasis, and the study of CD4+T lymphocyte subsets is a hot spot in immunological research in recent years as a CD4+T lymph node. A kind of cell subgroup is a special kind of auxiliary T cell found in recent years. The Tfh cells expressing CXCR5 are recruited into the lymphoid follicles in the chemotactic of CXCL13, participate in the differentiation of B cells and the formation of the germinal center, and regulate the immune function by producing IL-21 and other adhesion molecules such as ICOS and other cytokines. Tfr cells, as a regulatory counterpart of Tfh cells, also have phenotypic characteristics of Tfh and Foxp3+Treg cells, inhibit the formation of germinal centers and differentiate between B cells, and act as opposite roles to Tfh. Recent studies have shown that the dysfunctional immune system plays an important role in the development of breast cancer, non small cell lung cancer and sarcomas, while Tfh and Tfr are in place. The study of colorectal cancer patients was rarely reported at home and abroad. Flow cytometry and enzyme-linked immunosorbent assay were used to detect the expression of Tfh, Tfr and the content of IL-21 in the peripheral blood of colorectal cancer patients and healthy people, and to analyze the correlation with the clinicopathological parameters and to explore the significance of their expression in colorectal cancer. To detect the level of peripheral blood expression of Tfh and Tfr in colorectal cancer patients and healthy controls, and to determine the IL-21 content of the peripheral blood supernatant, and to analyze the relationship between Tfh and Tfr and the clinicopathological parameters and to explore the significance of their expression in colorectal cancer. Methods: 1 the flow cytometry was used to detect the colorectal cancer patients and the healthy control group. The expression of CD4+CXCR5+PD-1+Foxp3-Tfh and CD4+CXCR5+PD-1+Foxp3+Tfr cells in peripheral blood.2 by enzyme linked immunosorbent assay (ELISA) method, at the molecular level, the content of IL-21 in the peripheral blood supernatant of colorectal cancer patients and healthy controls was detected at the molecular level. Results: 1 of the experimental group of colorectal cancer patients with the same information and clinical characteristics A total of 62 cases, 34 male and 28 female, age range 23~83 years, average age 57.6 years, middle age 58.5 years old, pathology confirmed.TNM staging of adenocarcinoma: stage I, 21 cases, stage III 25 cases, stage IV 9 cases (4 cases of liver metastasis, 3 lung metastasis, peritoneal metastasis 1, 1 cases of brain metastasis), of which, I, II, III and part IV stage (4 case of liver metastases) There were 32 cases of lymph node metastasis and 30 cases without lymph node metastasis. The healthy control group was 30 cases, 6 men and 24 women, the age range was 26~78 years, the average age was 53.2 years and the median age was 54 years. The experimental group and the healthy control group had no statistical difference between the.2 experiment group and the control group at the age, sex, and smoking status. The expression of Tfh and Tfr cells in the blood was marked by CD4+CXCR5+PD-1+Foxp3- as a Tfh cell, and CD4+CXCR5+PD-1+Foxp3+ as a marker for Tfr cells. The expression of Tfh cells and Tfr cells in peripheral blood of the colorectal cancer patients and the control group was detected by flow cytometry. The.P1 gate of the peripheral blood of the peripheral blood was 2 of the peripheral blood lymphocytes, and the CD4+T cells were set up to be 2. The expression of Tfh and Tfr cells in peripheral blood, Q3 and Q4, respectively,.3 experimental group and control group, the expression of Tfh cells and Tfr cells in the peripheral blood of the control group, the percentage of Tfh cells in the peripheral blood of CRC patients was higher than that of the healthy control group (2.93 + 1.34, P=0.038), and the peripheral blood of the experimental group was lower than the healthy control group. The relationship between the peripheral blood Tfh cells and the clinicopathological parameters in the patients with.4 colorectal cancer (0.16 + 0.09vs0.2 + 0.11, P=0.047) with lymph node metastasis, Tfh cells accounted for the percentage of CD4+T cells with no lymph node metastasis (2.85 + 1.5 vs2.20 + 1.2). The percentage of Tfh cells in TNM staging was compared with the percentage of CD4 +T cells, stage I (1.39 + 0.64) and IV stage (1.97 + 0.82) compared, P=0.04, there were statistical differences, while phase I and stage II, phase II, stage II, stage III and IV were more than 0.05, and there was no statistical difference. Tfh cells in CEA5ng/ml and CEA5ng/ml two groups of CRC patients were compared with Tfh cells (3.06 + 2.79vs2.19 + 2.16, P=0.173), and there was no statistical difference in the.5 enzyme immunoadsorption experiment The IL-21 level of peripheral blood supernatant in the experimental group and the healthy control group was detected. The results showed that the serum IL-21 level of the colorectal cancer patients in the experimental group was higher than that of the healthy control group (196.48 + 69.85vs149.85 + 91.49, P0.05). Conclusion: the amount of Tfh cells in the peripheral blood of 1 colorectal cancer patients was increased, and the expression of Tfr was decreased, and the peripheral blood was in the peripheral blood. Supernatant IL-21 level increased.2 Tfh, Tfr cells may participate in the progression of colorectal cancer.
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R735.34

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