炎症性肠病促进乳腺癌肺转移的机制研究

发布时间：2019-03-06 16:08

【摘要】：目的通过建立小鼠炎症性肠病乳腺癌肺转移模型,观察炎症性肠病是否可加速乳腺癌的肺部转移过程,探索其可能机制,以期为临床上防治乳腺癌合并炎症性肠病患者发生肿瘤远处转移提供新的干预策略。方法1依据实验室前期实验动物造模方法,以Balb/c雌鼠为实验对象,建立2.5%葡聚糖硫酸钠(dextran sulfate sodium,DSS)诱导的炎症性肠病(inflammatory bowel disease,IBD)模型,并在此模型基础上通过尾静脉接种小鼠乳腺癌细胞4T1 100ul(1×106/ml)建立IBD乳腺癌肺转移动物模型,动物实验结果重复2次,每次实验动物中每组动物至少10只(每组分别选取5只动物进行统计学分析)。2通过结肠组织病理切片评价IBD乳腺癌肺转移模型中的结肠炎症情况,通过肺组织病理切片观察IBD乳腺癌肺转移模型中的肺转移情况。3应用流式细胞术、血细胞计数仪检测小鼠外周血中中性粒细胞情况,应用免疫组织化学染色方法检测小鼠肺组织中中性粒细胞浸润情况。4应用脱氧核糖核酸酶(deoxyribonuclease,DNase-I)破坏中性粒细胞胞外捕获网结构(neutrophil extracellular trap,NET),肺组织病理切片观察小鼠肺转移情况。5统计学方法:应用SAS9.1统计软件包对数据进行统计分析,所有计量资料用(均数±标准差)来表示。两组样本之间差异的显著性分析采用t检验,多组样本间差异的显著性分析采用One-way ANOVA单因素方差分析,多组之间两两比较(事后检验)采用Bonferroni法。在炎性细胞浸润和组织破坏程度评估分析中,两组样本(多值有序变量的成组设计资料)之间采用Wilcoxon秩和检验。P0.05视为有统计学意义。结果1 2.5%DSS可诱导Balb/c雌鼠明显的结肠炎症反应,实验过程中观察可见经DSS诱导的实验小鼠发生明显腹泻、血便,解剖见结肠组织明显充血,长度明显缩短(P0.01),病理切片观察结肠组织可见大量的炎性细胞浸润及组织损伤。2在DSS诱导的IBD乳腺癌肺转移模型中,肺组织病理切片可见经DSS诱导的小鼠,与正常饮水组相比,其乳腺癌肺转移明显增加(P0.01)。3血细胞计数、流式细胞术结果显示,DSS可诱导小鼠外周血中中性粒细胞的动员,其数量及比例均增加(P0.05),免疫组织化学染色结果显示,转移部位肺组织中浸润的中性粒细胞也明显增加(P0.01)。4应用DNase-I破坏中性粒细胞NET结构,可逆转DSS诱导的IBD促进乳腺癌肺转移增加(P0.05)。结论DSS诱导的小鼠IBD乳腺癌肺转移模型可见明显的乳腺癌肺转移,其机制可能为DSS诱导小鼠外周血中中性粒细胞动员并浸润至肺组织,动员的中性粒细胞可通过形成NET结构促进乳腺癌肺转移,应用脱氧核糖核酸酶(DNase-I)破坏中性粒细胞NET结构,可逆转DSS诱导的小鼠乳腺癌肺转移增加。
[Abstract]:Objective to investigate whether inflammatory bowel disease can accelerate the process of lung metastasis of breast cancer and explore its possible mechanism by establishing a model of lung metastasis in mice with inflammatory bowel disease. In order to provide a new intervention strategy for the prevention and treatment of distant metastasis in patients with breast cancer complicated with inflammatory bowel disease. Methods 1 the model of inflammatory bowel disease (inflammatory bowel disease,IBD) induced by 2.5% glucosulfate sodium sulfate (dextran sulfate sodium,DSS) was established in Balb/c female rats according to the experimental animal model. On the basis of this model, mice breast cancer cell line 4T1 100ul (1 脳 10 ~ 6 / ml) was injected into the tail vein to establish the animal model of lung metastasis of IBD breast cancer. The results of the animal experiment were repeated twice. At least 10 animals in each group were selected for each experiment (5 animals in each group were selected for statistical analysis). 2 colonic inflammation in IBD breast cancer lung metastasis model was evaluated by histopathological sections of colon. The lung metastasis in the model of IBD breast cancer was observed by pathological sections of lung tissue. 3The neutrophils in peripheral blood of mice were detected by flow cytometry and blood cell counter. Immunohistochemical staining was used to detect neutrophil infiltration in the lung tissue of mice. (4) the extracellular capture net structure (neutrophil extracellular trap,NET) of neutrophils was destroyed by deoxyribonuclease (deoxyribonuclease,DNase-I). The pathological sections of lung tissue were used to observe the lung metastasis in mice. 5 Statistical methods: the data were statistically analyzed by SAS9.1 software package, and all the data were expressed by the mean 卤standard deviation (mean 卤standard deviation). T-test was used to analyze the significance of the difference between the two groups, One-way ANOVA single factor analysis of variance was used to analyze the significance of the difference between multiple samples, and Bonferroni method was used to compare the differences between the two groups (post-test). In the evaluation analysis of inflammatory cell infiltration and tissue destruction, Wilcoxon rank sum test was used between the two groups of samples (group design data of multi-valued ordered variables). P0.05 was regarded as statistically significant. Results 1 2.5%DSS could induce obvious colonic inflammation in Balb/c female rats. In the course of the experiment, the mice induced by DSS had obvious diarrhea, blood stool, and the colon tissue was obviously hyperemia in anatomy, and in the course of the experiment, there were obvious diarrhea and blood stool in the experimental mice induced by DSS. The length was significantly shortened (P0.01), and a large number of inflammatory cell infiltration and tissue injury were observed in colonic tissue by pathological section. 2 in the lung metastasis model of IBD breast cancer induced by DSS, the mice induced by DSS could be seen in the pathological section of lung tissue. Compared with the normal drinking water group, the lung metastasis of breast cancer was significantly increased (P0.01). 3Blood cell count and flow cytometry showed that DSS could induce the mobilization of neutrophils in peripheral blood of mice, and the number and proportion of neutrophils were increased (P0.05). Immunohistochemical staining showed that the infiltration of neutrophils in the metastatic lung tissue was also significantly increased (P0.01). (4) the NET structure of neutrophils was destroyed by DNase-I. Reversible DSS-induced IBD enhanced lung metastasis in breast cancer (P0.05). Conclusion the lung metastasis of breast cancer in IBD mice induced by DSS may be related to the mobilization of neutrophils in peripheral blood and infiltration of neutrophils into lung tissue induced by DSS, and the mechanism may be that DSS can induce the mobilization of neutrophils in peripheral blood of mice. The mobilized neutrophils could promote the lung metastasis of breast cancer by forming NET structure and destroy the NET structure of neutrophils by using deoxyribonuclease (DNase-I) to reverse the increase of DSS-induced lung metastasis of breast cancer in mice.
【学位授予单位】：华北理工大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R574;R737.9

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