基于中药方剂配伍机理的中药有效成分在食管癌前病变进程中的免疫调节作用

发布时间：2018-01-10 11:17

本文关键词：基于中药方剂配伍机理的中药有效成分在食管癌前病变进程中的免疫调节作用　出处：《河北医科大学》2015年硕士论文　论文类型：学位论文

【摘要】：目的:本课题拟研究“连翘、天花粉蛋白、党参、香加皮”中有效成分组成的“连花参加”方逆转化学致癌剂4-硝基喹啉-1-氧化物(4-nitro-quinoline-1-oxide,4NQO)诱导小鼠(C57BL/6)食管癌前病变的效果及潜在的免疫调节机制。方法:1选取300只C57BL小鼠动物模型,按随机原则分为6组,分别为空白对照组(A1组)(不做任何处理)30只;甲基纤维素对照组(A2组)30只,给予甲基纤维素灌胃,不给药;诱癌剂组(B组),治疗组(C组),预防组(D组),全反式维甲酸阳性对照组(E组),用灭菌水配制成100μg/ml浓度,采取正常饮水法给B、C、D、E四组小鼠。受试药“连花参加”方的配制方法为连翘脂素(2mg):天花粉蛋白(1.4mg):香加皮三萜类化合物(1.4mg):党参蛋白(0.7mg),按此比例混合溶于1.5%的甲基纤维素,定容20ml。D组在实验开始就给与中药方剂“连花参加”方灌胃。C组第15周开始给予中药方剂“连花参加”方灌胃,E组也在15周进行“全反式维甲酸(ATRA)”灌胃。C、D、E组小鼠每周均灌胃三次,每次0.1ml。2该实验从第12周时分别把A1、A2、B、D组小鼠处死5只,观察结果显示小鼠食管未出现不典型增生。在15周时,与12周采取同样的处理,HE染色结果显示小鼠食管发现早期不典型增生。从15周开始,将B组、C组、D组、E组100μg/ml 4NQO水溶液统一更换为灭菌水。第18、21周,A1、A2组各处死5只,B、C、D、E组各处死15只,第24周时,处死剩余小鼠,均为断颈处死。3建立小鼠癌前病变的模型,HE检测B组不同时期组织的病理学变化,鉴定肿瘤进展的阶段。4提取小鼠脾细胞,观察用药前后对脾细胞中淋巴细胞的影响。用流式细胞法检测小鼠脾细胞中CD4+,CD8+,B7-H4(B7homologue4),CD4+CD25+foxp3+等T细胞及其表面分子在不同时间点的组间变化情况。结果:1成功建立了小鼠食管癌癌前病变的模型,HE染色发现第15周时4NQO诱癌组的小鼠开始有轻度不典型增生。并且随着时间的延长,病变程度逐渐加深。2通过HE染色结果显示,小鼠食管组织病变程度在24周时,有显著性差异,P0.05,C、D、E组的致癌率比B组显著降低,P0.05,说明“连花参加”方可减缓食管癌病变的进程。3实验结果显示在18周和21周时各组间CD4+CD25+foxp3+调节性T细胞占脾淋巴细胞的百分率表达水平无统计学差异;在24周时各组间有统计学差异,P0.05。4 B7-H4在CD4+上的表达率在21周和24周时出现显著性差异,P0.05。B7-H4在CD8+T细胞和CD4+CD25+foxp3+T细胞上的表达率在18周、21周和24周时均无显著性差异。结论:1成功建立了小鼠食管癌前病变动物模型。2“连花参加”方通过抑制Treg细胞在脾淋巴细胞上的表达和B7-H4在CD4+T细胞的表达,促进了机体抗肿瘤免疫作用,减缓了食管癌前病变进程。
[Abstract]:Objective: to study "Forsythia suspensa, Trichosanthin, Codonopsis pilosula". "Lianhuayan", an active component of Xiangjia Peel, was used to reverse the chemical carcinogen 4-nitro-quinoline-1-oxide (4-nitro-quinoline-1-oxide). (4) the effect of C57BL / 6) on precancerous lesion and its potential immunomodulatory mechanism in mice were induced by NQO. Methods 300 C57BL mice were randomly divided into 6 groups. There were 30 rats in group A 1 (without any treatment) in the blank control group. 30 rats in group A _ 2 of methylcellulose control group were given intragastric administration of methylcellulose without administration of the drug. Cancer inducers group B, treatment group C, prevention group D, all trans retinoic acid positive control group, group E, 100 渭 g / ml of sterilizing water were given to B by normal drinking water. C,D. Group E mice. The preparation method of "Lianhua participating" prescription was Forsythia suspensa 2mg / g: Trichosanthin 1.4 mg / g: trichosanthin = 1.4 mg / g: Codonopsis Codonopsis protein (1. 4 mg / g / L); trichosanthin 1. 4 mg / g; triterpenoid compound 1. 4 mg / g; (0.7 mg). In this proportion, methyl cellulose dissolved in 1.5% was mixed. At the beginning of the experiment, group D was given the traditional Chinese medicine prescription "continuous flower participation". Group C was given the traditional Chinese medicine prescription "continuous flower participation" by stomach at the 15th week. Group E was also given ATRA-ATRA for 15 weeks. Mice in group E were given oral administration of A1A _ 2 three times a week, 0.1ml.2 each time from the 12th week. Five mice in group B were killed. The results showed that there was no atypical hyperplasia in the esophagus of the mice, and the same treatment was taken at 15 weeks and 12 weeks. The results of HE staining showed that early atypical hyperplasia was found in the esophagus of mice. From 15 weeks on, 100 渭 g / ml 4NQO solution of group B and C was replaced with 100 渭 g / ml 4NQO solution as sterilizing water. In group A _ 1 A _ 2, 15 rats were killed everywhere in group A _ 1 A _ 2 at 21 weeks. At the 24th week, the remaining mice were killed, all of them were killed by cervical amputation to establish the model of precancerous lesion in mice. He was used to detect the pathological changes of the tissues in group B at different stages and to identify the stage of tumor progression. 4. The spleen cells were extracted from mice. To observe the effect on lymphocytes in splenocytes before and after treatment, and to detect the CD4 CD8 B7-H4 B7homologue 4 in spleen cells of mice by flow cytometry (FCM). The changes of T cells and their surface molecules such as CD4 CD25 foxp3 were observed at different time points. Results the mouse model of precancerous lesion of esophageal carcinoma was successfully established at 1: 1. He staining showed that the mice in 4NQO induced cancer group began to have mild atypical hyperplasia at the 15th week, and with the extension of time, the degree of lesion gradually deepened by HE staining results showed. At 24 weeks after 24 weeks, there was significant difference in carcinogenic rate of P0.05CU DU E group compared with B group (P0.05). The results show that "Lianhua attends" can slow down the progression of esophageal cancer. 3. 3. The results show that CD4 CD25 foxp3 can be found between groups at 18 and 21 weeks. There was no significant difference in the percentage of regulatory T cells in spleen lymphocytes. There was a significant difference in the expression rate of P0.05.4 B7-H4 on CD4 between the groups at 24 weeks and 21 weeks and 24 weeks. The expression rate of P0.05.B7-H4 on CD8 T cells and CD4 CD25 foxp3 T cells was 18 weeks. There was no significant difference between 21 weeks and 24 weeks. Conclusion the mouse model of precancerous lesion was successfully established by 1: 1. The expression of Treg cells on spleen lymphocytes and B7-H4 on CD4 T cells were inhibited. Promoted the body anti-tumor immunity, slowed down the process of esophageal precancerous lesions.
【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】：R735.1

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