翁连解毒汤对溃疡性结肠炎大鼠IL-13、TLR4表达的影响
发布时间：2018-07-28 11:12
【摘要】：目的:溃疡性结肠炎(UC)是一种病因尚不明确的直肠和结肠慢性非特异性炎症性疾病。病情轻重不一,反复发作,且病程较长。本实验通过观察翁连解毒汤对UC模型大鼠一般情况、疾病活动指数(DAI)评分、结肠组织病理学形态、白细胞介素-13(IL-13)及Toll样受体4(TLR4)表达的影响,探讨翁连解毒汤治疗溃疡性结肠炎的作用机理,并为临床应用提供实验依据。方法:60只Wistar雄性健康清洁级大鼠(体重220g-250g)适应性喂养7天后,随机分为空白对照组(CK组)、模型组(MG组)、美沙拉嗪组(MS组)、中药高剂量组(HD组)、中剂量组(MD组)及低剂量组(LD组)6组,每组10只,予常规饲养。除CK组外,其余各组大鼠均采用2,4,6-三硝基苯磺酸(TNBS)/乙醇联合造模法设计UC大鼠模型。造模成功后第4天开始,CK组不做特殊处理,MG组予4ml/kg生理盐水灌胃,MS组予美沙拉嗪混悬液灌胃,中药各组大鼠分别予相应剂量的药物灌胃,连续药物干预2周后观察治疗期间大鼠精神状态、食量、体重以及粪便性状等一般情况的变化,并进行DAI评分。治疗结束后,各组大鼠禁食不禁水24小时,利多卡因麻醉,股动脉取血,酶联免疫吸附法(ELISA)检测血清IL-13含量,免疫组化法检测肠粘膜组织中TLR4的表达情况。结果:1一般情况观察整个治疗期间CK组大鼠精神状态良好,喜觅食,体毛光泽整洁,动作敏捷,体重增加,粪便为正常的黑色颗粒状。其余各组大鼠在造模成功后均出现不同程度的稀便或粘液脓血便,且肛周污秽,扎堆嗜蜷卧,不喜活动,精神差,被毛凌乱无光泽,进食量减少。药物干预后除MG组大鼠外,HD组、MD组、LD组、MS组四组大鼠精神状态、食量、体重等一般情况逐渐改善,稀便及黏液脓血便较前减少。其中以hd组大鼠症状改善最明显,ms组与md组大鼠相比,好转情况无明显差别,ld组大鼠情况不及以上三组。而mg组大鼠随着实验进度状态每况愈下,稀便或黏液脓血便加重,体重均有不同程度的减轻,精神萎靡,嗜蜷卧,被毛暗淡无光泽,不喜食。2dai评分结果hd组、md组、ld组、ms组均低于mg组,差异有统计学意义(p0.05);hd组相较于md组、ld组及ms组三组均低,差异有统计学意义(p0.05);md组与ms组均低于ld组,差异有统计学意义(p0.05);md组与ms组比较,差异无统计学意义(p0.05)。3结肠组织病理学观察ck组大鼠组织表面上皮完整,隐窝长且平直,未见分支、扭曲,杯状细胞数量多,固有膜内偶见炎细胞,黏膜肌层未见增厚,黏膜下层未见血管扩张充血和水肿;mg组可见溃疡表面被覆炎性渗出物及坏死组织,固有膜内大量炎细胞弥漫浸润;hd组大鼠组织表面上皮完整,隐窝长且平直,偶见分支,杯状细胞数量较多,固有膜内可见少量炎细胞;md组、ms组差异不明显,组织表面上皮不规则,隐窝分支少,杯状细胞数量较多,固有膜内可见较少炎细胞;ld组表面上皮不规则,隐窝呈分支状并且扭曲,杯状细胞数量明显减少,隐窝基底部与黏膜肌层距离增加,其间可见大量炎细胞片状浸润,黏膜肌层增厚。4指标检测il-13的表达:ck组il-13表达均高于hd组、md组、ld组、ms组以及mg组,差异有统计学意义(p0.05);hd组高于md组、ld组、ms组及mg组,差异有统计学意义(p0.05);md组、ms组高于ld组、mg组,差异有统计学意义(p0.05);ld组高于mg组,差异有统计学意义(p0.05);md组与ms组相比,无统计学意义(p0.05)。tlr4的表达:ck组tlr4表达低于hd组、md组、ld组、ms组以及mg组,差异具有统计学意义(p0.05);hd组低于md组、ld组、ms组及mg组,差异具有统计学意义(p0.05);md组、ms组低于ld组、mg组,差异具有统计学意义(p0.05);ld组低于mg组,差异具有统计学意义(p0.05);md组与ms组比较无统计学意义(P0.05)。结论:翁连解毒汤能够显著改善溃疡性结肠炎大鼠的一般情况和肠黏膜组织的损伤情况,中药高剂量组效果优于其它各治疗组,中剂量组与美沙拉嗪组疗效相当,低剂量组疗效次之。其作用机制可能是通过降低肠黏膜组织中TLR4的表达、上调IL-13的含量来实现的,通过影响某种信号转导通路,增加抑炎因子的释放,促使促炎因子和抑炎因子趋于动态平衡,从而阻断细胞损害过程,刺激局部组织修复再生,促进溃疡的愈合。
[Abstract]:Objective: ulcerative colitis (UC) is a chronic nonspecific inflammatory disease of the rectum and colon which is still unknown. The severity of the disease is different, repeated episodes, and the course of the disease is longer. This experiment was conducted by observing the general condition of the UC model rats, the disease activity index (DAI), the pathological morphology of the colon, and the interleukin -13. IL-13) and the effect of Toll like receptor 4 (TLR4) expression to explore the mechanism of Weng Lian Jiedu Decoction in the treatment of ulcerative colitis and provide experimental basis for clinical application. Methods: 60 Wistar male healthy clean rats (body weight 220g-250g) were fed for 7 days after adaptive feeding, and were randomly divided into blank control group (CK group), model group (MG group), mesalazine group (MS). Group (group HD), medium dose group (group HD), middle dose group (group MD) and low dose group (group LD), 10 rats in each group were given routine feeding. Except for group CK, all the other rats were designed with 2,4,6- three nitrobenzene sulfonic acid (TNBS) / ethanol combined model method to design UC rat model. After building the work, the group CK did not do special treatment, MG group was given 4ml/kg saline irrigation. The stomach and MS group were given the mesalazine suspension for gastric perfusion, and the rats in each group were given the corresponding dose of medicine. After 2 weeks of continuous drug intervention, the general situation of mental state, food intake, weight and fecal traits were observed and the DAI scores were observed. After the treatment of the knot, the rats in each group were fasting for 24 hours, lidocaine anesthesia. The content of serum IL-13 was detected by the femoral artery and enzyme linked immunosorbent assay (ELISA), and the expression of TLR4 in the intestinal mucosa was detected by immunohistochemical method. Results: 1 the general situation observed that the mental state of the rats in the CK group was good during the whole treatment period, looking for food, the shiny of the body hair was neat, the movement was agile, the weight was increased, and the feces as normal black granules. After the success of the model, the rats in all groups had different degrees of dilute stool or mucous pus and stool, and the perianal filth was filth, and had a bad mood, poor spirit, lustrous, and less eating. After the intervention of the MG group, the mental state of the four groups of rats in group HD, MD, LD and MS gradually improved, and the general condition of the diet, weight and weight gradually improved. The dilute stool and mucus pus and blood stool decreased more than that in the HD group. There was no obvious difference between the group MS and the MD group. In group LD, the condition of the rats was less than the above three groups. And the rats in group mg were worse with the progress of the experiment, and the weight of the dilute stool or mucus pus and blood was added, and the body weight was reduced in varying degrees, and the spirit was dispirited. In group MD, group MD, group LD, MS group were lower than group mg, and the difference was statistically significant (P0.05); HD group was lower than group MD, LD group and MS group were lower in HD group than in group MD (P0.05), and the difference was statistically significant (.2dai). The study significance (P0.05).3 colon histopathology observed that the tissue surface epithelium of group CK rats was complete, the recess long and straight, no branches, distortion, the number of goblet cells, innate inflammatory cells in the lamina propria, no thickening of the mucous membrane, and no vasodilator congestion and edema in the submucosa; in group mg, the surface of the ulcer was covered with inflammatory exudation and necrosis. Tissue, a large number of inflammatory cells infiltrated in the innate membrane; in group HD, the surface epithelium of the rats was complete, the crypts were long and straight, the number of goblet cells was more, the number of goblet cells was more, and a small amount of inflammatory cells were seen in the propria. In group MD, the differences in the MS group were not obvious, the epithelial epithelium was irregular, the crypts were small, the number of goblet cells were more, and the propri was less inflammatory. The surface epithelium of the LD group was irregular, the crypts were branched and twisted, the number of goblet cells decreased obviously, the distance between the basement and mucous layer of the recess increased, and a large number of inflammatory cells were infiltrated, and the.4 index of the muscularis layer thickened to detect the expression of IL-13. The IL-13 table in group CK was higher than that of the HD group, MD, LD, MS and Mg groups were different. Group HD was higher than group MD, group LD, MS group and mg group, the difference was statistically significant (P0.05); MD group, MS group was higher than LD group and mg group, the difference was statistically significant (P0.05). Group s and mg group, the difference was statistically significant (P0.05), and group HD was lower than group MD, LD group, MS group and mg group, and the difference was statistically significant (P0.05); MD group and MS group were lower than LD group, and the difference had statistical significance. Jiedu Decoction can significantly improve the general condition of ulcerative colitis and the damage of intestinal mucosa. The effect of high dose group of Chinese medicine is better than that of other treatment groups. The effect of medium dose group is equal to that of mesalazine group, and the effect of low dose group is second. The mechanism of its action may be to reduce the expression of TLR4 in intestinal mucosa and up regulation of IL-13. The content is achieved by affecting a signal transduction pathway, increasing the release of anti inflammatory factors and promoting the dynamic balance of proinflammatory and anti inflammatory factors, thus blocking the process of cell damage, stimulating the local tissue to repair and regenerating and promoting the healing of the ulcer.
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R285.5
本文编号：2149942
[Abstract]:Objective: ulcerative colitis (UC) is a chronic nonspecific inflammatory disease of the rectum and colon which is still unknown. The severity of the disease is different, repeated episodes, and the course of the disease is longer. This experiment was conducted by observing the general condition of the UC model rats, the disease activity index (DAI), the pathological morphology of the colon, and the interleukin -13. IL-13) and the effect of Toll like receptor 4 (TLR4) expression to explore the mechanism of Weng Lian Jiedu Decoction in the treatment of ulcerative colitis and provide experimental basis for clinical application. Methods: 60 Wistar male healthy clean rats (body weight 220g-250g) were fed for 7 days after adaptive feeding, and were randomly divided into blank control group (CK group), model group (MG group), mesalazine group (MS). Group (group HD), medium dose group (group HD), middle dose group (group MD) and low dose group (group LD), 10 rats in each group were given routine feeding. Except for group CK, all the other rats were designed with 2,4,6- three nitrobenzene sulfonic acid (TNBS) / ethanol combined model method to design UC rat model. After building the work, the group CK did not do special treatment, MG group was given 4ml/kg saline irrigation. The stomach and MS group were given the mesalazine suspension for gastric perfusion, and the rats in each group were given the corresponding dose of medicine. After 2 weeks of continuous drug intervention, the general situation of mental state, food intake, weight and fecal traits were observed and the DAI scores were observed. After the treatment of the knot, the rats in each group were fasting for 24 hours, lidocaine anesthesia. The content of serum IL-13 was detected by the femoral artery and enzyme linked immunosorbent assay (ELISA), and the expression of TLR4 in the intestinal mucosa was detected by immunohistochemical method. Results: 1 the general situation observed that the mental state of the rats in the CK group was good during the whole treatment period, looking for food, the shiny of the body hair was neat, the movement was agile, the weight was increased, and the feces as normal black granules. After the success of the model, the rats in all groups had different degrees of dilute stool or mucous pus and stool, and the perianal filth was filth, and had a bad mood, poor spirit, lustrous, and less eating. After the intervention of the MG group, the mental state of the four groups of rats in group HD, MD, LD and MS gradually improved, and the general condition of the diet, weight and weight gradually improved. The dilute stool and mucus pus and blood stool decreased more than that in the HD group. There was no obvious difference between the group MS and the MD group. In group LD, the condition of the rats was less than the above three groups. And the rats in group mg were worse with the progress of the experiment, and the weight of the dilute stool or mucus pus and blood was added, and the body weight was reduced in varying degrees, and the spirit was dispirited. In group MD, group MD, group LD, MS group were lower than group mg, and the difference was statistically significant (P0.05); HD group was lower than group MD, LD group and MS group were lower in HD group than in group MD (P0.05), and the difference was statistically significant (.2dai). The study significance (P0.05).3 colon histopathology observed that the tissue surface epithelium of group CK rats was complete, the recess long and straight, no branches, distortion, the number of goblet cells, innate inflammatory cells in the lamina propria, no thickening of the mucous membrane, and no vasodilator congestion and edema in the submucosa; in group mg, the surface of the ulcer was covered with inflammatory exudation and necrosis. Tissue, a large number of inflammatory cells infiltrated in the innate membrane; in group HD, the surface epithelium of the rats was complete, the crypts were long and straight, the number of goblet cells was more, the number of goblet cells was more, and a small amount of inflammatory cells were seen in the propria. In group MD, the differences in the MS group were not obvious, the epithelial epithelium was irregular, the crypts were small, the number of goblet cells were more, and the propri was less inflammatory. The surface epithelium of the LD group was irregular, the crypts were branched and twisted, the number of goblet cells decreased obviously, the distance between the basement and mucous layer of the recess increased, and a large number of inflammatory cells were infiltrated, and the.4 index of the muscularis layer thickened to detect the expression of IL-13. The IL-13 table in group CK was higher than that of the HD group, MD, LD, MS and Mg groups were different. Group HD was higher than group MD, group LD, MS group and mg group, the difference was statistically significant (P0.05); MD group, MS group was higher than LD group and mg group, the difference was statistically significant (P0.05). Group s and mg group, the difference was statistically significant (P0.05), and group HD was lower than group MD, LD group, MS group and mg group, and the difference was statistically significant (P0.05); MD group and MS group were lower than LD group, and the difference had statistical significance. Jiedu Decoction can significantly improve the general condition of ulcerative colitis and the damage of intestinal mucosa. The effect of high dose group of Chinese medicine is better than that of other treatment groups. The effect of medium dose group is equal to that of mesalazine group, and the effect of low dose group is second. The mechanism of its action may be to reduce the expression of TLR4 in intestinal mucosa and up regulation of IL-13. The content is achieved by affecting a signal transduction pathway, increasing the release of anti inflammatory factors and promoting the dynamic balance of proinflammatory and anti inflammatory factors, thus blocking the process of cell damage, stimulating the local tissue to repair and regenerating and promoting the healing of the ulcer.
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R285.5
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