【作者】 刘云；

【导师】 郭喜军；

【作者基本信息】 河北医科大学 ， 中医内科学（专业学位）， 2017， 硕士

【摘要】 目的:溃疡性结肠炎(ulcerative colitis,UC)是发生在直肠和结肠的、累及黏膜及黏膜下层的慢性非特异性炎症性疾病,发病率高,且易复发,病程较长。本实验通过观察翁连解毒汤方对溃疡性结肠炎大鼠的一般状态、疾病活动指数(DAI)评分、血清IL-17含量及结肠组织TLR2表达的影响,探讨翁连解毒汤治疗溃疡性结肠炎的作用机制,为临床应用提供实验依据。方法:将60只洁净雄性健康Wistar大鼠根据随机数字表随机分为6组,每组10只,分别为空白对照组(NG)、模型组(MG)、西药美沙拉嗪组(XG)、中药高剂量组(HG)、中药中剂量组(ZG)、中药低剂量组(LG)。空白对照组(NG)正常饲养,其余5组大鼠均采用三硝基苯磺酸(TNBS)/乙醇联合造模方法建立溃疡性结肠炎大鼠模型。自造模后第4天,各组大鼠除NG、MG正常饲养外,余治疗组大鼠除基础饲养外合用药物灌胃,其中XG组大鼠采取美沙拉嗪混悬液灌胃,HG、ZG和LG大鼠采用不同的浓度的翁连解毒汤灌胃治疗。连续治疗2周后,观察大鼠在各个时期的精神状态、饮食、被毛、粪便性状、体重等一般状态的变化并记录,做出疾病活动指数评分表。治疗结束后,各组大鼠禁食不禁水24小时,利多卡因麻醉大鼠取股动脉血,用双抗体夹心ABC-ELLISA法检测血清IL-17含量,解剖并取其肉眼观察病变较重部位的结肠组织,常规石蜡包埋、切片、HE染色,检测TLR2的表达情况。结果:1一般情况观察:NG大鼠一般状态良好,余组大鼠精神萎靡,反应迟钝,进食水量减少,体重下降,粪便稀薄夹杂粘液、脓血。药物灌胃治疗后各组大鼠精神状态好转,进食量明显增多,体重下降减慢,粘液、脓血便减少或消失,其中HG改善最明显,ZG、XG改善无明显差异,较HG略差,较LG较好。DAI评分结果提示:XG、HG、ZG、LG与MG相比较低,差异有统计学意义(P<0.05);XG、ZG、LG与HG相比较高,差异有统计学意义(P<0.05);XG与ZG无统计学意义(P>0.05);XG、ZG与LG相比较低,差异有统计学意义(P>0.05)。2结肠黏膜肉眼与病理学观察肉眼观察:NG大鼠上皮黏膜光滑完整。MG大鼠黏膜充血水肿糜烂面最明显。HG大鼠黏膜较光滑,溃疡面极少。XG和ZG大鼠黏膜欠光滑,可见较少的溃疡面。LG大鼠黏膜组织轻度充血水肿,溃疡灶较多。组织病理学观察:NG大鼠黏膜光滑,腺体结构排列规则。MG大鼠黏膜粗糙,可见多发、弥漫性糜烂或溃疡面,局部腺体破坏最明显,可见大量炎性细胞浸润。HG组黏膜较光滑完整,新生腺体最多。XG和ZG大鼠黏膜欠光滑,可见少许炎性细胞浸润,溃疡面较少。LG大鼠黏膜组织轻度充血水肿,遗留溃疡灶较多,少数处于愈合期,溃疡面及边缘覆于少量新生腺体,见附图Fig.1-Fig.12。3指标检测IL-17含量比较:MG、XG、HG、ZG、LG与NG相比含量较高,差异有统计学意义(P<0.05);HG、XG、ZG、LG与MG相比,含量较低,差异有统计学意义(P<0.05);XG、ZG、LG与HG相比含量较高,差异有统计学意义(P<0.05);XG、ZG与LG相比含量较低,且差异有统计学意义(P<0.05);XG与ZG相比较,差异无统计学意义(P>0.05)。TLR2表达比较:MG、XG、HG、ZG、LG与NG相比表达较高,差异有统计学意义(P<0.05);XG、HG、ZG、LG与MG相比,表达较低,差异具有统计学意义(P<0.05);HG较XG、ZG、LG相比,表达较低,差异有统计学意义(P<0.05);ZG、XG与LG相比,表达较低,差异有统计学意义(P<0.05);XG与ZG相比较,差异无统计学意义(P>0.05)。结论:翁连解毒汤能够改善溃疡性结肠炎大鼠的一般情况并且能够修复损伤的黏膜,降低DAI评分,并呈现一定的量-效关系,其特点为随着中药浓度的升高而疗效增加,其作用机制可能通过降低TLR2的表达、减少IL-17的含量,从而阻断炎症反应,减少炎性细胞浸润,刺激局部组织再生与修复,达到愈合溃疡面的作用。更多还原

【Abstract】 Objective : ulcerative colitis(ulcerative colitis,UC)is a chronic nonspecific inflammatory bowel disease,involving rectum and colonic mucosa and submucosa,having the characteristics of high incidence,recurring easily and long course.We can observe the effects of Weng LianJieDu decoction on the general state of rats,disease activity index(DAI),the content of interleukin-17(IL-17)and the expression of toll-like receptor-2,in order to explore the mechanism of WengLianJieDu decoction to UC rats and try to provide the experimental experience for clinical treatment.Methods : 60 clean and healthy male Wistar rats were divided into 6groups by random number able,10 rats of each group: the blank control group(NG),model group(MG),taking western medicine group(XG),taking ShuangHongQinYuTang decoction groups which includes high concentration(HG)、 middle concentration(ZG)and low concentration(LG).The blank control group were fed normally,others were manufactured into ulcerative colitis models by TNBS/ethanol.4 days after modeling,we selected a rat randomly from the group(except NG group),which was beheaded anatomy and we took the heavy colon tissue with naked eye in which the mucosa appeared erosion and ulcers,hyperemia and edema,thus the rats were modeled successfully.In addition to the blank control group and the model group of normal feeding,the rest were treated with corresponding Chinese medicine respectively.XG were gavaged with mesalazine suspension and HG,ZG and LG were gavaged with different concentrations of mixed suspension of ShuangHong Qin YuTang decoction.After continual treatment for 2 weeks,we need observe and note the rats mental state,food consumption,coat color,faeces character and weight of rats in different periods and relying on these to carry on DAI score.At the end of the treatment,each rat of all the groups can only drank water for 24 hours.After abrosia 24 hours,we anesthetized rats with lidocaine,took the femoral arterial blood and detected the content of IL-17 by double antibody sandwich method,at the same time we dissected rats and took colon tissue of the heavier lesion site,which need paraffin embedding,section cutting and HE dyeing to help us test the expression of TLR2.Results: 1 general state: NG group of rats were in good conditions but others’ spirits were dropping and reacted slowly,loss of weight and mucous and blood were found in their thin stool.After gavage treatment,the therapy group had good changes which include energetic spirit,good appetite,weight gain and the normal feces in which mucous and purulent disappeared.Among all therapy groups,HG group changed most obviously and restored fastest,XG and ZG had no obvious difference,comparing to HG less poored but better than LG group.DAI score results shows: XG,HG,ZG,LG groups DAI scores were all lower than MG and the differences were statistically significant(P<0.05);HG was lower than XG and ZG,and the difference was statistically significant(P<0.05);LG group compared with XG and ZG,the DAI score was higher,but between XG to ZG there was no obvious statistically significance.(P<0.05);2 The colonic mucosa observation with naked eye and Pathology.The naked eye: the colonic epithelium mucosa of NG group rats was normal.MG rats colonic mucosa was the heaviest in hyperemia and edema and in glands’ damages and the most ulcers.HG rats colonic mucosa was smoother and had more new glands and the fewest ulcers than others’ groups.XG and ZG rats colonic mucosa were not smooth and had fewer ulcers than LG.LG group rats colonic mucosa were slight hyperemia and edema and had many ulcers.The observation of Pathology: NG group rats mucosa was smooth and the glands were normal.There are many ulcers,broken glands and inflammatory cells in MG group rats rough mucosa.HG group rats mucosa was more smoother and new glands.There were less inflammatory cells and ulcers in XG and ZG groups mucosa.LG group mucosa left more lesions and mild hyperemia and edema and there were some new glands around the ulcers;3 Index detection The comparison of content of IL-17: MG,XG,HG,ZG and LG the content of IL-17 all were higher than NG and the differences were statistically significant(P<0.05);XG,HG,ZG and LG the content of IL-17 all were lower than MG and the differences were statistically significant(P<0.05);XG,ZG and LG the content of IL-17 all were higher than HG and the differences were statistically significant(P<0.05);XG and ZG groups were lower than LG,the differences were statistically significant(P<0.05);XG group and ZG had no statistically significance(P>0.05).The content of TLR-2: the expression of TLR-2 in MG,XG,HG,ZG and LG all were higher than NG and the differences were statistically significant(P<0.05);the expression of TLR-2 in all were lower than MG and the differences were statistically significant(P<0.05);XG,ZG and LG the expression of TLR-2 all were higher than HG and the differences were statistically significant(P<0.05);XG and ZG group were lower than LG,the differences were statistically significant(P<0.05);XG group and ZG had no statistically significance(P>0.05).Conclusions:WengLianJieDu decoction can improve the ulcerative colitis rats general state,restore the damaged colonic mucous and reduce the score of DAI,and the efficacy changes with the different concentration of decoction.The mechanism of the Weng LianJieDu is that the decoction may reduce the expression of TLR-2 and the content of IL-17 in order to block inflammation,reduce the inflammatory cells infiltration,stimulate local tissue regeneration and heal the ulcers.更多还原