丁氏潰結(jié)灌腸液對DSS誘導(dǎo)的潰瘍性結(jié)腸炎大鼠腸道屏障作用的相關(guān)研究
發(fā)布時間：2018-08-03 07:13
【摘要】：目的:研究丁氏潰結(jié)灌腸液對葡聚糖硫酸鈉(Dextran Sulfate Sodium,DSS)誘導(dǎo)的潰瘍性結(jié)腸炎(Ulcerative Colitis,UC)大鼠的結(jié)腸組織中緊密連接蛋白(Occludin)、粘蛋白2(MUC2)和誘導(dǎo)型一氧化氮合酶(iNOS)、血清免疫球蛋白A(IgA)的影響,探討其可能的作用機(jī)制。方法:隨機(jī)挑選10只大鼠,歸入到空白對照組,其余的50只通過飲用DSS水溶液的方法建立大鼠UC模型。模型復(fù)制成功后,將參加造模的大鼠隨機(jī)的分為5組,即模型組、丁氏潰結(jié)灌腸液低劑量組、丁氏潰結(jié)灌腸液中劑量組、丁氏潰結(jié)灌腸液高劑量組、美沙拉秦灌腸液組。除空白對照組外,其余各組給予生理鹽水和相應(yīng)藥物連續(xù)灌腸14d。治療14d后采用腹主動脈取血的方法,將全部大鼠處死,并剖取結(jié)腸標(biāo)本。觀察大鼠的體重、大便性狀、便血情況及結(jié)腸組織的損傷變化并評分,評估疾病活動指數(shù)(Disease Activity Index,DAI),肉眼觀察結(jié)腸組織大體情況并評分,光鏡下觀察結(jié)腸組織病理損傷并評分,酶聯(lián)免疫吸附(ELISA)法測定血清IgA含量、蛋白質(zhì)印跡(Western Blot)法檢測結(jié)腸組織Occludin蛋白水平、實時熒光定量PCR(Real-Time PCR)檢測結(jié)腸組織MUC2mRNA、iNOS mRNA表達(dá)水平。結(jié)果:1、造模后大鼠精神萎靡、體重下降、大便溏稀伴有黏液膿血,DAI評分與空白對照組比較增高(P0.05)。灌腸治療后,丁氏潰結(jié)灌腸液組和美沙拉秦灌腸液組大鼠上述癥狀好轉(zhuǎn),DAI評分與模型組比較降低(P0.05)。其中丁氏潰結(jié)灌腸液低劑量組情況不及其他用藥祖,但也好于模型組。2、肉眼觀察,模型組大鼠結(jié)腸粘膜出現(xiàn)不完整損傷,明顯充血、水腫,并伴有潰瘍形成,與空白對照組比較大體評分增高(P0.05)。丁氏潰結(jié)灌腸液各組及美沙拉秦灌腸液組大鼠結(jié)腸粘膜損傷后得以修復(fù),僅出現(xiàn)了充血、水腫、輕度糜爛,未見到明顯的潰瘍形成,與模型組比較大體評分下降(P0.05)。3、光鏡下觀察,模型組大鼠結(jié)腸粘膜損傷后出現(xiàn)壞死脫落,伴發(fā)不同程度的潰瘍灶,大量炎癥性細(xì)胞浸潤,粘膜腺體被破壞,排列稀疏紊亂,甚至缺如,并有程度不一的擴(kuò)張,與空白對照組大鼠比較病理評分顯著增加(P0.01)。丁氏潰結(jié)灌腸液各組和美沙拉秦灌腸液組大鼠結(jié)腸粘膜未有明顯的潰瘍形成,粘膜水腫充血和炎癥細(xì)胞浸潤減少,與模型組比較病理評分明顯的降低(P0.01)。4、模型組大鼠血清IgA含量、結(jié)腸組織Occludin蛋白表達(dá)水平,與空白對照組相比顯著下降(P0.05),經(jīng)丁氏潰結(jié)灌腸液及美沙拉秦灌腸液治療后,各組血清IgA含量、結(jié)腸組織Occludin蛋白表達(dá)水平升高,與模型組比較有統(tǒng)計學(xué)差異(P0.05)。5、模型組大鼠結(jié)腸組織iNOSmRNA的表達(dá)水平,與空白對照組比較明顯的升高(P0.01),經(jīng)丁氏潰結(jié)灌腸液和美沙拉秦灌腸液治療后,各組iNOSmRNA的表達(dá)呈下降趨勢,與模型組比較有統(tǒng)計學(xué)意義(P0.05)。模型組大鼠結(jié)腸組織MUC2 mRNA的表達(dá)水平,與空白對照組比較明顯的減低(P0.01),經(jīng)丁氏潰結(jié)灌腸液和美沙拉秦灌腸液治療后,MUC2mRNA的表達(dá)水平顯示升高,與模型組比較有統(tǒng)計學(xué)差異(P0.05)。在mRNA表達(dá)水平上的比較,丁氏潰結(jié)灌腸液中高劑量組與美沙拉秦灌腸液組之間無明顯差異。結(jié)論:1、丁氏潰結(jié)灌腸液對DSS誘導(dǎo)的UC有一定治療作用,可有效減輕其大便不成形、便血、體重下降等癥狀,使腸粘膜組織炎癥好轉(zhuǎn),降低DAI評分和大體、病理評分。2、對于DSS誘導(dǎo)的UC腸道損傷,丁氏潰結(jié)灌腸液可顯著升高Occludin蛋白、MUC2 mRNA表達(dá)水平,增高血清IgA含量,降低iNOSmRNA表達(dá)水平,從腸道機(jī)械屏障、免疫屏障及化學(xué)屏障方面,減輕大鼠腸粘膜炎癥損傷。
[Abstract]:Objective: To study the effects of Ding's ulceration enema on the colonic tissue, mucin 2 (MUC2) and inducible nitric oxide synthase (iNOS) and serum immunoglobulin A (IgA) in the colonic tissue of rats with Dextran Sulfate Sodium (DSS) induced ulcerative colitis (UC). Mechanism. Methods: 10 rats were randomly selected and returned to the blank control group. The other 50 rats were established by drinking DSS water solution. After the model replication was successful, the model rats were randomly divided into 5 groups, that is, the model group, the low dose group of Ding's ulceration enema, the middle dose group of the Ding's ulcerative enema solution, and the Ding's ulceration enema. In the high dose group, the Mesalazine Enemas group. Except for the blank control group, the other groups were given the normal saline and the corresponding drug continuous enema 14D. for the treatment of 14d after the treatment of the abdominal aorta. All rats were killed and the colon specimens were dissected. The body weight, stool, blood condition and colonic tissue injury of the rats were observed and evaluated. The disease activity index (Disease Activity Index, DAI) was evaluated and the colonic tissue was observed and graded by naked eyes. The pathological damage of colon tissue was observed under light microscope and the serum IgA content was measured by enzyme linked immunosorbent assay (ELISA). The level of Occludin protein in colon tissue was detected by the method of Western blot (Western Blot), and real time fluorescent quantitative PCR (Real) -Time PCR) detected the expression level of MUC2mRNA and iNOS mRNA in colonic tissue. Results: 1, the rats were mentally depressed, weight loss, loose stool with mucous blood, DAI score and blank control group increased (P0.05). After enema treatment, the symptoms of DINK and clyster group and mesalazine enema group were improved, DAI score and model Group comparison decreased (P0.05). Among them, the low dose group of Ding's ulcerative enema was inferior to that of other drugs, but it was also better than.2 in the model group. The colonic mucosa of the model group had incomplete injury, obvious congestion, edema, and the formation of ulceration, and the gross score was higher in the blank control group (P0.05). The colonic mucosa of the Mesalazine Enemas group was repaired. Only the hyperemia, edema, mild erosion, no obvious ulcers were found, and the gross scores of the model group decreased (P0.05).3. Under the light microscope, the colonic mucosa of the model rats showed necrosis and shedding, accompanied by varying degrees of ulcer, and a large number of inflammatory lesions. Cell infiltration, mucous glands were destroyed, the arrangement of a sparse disorder, or even the absence of a degree of expansion, compared with the blank control group, the pathological score of the rats was significantly increased (P0.01). There was no obvious ulcer formation in the colonic mucosa of the group of Ding's ulcerative enema and the mesalazine enema group, and the congestion of mucous edema and the infiltration of inflammatory cells. Less, compared with the model group, the pathological score was significantly lower (P0.01).4, the serum IgA content of the model group and the expression level of Occludin protein in the colon tissue were significantly lower than that in the blank control group (P0.05). The serum IgA content and the expression of Occludin protein in the colon tissues were increased after the treatment of the Ding ulceration enema and Mesalazine Enemas. Compared with the model group, there was a statistically significant difference (P0.05).5. The expression level of iNOSmRNA in the colon tissue of the model group was significantly higher than that in the blank control group (P0.01). After the treatment of the Ding ulceration enema and the mesalazine enema, the expression of iNOSmRNA decreased in each group, and was statistically significant (P0.05) compared with the model group (P0.05). The model group was larger than the model group. The expression level of MUC2 mRNA in the rat colonic tissue was significantly lower than that in the blank control group (P0.01). The expression level of MUC2mRNA increased after the treatment of Ding's ulcerative enema and mesalazine enema, compared with the model group (P0.05). In the mRNA expression level, the high dose group and the beauty in the Ding's ulcerative enema solution were compared. There was no obvious difference between the Shalin enema group. Conclusion: 1, Ding's ulcerative enema has a certain therapeutic effect on DSS induced UC. It can effectively alleviate the symptoms of unformed bowel, blood, weight loss and so on, improve the inflammation of the intestinal mucosa, reduce the DAI score and gross, the pathological score of.2, DSS induced UC intestinal injury, and Ding's ulceration enema The solution could significantly increase the expression of Occludin protein, MUC2 mRNA, increase the content of serum IgA, reduce the level of iNOSmRNA expression, and reduce the injury of intestinal mucositis in rats, from the intestinal mechanical barrier, immune barrier and chemical barrier.
【學(xué)位授予單位】：南京中醫(yī)藥大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R285.5
[Abstract]:Objective: To study the effects of Ding's ulceration enema on the colonic tissue, mucin 2 (MUC2) and inducible nitric oxide synthase (iNOS) and serum immunoglobulin A (IgA) in the colonic tissue of rats with Dextran Sulfate Sodium (DSS) induced ulcerative colitis (UC). Mechanism. Methods: 10 rats were randomly selected and returned to the blank control group. The other 50 rats were established by drinking DSS water solution. After the model replication was successful, the model rats were randomly divided into 5 groups, that is, the model group, the low dose group of Ding's ulceration enema, the middle dose group of the Ding's ulcerative enema solution, and the Ding's ulceration enema. In the high dose group, the Mesalazine Enemas group. Except for the blank control group, the other groups were given the normal saline and the corresponding drug continuous enema 14D. for the treatment of 14d after the treatment of the abdominal aorta. All rats were killed and the colon specimens were dissected. The body weight, stool, blood condition and colonic tissue injury of the rats were observed and evaluated. The disease activity index (Disease Activity Index, DAI) was evaluated and the colonic tissue was observed and graded by naked eyes. The pathological damage of colon tissue was observed under light microscope and the serum IgA content was measured by enzyme linked immunosorbent assay (ELISA). The level of Occludin protein in colon tissue was detected by the method of Western blot (Western Blot), and real time fluorescent quantitative PCR (Real) -Time PCR) detected the expression level of MUC2mRNA and iNOS mRNA in colonic tissue. Results: 1, the rats were mentally depressed, weight loss, loose stool with mucous blood, DAI score and blank control group increased (P0.05). After enema treatment, the symptoms of DINK and clyster group and mesalazine enema group were improved, DAI score and model Group comparison decreased (P0.05). Among them, the low dose group of Ding's ulcerative enema was inferior to that of other drugs, but it was also better than.2 in the model group. The colonic mucosa of the model group had incomplete injury, obvious congestion, edema, and the formation of ulceration, and the gross score was higher in the blank control group (P0.05). The colonic mucosa of the Mesalazine Enemas group was repaired. Only the hyperemia, edema, mild erosion, no obvious ulcers were found, and the gross scores of the model group decreased (P0.05).3. Under the light microscope, the colonic mucosa of the model rats showed necrosis and shedding, accompanied by varying degrees of ulcer, and a large number of inflammatory lesions. Cell infiltration, mucous glands were destroyed, the arrangement of a sparse disorder, or even the absence of a degree of expansion, compared with the blank control group, the pathological score of the rats was significantly increased (P0.01). There was no obvious ulcer formation in the colonic mucosa of the group of Ding's ulcerative enema and the mesalazine enema group, and the congestion of mucous edema and the infiltration of inflammatory cells. Less, compared with the model group, the pathological score was significantly lower (P0.01).4, the serum IgA content of the model group and the expression level of Occludin protein in the colon tissue were significantly lower than that in the blank control group (P0.05). The serum IgA content and the expression of Occludin protein in the colon tissues were increased after the treatment of the Ding ulceration enema and Mesalazine Enemas. Compared with the model group, there was a statistically significant difference (P0.05).5. The expression level of iNOSmRNA in the colon tissue of the model group was significantly higher than that in the blank control group (P0.01). After the treatment of the Ding ulceration enema and the mesalazine enema, the expression of iNOSmRNA decreased in each group, and was statistically significant (P0.05) compared with the model group (P0.05). The model group was larger than the model group. The expression level of MUC2 mRNA in the rat colonic tissue was significantly lower than that in the blank control group (P0.01). The expression level of MUC2mRNA increased after the treatment of Ding's ulcerative enema and mesalazine enema, compared with the model group (P0.05). In the mRNA expression level, the high dose group and the beauty in the Ding's ulcerative enema solution were compared. There was no obvious difference between the Shalin enema group. Conclusion: 1, Ding's ulcerative enema has a certain therapeutic effect on DSS induced UC. It can effectively alleviate the symptoms of unformed bowel, blood, weight loss and so on, improve the inflammation of the intestinal mucosa, reduce the DAI score and gross, the pathological score of.2, DSS induced UC intestinal injury, and Ding's ulceration enema The solution could significantly increase the expression of Occludin protein, MUC2 mRNA, increase the content of serum IgA, reduce the level of iNOSmRNA expression, and reduce the injury of intestinal mucositis in rats, from the intestinal mechanical barrier, immune barrier and chemical barrier.
【學(xué)位授予單位】：南京中醫(yī)藥大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R285.5
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