同型半胱氨酸對(duì)實(shí)驗(yàn)性結(jié)腸炎大鼠腸纖維化的影響及機(jī)制研究
發(fā)布時(shí)間：2019-04-18 17:36
【摘要】：背景：克羅恩�。–rohn’s disease, CD）是一種病因未明的慢性非特異性腸道肉芽腫性疾病，腸纖維性狹窄是CD常見(jiàn)并發(fā)癥之一，目前尚無(wú)有效的治療藥物，手術(shù)是最終治療手段，術(shù)后仍有約70%患者再次發(fā)生狹窄，，反復(fù)發(fā)生的腸狹窄嚴(yán)重影響患者的生活質(zhì)量。同型半胱氨酸（homocysteine, Hcy）是一種含硫氨基酸，是甲硫氨酸去甲基化過(guò)程的中間產(chǎn)物，研究顯示Hcy與心、肝、腎、肺等多種組織器官纖維化過(guò)程密切相關(guān)，多個(gè)研究發(fā)現(xiàn)在CD患者血漿和腸黏膜中Hcy水平明顯增高，但對(duì)于Hcy是否參與CD腸纖維化的病理過(guò)程目前國(guó)內(nèi)外尚無(wú)報(bào)道。因此，探討Hcy對(duì)CD腸纖維化的影響及其可能機(jī)制對(duì)于CD腸纖維化狹窄的防治具有重要的理論和實(shí)際意義。 目的：探討同型半胱氨酸（Hcy）對(duì)實(shí)驗(yàn)性結(jié)腸炎大鼠腸纖維化的影響及其可能機(jī)制。 方法：1.采用2,4,6-三硝酸苯磺酸(TNBS)灌腸制備大鼠CD模型。對(duì)照組以等體積的生理鹽水（NS）灌腸；2.SD大鼠隨機(jī)分為四組（n=8）：正常對(duì)照組（NS灌腸+皮下注射NS）、正常對(duì)照+Hcy注射組（NS灌腸+皮下注射Hcy）、TNBS模型對(duì)照組（TNBS/乙醇灌腸+皮下注射NS）、TNBS模型+Hcy注射組（TNBS/乙醇灌腸+皮下注射Hcy）；3.記錄大鼠體重變化并進(jìn)行疾病活動(dòng)指數(shù)評(píng)分（DAI），實(shí)驗(yàn)結(jié)束后取部分結(jié)腸行大體損傷評(píng)分（CMDI），HE染色和Masson染色行結(jié)腸組織學(xué)損傷評(píng)分（HI）及纖維化評(píng)分；4.高效液相熒光檢測(cè)法（HPLC-FD）檢測(cè)大鼠血漿及結(jié)腸組織Hcy水平，結(jié)腸勻漿檢測(cè)MDA含量及SOD、GSH-PX、MPO活性，酶聯(lián)免疫法（ELISA）測(cè)定結(jié)腸勻漿IL-1β、IL-6、TNF-α、TGF-β1、CTGF、MMP-2、MMP-9、Ⅰ型和Ⅲ型膠原水平，逆轉(zhuǎn)錄聚合酶鏈反應(yīng)（RT-PCR）檢測(cè)TGF-β1、MMP-2、MMP-9和TIMP-1mRNA表達(dá)水平，免疫組化SP法檢測(cè)結(jié)腸組織MMP-2和MMP-9蛋白表達(dá)水平。 結(jié)果：與正常對(duì)照組比較，TNBS結(jié)腸炎大鼠體重明顯減輕，出現(xiàn)不同程度的肉眼血便，DAI評(píng)分增高（p0.05）；CMDI、HI及纖維化評(píng)分均顯著增高。結(jié)腸組織MPO活性增高，SOD、GSH-PX活性降低，MDA、IL-1β、IL-6、TNF-α、TGF-β1、CTGF、MMP-2、MMP-9、Ⅰ型和Ⅲ型膠原水平增高，MMP-2和MMP-9蛋白表達(dá)增強(qiáng)，TGF-β1、MMP-2、MMP-9和TIMP-1mRNA表達(dá)增強(qiáng)（p0.05）。與TNBS模型對(duì)照組比較，皮下注射Hcy的結(jié)腸炎大鼠血漿及結(jié)腸組織Hcy水平明顯增高，DAI、CMDI、HI及纖維化評(píng)分增高，結(jié)腸組織SOD、GSH-PX活性降低，MPO活性、MDA、IL-1β、IL-6、TNF-α、TGF-β1、CTGF、MMP-2、MMP-9、Ⅰ型和Ⅲ型膠原水平均明顯增高（p0.05）。 結(jié)論：Hcy可加重實(shí)驗(yàn)性結(jié)腸炎大鼠腸纖維化，機(jī)制可能與增加結(jié)腸氧化及炎癥損傷、促進(jìn)TGF-β1、CTGF等促纖維化因子表達(dá)以及通過(guò)影響MMPs/TIMPs平衡導(dǎo)致細(xì)胞外基質(zhì)代謝失衡有關(guān)。
[Abstract]:Background: Crohn's disease (Crohn's disease, CD) is a chronic nonspecific intestinal granulomatous disease with unknown etiology. Intestinal fibrous stenosis is one of the common complications of CD. About 70% of the patients suffered from recurrent stricture, and recurrent intestinal stenosis seriously affected the quality of life of the patients. Homocysteine (homocysteine, Hcy) is a sulfur-containing amino acid, which is the intermediate product of methionine demethylation. Studies have shown that Hcy is closely related to fibrosis of heart, liver, kidney, lung and other tissues and organs. Many studies have found that the level of Hcy in plasma and intestinal mucosa of patients with CD is significantly increased, but there is no report on whether Hcy is involved in the pathological process of CD intestinal fibrosis at home and abroad. Therefore, it is of great theoretical and practical significance to explore the effect of Hcy on intestinal fibrosis in CD and its possible mechanism for the prevention and treatment of intestinal fibrosis stenosis in CD. Objective: to investigate the effect of homocysteine (Hcy) on intestinal fibrosis in rats with experimental colitis and its possible mechanism. Methods: 1. The CD model of rats was established by (TNBS) enema with 2,4, 6-trinitrate benzenesulfonic acid. The control group was enema with the same volume of saline (NS). 2.SD rats were randomly divided into four groups (n = 8): normal control group (NS enema subcutaneously injected NS), normal control Hcy injection group; NS enema subcutaneously injected Hcy), TNBS model control group; TNBS/ ethanol enema subcutaneously injected NS),). TNBS model Hcy injection group (TNBS/ ethanol enema subcutaneously injected Hcy); 3. The body weight of the rats was recorded and the disease activity index score (DAI),) was carried out. After the end of the (DAI), experiment, partial colon was taken for gross injury score (CMDI), HE staining and Masson staining for colon histology injury score (HI) and fibrosis score; 4. High performance liquid phase fluorescence assay (HPLC-FD) was used to detect the level of Hcy in plasma and colon tissue of rats. The content of MDA and the activity of SOD,GSH-PX,MPO in colonic homogenate were measured. The levels of IL-1 尾 and IL-6,TNF- 偽 in colonic homogenate were determined by enzyme linked immunosorbent assay (ELISA). The levels of TGF- 尾 1, CTGF,MMP-2,MMP-9, type 鈪
本文編號(hào)：2460209
[Abstract]:Background: Crohn's disease (Crohn's disease, CD) is a chronic nonspecific intestinal granulomatous disease with unknown etiology. Intestinal fibrous stenosis is one of the common complications of CD. About 70% of the patients suffered from recurrent stricture, and recurrent intestinal stenosis seriously affected the quality of life of the patients. Homocysteine (homocysteine, Hcy) is a sulfur-containing amino acid, which is the intermediate product of methionine demethylation. Studies have shown that Hcy is closely related to fibrosis of heart, liver, kidney, lung and other tissues and organs. Many studies have found that the level of Hcy in plasma and intestinal mucosa of patients with CD is significantly increased, but there is no report on whether Hcy is involved in the pathological process of CD intestinal fibrosis at home and abroad. Therefore, it is of great theoretical and practical significance to explore the effect of Hcy on intestinal fibrosis in CD and its possible mechanism for the prevention and treatment of intestinal fibrosis stenosis in CD. Objective: to investigate the effect of homocysteine (Hcy) on intestinal fibrosis in rats with experimental colitis and its possible mechanism. Methods: 1. The CD model of rats was established by (TNBS) enema with 2,4, 6-trinitrate benzenesulfonic acid. The control group was enema with the same volume of saline (NS). 2.SD rats were randomly divided into four groups (n = 8): normal control group (NS enema subcutaneously injected NS), normal control Hcy injection group; NS enema subcutaneously injected Hcy), TNBS model control group; TNBS/ ethanol enema subcutaneously injected NS),). TNBS model Hcy injection group (TNBS/ ethanol enema subcutaneously injected Hcy); 3. The body weight of the rats was recorded and the disease activity index score (DAI),) was carried out. After the end of the (DAI), experiment, partial colon was taken for gross injury score (CMDI), HE staining and Masson staining for colon histology injury score (HI) and fibrosis score; 4. High performance liquid phase fluorescence assay (HPLC-FD) was used to detect the level of Hcy in plasma and colon tissue of rats. The content of MDA and the activity of SOD,GSH-PX,MPO in colonic homogenate were measured. The levels of IL-1 尾 and IL-6,TNF- 偽 in colonic homogenate were determined by enzyme linked immunosorbent assay (ELISA). The levels of TGF- 尾 1, CTGF,MMP-2,MMP-9, type 鈪
本文編號(hào)：2460209
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