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  本文關(guān)鍵詞： 梗阻性黃疸 腸粘膜 瘦素 瘦素受體 增殖　出處：《濱州醫(yī)學院》2015年碩士論文　論文類型：學位論文

【摘要】：目的 建立可復性梗阻性黃疸大鼠模型,探討瘦素對可復性梗阻性黃疸大鼠腸粘膜增殖的影響及機制方法用”支撐管膽管結(jié)扎加支撐管拔除法”建立可復性梗阻性黃疸大鼠模型及膽管結(jié)扎法建立梗阻性黃疸模型。選用成年健康雄性Wistar大鼠64只,隨機分成四組,可復性梗阻性黃疸組(ROJ)、梗阻性黃疸組(OJ)、可復性梗阻性黃疸+瘦素組(ROJ+LP)、梗阻性黃疸+瘦素組(OJ+LP)。ROJ組大鼠術(shù)后第5天,自左下腹部切口打開皮下輕輕拔出PICC導管完成膽管再通。瘦素組動物自術(shù)后第1天腹腔注射重組瘦素10ug/kg,bid,其余組動物相應(yīng)時間點腹腔注射等量生理鹽水作為對照。四組大鼠分別于術(shù)后5天和8天各處死8只,分別標記為ROJ5組、ROJ8組、OJ5組和OJ8組、ROJ5+LP組、ROJ8+LP組、OJ5+LP組和OJ8+LP組。放血法處死,剖腹探查觀察膽管梗阻情況,并取標本。光學顯微鏡下HE切片觀察腸黏膜結(jié)構(gòu)的形態(tài)學變化,測量空腸黏膜上皮的絨毛高度及隱窩深度；免疫組化檢測腸黏膜瘦素、受體以及Ki67抗原的表達情況。結(jié)果1.一般情況：ROJ組、OJ組及OJ+LP組分別于術(shù)后第4,6,2天各死亡1只,ROJ+LP組分別于術(shù)后第3、5天各死亡1只,死因為術(shù)后腸梗阻、腸壞死及腹腔積液。2.腸粘膜組織學觀察：ROJ5組腸黏膜以非特異性炎細胞浸潤、間質(zhì)的充血水腫的炎癥改變?yōu)橹?腸黏膜萎縮變薄,絨毛的數(shù)量及密度下降,絨毛高度降低,間隙增大不連續(xù),有凹陷存在,并有部分絨毛的脫落,隱窩深度變淺,腺體排列稀疏；ROJ8組腸絨毛結(jié)構(gòu)較完整,上皮細胞基本呈高柱狀,無明顯間質(zhì)水腫及中性粒細胞的浸潤,基本維持正常的腸粘膜結(jié)構(gòu)。0J5組腸粘膜形態(tài)基本同ROJ5組,0J8組的腸粘膜形態(tài)相比0J5組嚴重紊亂,炎細胞浸潤、間質(zhì)的充血水腫多見。ROJ5+LP組較ROJ5組腸粘膜損害程度差,有輕度絨毛排列紊亂,ROJ8+LP組腸粘膜形態(tài)未見明顯異常表現(xiàn)。OJ5+LP組腸粘膜形態(tài)基本同ROJ5+LP組。OJ8+LP組仍有腸粘膜形態(tài)學異常改變,但程度較0J8組減輕。3.腸粘膜絨毛高度和隱窩深度：ROJ5組的絨毛高度明顯低于ROJ8組,P0.01；0J5組的絨毛高度高于0J8組,P=0.05；ROJ5+LP組的絨毛高度明顯低于ROJ8+LP組,P0.05；OJ5+LP組的絨毛高度高于OJ8+LP組,P0.05。ROJ8組明顯低于ROJ8+LP組,P0.05；0J8組的絨毛高度明顯低于OJ8+LP組,P0.05,ROJ5組的絨毛高度明顯低于ROJ5+LP組,P0.01；0J5組的絨毛高度明顯低于OJ5+LP組,P0.05。ROJ8組的絨毛高度明顯高于0J8組,P0.01,ROJ8+LP組的絨毛高度明顯高于OJ8+LP組,P0.01。ROJ5組的隱窩深度低于ROJ8組；0J5組的隱窩深度高于0J8組；ROJ5+LP組的隱窩深度低于ROJ8+LP組;OJ5+LP組的隱窩深度高于0J8+LP組。ROJ8+LP組的腸粘膜隱窩深度高于ROJ8組,P0.01。ROJ5+LP組的腸粘膜隱窩深度明顯高于ROJ5組,P0.01；OJ8+LP組的腸粘膜隱窩深度明顯高于OJ8組,P0.0l,OJ5+LP組的腸粘膜隱窩深度明顯高于0J5組,P0.01。ROJ8組的隱窩深度明顯高于0J8,P0.01,ROJ+LP組的絨毛高度明顯高于OJ+LP組,P0.05。4.腸粘膜細胞增殖變化：腸黏膜增殖陽性細胞主要集中于腸粘膜隱窩處。ROJ5組的腸粘膜增殖指數(shù)明顯低于ROJ8組；0J5組的腸粘膜增殖指數(shù)高于OJ8組,ROJ5+LP組的腸粘膜增殖指數(shù)低于ROJ8+LP組,P0.05;OJ5+LP組的腸粘膜增殖指數(shù)高于0J8+LP組。術(shù)后第8天,ROJ8組的腸粘膜增殖指數(shù)明顯低于ROJ8+LP組,P0.01；OJ8組的腸粘膜增殖指數(shù)明顯低于OJ8+LP組,P0.01;OJ5組的腸粘膜增殖指數(shù)明顯低于OJ5+LP組,P0.01；組內(nèi)比較：ROJ8+LP的腸粘膜增殖指數(shù)明顯高于ROJ5+LP組,P0.05,差異有統(tǒng)計學意義。5.腸粘膜瘦素蛋白及其瘦體變化：瘦素及受體的陽性產(chǎn)物主要定位于胞漿中,呈棕黃色的顆粒,細胞間質(zhì)也有少量著色。陽性細胞主要分布于腸黏膜上皮、隱窩細胞、腸腺細胞及固有層細胞。Leptin組及對照組腸黏膜中均有瘦素表達,但表達程度不一,高表達主要見于Leptin組,對照組腸黏膜組織表達相對較低或不表達。ROJ5組的0B蛋白表達明顯ROJ8組；0J5組的0B蛋白表達高于0J8組,ROJ5+LP組的0B蛋白表達低于ROJ8+LP組;OJ5+LP組的0B蛋白表達高于OJ8+LP組。術(shù)后第8天,ROJ8組的腸粘膜0B蛋白表達明顯低于ROJ+LP組,P0.01；OJ8組的腸粘膜0B蛋白表達明顯低于OJ8+LP組,P0.05。同時ROJ8+LP組的腸粘膜0B蛋白表達明顯高于OJ8+LP組,P0.05；術(shù)后第5天,ROJ5組的腸粘膜0B蛋白表達明顯低于ROJ5+LP組,P=0.01；0.15組的腸粘膜0B蛋白表達明顯低于0J5+LP組,P0.01。ROJ5組的瘦素受體表達低于ROJ8組；0J5組的瘦素受體表達高于0J8組,ROJ5+LP組的瘦素受體表達低于ROJ8+LP組;OJ5+LP組的瘦素受體表達高于OJ8+LP組。術(shù)后第8天,ROJ8組的瘦素受體表達明顯低于ROJ8+LP組,P0.01;OJ8組的瘦素受體表達明顯低于OJ8+LP組,P0.05；術(shù)后第5天,ROJ5組的瘦素受體表達明顯低于ROJ5+LP組,P0.01；OJ5組的瘦素受體表達明顯低于OJ5+LP組,P0.01。結(jié)論1.”支撐管膽管結(jié)扎加支撐管拔除法”制作的黃疸模型操作簡便,解除黃疸不需二次開腹手術(shù),創(chuàng)傷小,成功率、死亡率低,是一種穩(wěn)定可靠的的可復性梗阻性黃疸大鼠模型.2梗阻性黃疸可致腸粘膜增殖能力的下降,主要體現(xiàn)在腸粘膜增殖指數(shù)Ki67的下降,絨毛高度和隱窩深度的降低。黃疸解除后各項指標能夠得到改善。3外源性瘦素對梗阻性黃疸有促進腸粘膜細胞增殖的保護作用,在黃疸解除后,其作用仍能持續(xù)發(fā)揮。瘦素可加快腸粘膜增殖能力損害的修復。4外源性瘦素對腸粘膜細胞增殖作用的機理可能通過促進腸粘膜瘦素含量的增加及其受體表達的增強來實現(xiàn)。
[Abstract]:Objective to establish a complex rat model of obstructive jaundice, to explore the effect of method of reversible obstructive jaundice rat intestinal mucosal proliferation and the mechanism of "supporting tube with the support tube division of bile duct ligation to establish established obstructive jaundice model of rat model of obstructive jaundice and bile duct ligation of leptin. Healthy adult male Wistar 64 rats were randomly divided into four groups, reversible obstructive jaundice group (ROJ), obstructive jaundice group (OJ), reversible obstructive jaundice + leptin group (ROJ+LP), obstructive jaundice group (OJ+LP) and leptin fifth days after surgery of.ROJ group from the left lower abdominal incision, open skin gently pull out the PICC catheter to complete bile duct recanalization. Leptin group from animal first days after intraperitoneal injection of recombinant leptin 10ug/kg, bid, other animal group at corresponding time intraperitoneal injection of saline as a control group. The four groups of rats were sacrificed at 5 days and 8 days respectively 8 animals were killed, were labeled as ROJ5 group, ROJ8 group, OJ5 group and OJ8 group, ROJ5+LP group, ROJ8+LP group, OJ5+LP group and OJ8+LP group. Bloodletting method executed, laparotomy observation of bile duct obstruction, and the specimens. The morphological changes of intestinal mucosa structure of HE were observed under optical microscope, villus height and crypt depth the measurement of jejunal mucosa epithelium; immunohistochemical detection of intestinal mucosal leptin receptor and Ki67 antigen expression. Results 1. general condition: ROJ group, OJ group and OJ+LP group respectively at 4,6,2 days after operation the death of 1, ROJ+LP group was 3,5 days after the death of 1, cause of death observation of postoperative intestinal obstruction, intestinal necrosis and ascites.2. intestinal mucosa mucosa of intestinal ROJ5 group by non-specific inflammatory cell infiltration, inflammatory interstitial hyperemia and edema changes, intestinal mucosa atrophy thinning, the number and density of villus decreased, villus height decreased, the gap increases Is not continuous, depression exists, and shedding of the villus and crypt depth becomes shallow, glands are sparse; in group ROJ8, the intestinal villus structure is complete, the epithelial cells were columnar, no obvious interstitial edema and infiltration of neutrophils, maintain normal intestinal mucosa structure of intestinal mucosa of.0J5 group basically the same as ROJ5 group, morphology of intestinal mucosa in 0J8 group compared with 0J5 group of serious disorders, inflammatory cell infiltration, interstitial edema is more common in.ROJ5+LP group than in ROJ5 group of intestinal mucosa damage, mild villous derangement, ROJ8+LP group of intestinal mucosa of.OJ5+LP group showed no obvious abnormal intestinal mucosa morphology similar to that of ROJ5+LP group.OJ8+LP there were still some abnormal changes of intestinal mucosa morphology, but compared with the 0J8 group.3., intestinal villus height and crypt depth: ROJ5 group of villus height was significantly lower than ROJ8 group, P0.01 0J5 group; the villus height higher than that of group 0J8, P=0.05; ROJ 5+LP group of villus height was significantly lower than ROJ8+LP group, P0.05 OJ5+LP group; the villus height is higher than OJ8+LP group, P0.05.ROJ8 group was significantly lower than ROJ8+LP group, P0.05 0J8 group; the villus height was significantly lower than OJ8+LP group, P0.05 ROJ5 group, the villus height was significantly lower than ROJ5+LP group, P0.01 0J5 group; the villus height was significantly lower than OJ5+LP group. P0.05.ROJ8 group of villus height was significantly higher than 0J8 group, P0.01 ROJ8+LP group, the villus height was significantly higher than OJ8+LP group, P0.01.ROJ5 group, the crypt depth was lower than that of ROJ8 group; the crypt depth of 0J5 group was higher than that of 0J8 group; ROJ5+LP group of the crypt depth is lower than ROJ8+LP group; intestinal crypt depth of crypt depth in OJ5+LP group was higher than that of 0J8+LP group and.ROJ8+LP group higher than ROJ8 group, intestinal crypt depth in P0.01.ROJ5+LP group was significantly higher than ROJ5 group, P0.01; intestinal crypt depth in OJ8+LP group was significantly higher than OJ8 group, P0.0l OJ5+LP group, intestinal crypt depth The degree was significantly higher than 0J5 group, the crypt depth of P0.01.ROJ8 group was significantly higher than 0J8, P0.01, ROJ+LP group of villus height was significantly higher than OJ+LP group, the proliferation of P0.05.4. cells in the intestinal mucosa of intestinal mucosal changes: the proliferation of positive cells mainly in the intestinal crypt at.ROJ5 group intestinal mucosal proliferation index was significantly lower than that of ROJ8 group; intestinal mucosal proliferation index of 0J5 group higher than OJ8 group, intestinal mucosal proliferation index of ROJ5+LP was lower than that of group ROJ8+LP, P0.05; intestinal mucosal proliferation index of OJ5+LP group was higher than that of 0J8+LP group. After eighth days, intestinal mucosal proliferation index of ROJ8 group was significantly lower than that of group ROJ8+LP, P0.01; intestinal mucosal proliferation index in OJ8 group was significantly lower than that of OJ8+LP group, P0.01; intestinal mucosa the proliferation index of OJ5 group was significantly lower than that of group OJ5+LP, P0.01; group comparison: intestinal mucosal proliferation index of ROJ8+LP was significantly higher than that in group ROJ5+LP, P0.05, there was a significant difference between the.5. of intestinal mucosa leptin and leptin Change: the positive products of leptin and receptor mainly located in the cytoplasm, brownish yellow granules, interstitial cells also have a small amount of staining. Positive cells were mainly distributed in the intestinal epithelium, crypt cells, both leptin expression of intestinal gland cells and lamina propria cells in.Leptin group and control group in the intestinal mucosa, but the expression level a high expression mainly in the Leptin control group, the expression is relatively low or no expression of 0B protein in.ROJ5 group significantly ROJ8 group intestinal mucosa; expression of 0B protein in 0J5 group was higher than that in group 0J8, the expression of 0B protein in ROJ5+LP group was lower than that of ROJ8+LP group; OJ5+LP group the expression of 0B protein is higher than that of OJ8+LP group for eighth days. After operation, the expression of 0B protein in the intestinal mucosa of ROJ8 group was significantly lower than ROJ+LP group, the expression of 0B protein P0.01; intestinal mucosa of OJ8 group was significantly lower than that in group OJ8+LP, P0.05. and 0B in intestinal mucosa protein expression in ROJ8+LP group was significantly higher than that of OJ8+LP group, P0.05 fifth days after operation, The expression of 0B protein in intestinal mucosa of ROJ5 group was significantly lower than that of group ROJ5+LP, P=0.01; the expression of 0B protein in intestinal mucosa of the 0.15 group was significantly lower than 0J5+LP group, P0.01.ROJ5 group was lower than that in ROJ8 group the expression of leptin receptor; leptin receptor in 0J5 group than in the 0J8 group, the expression of leptin receptor in ROJ5+LP group than in the ROJ8+LP group; OJ5+LP group the expression of leptin receptor higher than that of OJ8+LP group. After eighth days, the expression of leptin receptor in ROJ8 group was significantly lower than that of ROJ8+LP group, P0.01 group; OJ8 expression of leptin receptor was significantly lower in group OJ8+LP, P0.05; fifth days after operation, ROJ5 group the expression of leptin receptor was significantly lower than that of ROJ5 group +LP, P0.01; group OJ5 expression of leptin receptor was lower than that in OJ5+LP group conclusion P0.01., 1. support tube bile duct ligation plus support tube division "produced by the jaundice model is simple, do not need to relieve jaundice two times open surgery, trauma, success rate, low mortality rate, is a stable and reliable. Decline of reversible obstructive jaundice model of obstructive jaundice can induce.2 rat intestinal mucosal proliferation, decline mainly reflected in the intestinal mucosa Ki67 proliferation index decreased, villus height and crypt depth. Jaundice after the termination of the indicators can be improved.3 exogenous leptin has a protective effect to promote the proliferation of cells in the intestinal mucosa of obstruction in jaundice, jaundice after the termination of its function still can continue to play. The mechanism of leptin.4 repair exogenous leptin can accelerate the proliferation of intestinal mucosal damage of intestinal mucosal cell proliferation may be achieved through the promotion of intestinal mucosa leptin content increased with the enhancement of expression and its receptor.

【學位授予單位】：濱州醫(yī)學院
【學位級別】：碩士
【學位授予年份】：2015
【分類號】：R657.3

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