外源性瘦素对可复性梗阻性黄疸大鼠肠粘膜增殖的影响

发布时间：2018-02-14 04:48

本文关键词： 梗阻性黄疸肠粘膜瘦素瘦素受体增殖　出处：《滨州医学院》2015年硕士论文　论文类型：学位论文

【摘要】：目的建立可复性梗阻性黄疸大鼠模型,探讨瘦素对可复性梗阻性黄疸大鼠肠粘膜增殖的影响及机制方法用”支撑管胆管结扎加支撑管拔除法”建立可复性梗阻性黄疸大鼠模型及胆管结扎法建立梗阻性黄疸模型。选用成年健康雄性Wistar大鼠64只,随机分成四组,可复性梗阻性黄疸组(ROJ)、梗阻性黄疸组(OJ)、可复性梗阻性黄疸+瘦素组(ROJ+LP)、梗阻性黄疸+瘦素组(OJ+LP)。ROJ组大鼠术后第5天,自左下腹部切口打开皮下轻轻拔出PICC导管完成胆管再通。瘦素组动物自术后第1天腹腔注射重组瘦素10ug/kg,bid,其余组动物相应时间点腹腔注射等量生理盐水作为对照。四组大鼠分别于术后5天和8天各处死8只,分别标记为ROJ5组、ROJ8组、OJ5组和OJ8组、ROJ5+LP组、ROJ8+LP组、OJ5+LP组和OJ8+LP组。放血法处死,剖腹探查观察胆管梗阻情况,并取标本。光学显微镜下HE切片观察肠黏膜结构的形态学变化,测量空肠黏膜上皮的绒毛高度及隐窝深度；免疫组化检测肠黏膜瘦素、受体以及Ki67抗原的表达情况。结果1.一般情况：ROJ组、OJ组及OJ+LP组分别于术后第4,6,2天各死亡1只,ROJ+LP组分别于术后第3、5天各死亡1只,死因为术后肠梗阻、肠坏死及腹腔积液。2.肠粘膜组织学观察：ROJ5组肠黏膜以非特异性炎细胞浸润、间质的充血水肿的炎症改变为主,肠黏膜萎缩变薄,绒毛的数量及密度下降,绒毛高度降低,间隙增大不连续,有凹陷存在,并有部分绒毛的脱落,隐窝深度变浅,腺体排列稀疏；ROJ8组肠绒毛结构较完整,上皮细胞基本呈高柱状,无明显间质水肿及中性粒细胞的浸润,基本维持正常的肠粘膜结构。0J5组肠粘膜形态基本同ROJ5组,0J8组的肠粘膜形态相比0J5组严重紊乱,炎细胞浸润、间质的充血水肿多见。ROJ5+LP组较ROJ5组肠粘膜损害程度差,有轻度绒毛排列紊乱,ROJ8+LP组肠粘膜形态未见明显异常表现。OJ5+LP组肠粘膜形态基本同ROJ5+LP组。OJ8+LP组仍有肠粘膜形态学异常改变,但程度较0J8组减轻。3.肠粘膜绒毛高度和隐窝深度：ROJ5组的绒毛高度明显低于ROJ8组,P0.01；0J5组的绒毛高度高于0J8组,P=0.05；ROJ5+LP组的绒毛高度明显低于ROJ8+LP组,P0.05；OJ5+LP组的绒毛高度高于OJ8+LP组,P0.05。ROJ8组明显低于ROJ8+LP组,P0.05；0J8组的绒毛高度明显低于OJ8+LP组,P0.05,ROJ5组的绒毛高度明显低于ROJ5+LP组,P0.01；0J5组的绒毛高度明显低于OJ5+LP组,P0.05。ROJ8组的绒毛高度明显高于0J8组,P0.01,ROJ8+LP组的绒毛高度明显高于OJ8+LP组,P0.01。ROJ5组的隐窝深度低于ROJ8组；0J5组的隐窝深度高于0J8组；ROJ5+LP组的隐窝深度低于ROJ8+LP组;OJ5+LP组的隐窝深度高于0J8+LP组。ROJ8+LP组的肠粘膜隐窝深度高于ROJ8组,P0.01。ROJ5+LP组的肠粘膜隐窝深度明显高于ROJ5组,P0.01；OJ8+LP组的肠粘膜隐窝深度明显高于OJ8组,P0.0l,OJ5+LP组的肠粘膜隐窝深度明显高于0J5组,P0.01。ROJ8组的隐窝深度明显高于0J8,P0.01,ROJ+LP组的绒毛高度明显高于OJ+LP组,P0.05。4.肠粘膜细胞增殖变化：肠黏膜增殖阳性细胞主要集中于肠粘膜隐窝处。ROJ5组的肠粘膜增殖指数明显低于ROJ8组；0J5组的肠粘膜增殖指数高于OJ8组,ROJ5+LP组的肠粘膜增殖指数低于ROJ8+LP组,P0.05;OJ5+LP组的肠粘膜增殖指数高于0J8+LP组。术后第8天,ROJ8组的肠粘膜增殖指数明显低于ROJ8+LP组,P0.01；OJ8组的肠粘膜增殖指数明显低于OJ8+LP组,P0.01;OJ5组的肠粘膜增殖指数明显低于OJ5+LP组,P0.01；组内比较：ROJ8+LP的肠粘膜增殖指数明显高于ROJ5+LP组,P0.05,差异有统计学意义。5.肠粘膜瘦素蛋白及其瘦体变化：瘦素及受体的阳性产物主要定位于胞浆中,呈棕黄色的颗粒,细胞间质也有少量着色。阳性细胞主要分布于肠黏膜上皮、隐窝细胞、肠腺细胞及固有层细胞。Leptin组及对照组肠黏膜中均有瘦素表达,但表达程度不一,高表达主要见于Leptin组,对照组肠黏膜组织表达相对较低或不表达。ROJ5组的0B蛋白表达明显ROJ8组；0J5组的0B蛋白表达高于0J8组,ROJ5+LP组的0B蛋白表达低于ROJ8+LP组;OJ5+LP组的0B蛋白表达高于OJ8+LP组。术后第8天,ROJ8组的肠粘膜0B蛋白表达明显低于ROJ+LP组,P0.01；OJ8组的肠粘膜0B蛋白表达明显低于OJ8+LP组,P0.05。同时ROJ8+LP组的肠粘膜0B蛋白表达明显高于OJ8+LP组,P0.05；术后第5天,ROJ5组的肠粘膜0B蛋白表达明显低于ROJ5+LP组,P=0.01；0.15组的肠粘膜0B蛋白表达明显低于0J5+LP组,P0.01。ROJ5组的瘦素受体表达低于ROJ8组；0J5组的瘦素受体表达高于0J8组,ROJ5+LP组的瘦素受体表达低于ROJ8+LP组;OJ5+LP组的瘦素受体表达高于OJ8+LP组。术后第8天,ROJ8组的瘦素受体表达明显低于ROJ8+LP组,P0.01;OJ8组的瘦素受体表达明显低于OJ8+LP组,P0.05；术后第5天,ROJ5组的瘦素受体表达明显低于ROJ5+LP组,P0.01；OJ5组的瘦素受体表达明显低于OJ5+LP组,P0.01。结论1.”支撑管胆管结扎加支撑管拔除法”制作的黄疸模型操作简便,解除黄疸不需二次开腹手术,创伤小,成功率、死亡率低,是一种稳定可靠的的可复性梗阻性黄疸大鼠模型.2梗阻性黄疸可致肠粘膜增殖能力的下降,主要体现在肠粘膜增殖指数Ki67的下降,绒毛高度和隐窝深度的降低。黄疸解除后各项指标能够得到改善。3外源性瘦素对梗阻性黄疸有促进肠粘膜细胞增殖的保护作用,在黄疸解除后,其作用仍能持续发挥。瘦素可加快肠粘膜增殖能力损害的修复。4外源性瘦素对肠粘膜细胞增殖作用的机理可能通过促进肠粘膜瘦素含量的增加及其受体表达的增强来实现。
[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.

【学位授予单位】：滨州医学院
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R657.3

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