谷氨酰胺对大鼠肠缺血再灌注损伤的作用及机制

发布时间：2018-08-30 13:06

【摘要】：目的：探讨谷氨酰胺对肠缺血再灌注损伤大鼠肠粘膜形态病理学的保护作用以及分子机制，并观察不同的给药方法、不同给药时间对谷氨酰胺作用的影响。方法：1.用手术的方法构建大鼠肠缺血再灌注损伤模型。 2.酶联免疫吸附试验（ELISA）检测大鼠血标本中TNF-α含量的变化。 3.肠标本的石蜡包埋、切片、以及HE染色，观察谷氨酰胺对大鼠肠缺血再灌注损伤形态学的影响。 4.用RT-PCR和Western-blot的方法检测肠缺血再灌注损伤的各组大鼠肠标本组织中NF-κB和PPAR-γ基因的mRNA和蛋白表达情况。结果：1.肠缺血大鼠模型：正常喂养组（NG组）、给予尾静脉注射谷氨酰胺组(GIG组)、给予谷氨酰胺灌胃组(GOG组)、术中肠系膜上静脉给予谷氨酰胺组（GIOG组）、术中肠腔给予谷氨酰胺组（GIEC组）,模型构建成功。 2.我们用ELISA的方法分别检测五组大鼠(NG、GIG、GOG、GIOG、GIEC)血清中TNF-α的含量，我们发现1个小时、12个小时NG、GIEC组大鼠血清中TNF-α含量明显高于GIG、GOG组（P＜0.05），同时也比GIOG组含量高。 3.我们用HE染色的方法，观察了谷氨酰胺对大鼠肠粘膜缺血再灌注损伤形态学的影响。结果显示NG、GIEC组大鼠在1小时和12小时肠粘膜的形态损伤非常严重。而GOG、GIG组大鼠1小时和12小时肠粘膜的损伤程度比NG、GIEC组轻。GIOG组大鼠1小时和12小时肠粘膜的损伤程度比GOG、GIG严重。 4.我们使用逆转录聚合酶链反应技术（RT-PCR）和蛋白免疫印迹技术（Western-blot），去评估各组(NG、GIG、GOG、GIOG、GIEC)大鼠肠粘膜标本中NF-κB和PPAR-γ基因的mRNA和蛋白表达情况。NG、GIEC组NF-κB和PPAR-γ基因的mRNA和蛋白表达量最高，两组之间并没有差异（P＞0.05）；GIG、GOG组NF-κB和PPAR-γ基因的mRNA和蛋白表达量最低，与NG、GIEC组NF-κB和PPAR-γ基因的mRNA和蛋白表达量有显著的差异（P＜0.05）。结论：1.对于肠缺血再灌注损伤的大鼠模型，提前4天预给予谷氨酰胺，，不管是静脉注射组，还是口服组，谷氨酰胺都能够对大鼠的肠粘膜起到保护作用，能够降低血清TNF-α水平；而在肠缺血时给药，因为静脉注射使细胞内很快能够得到丰富的谷氨酰胺，所以能够起到一定的保护作用，但是肠腔内给药，因为缺血再灌注损伤的存在并不能吸收，所以未对肠粘膜起到保护作用，也不能降低血清TNF-α的水平。 2、对于肠缺血再灌注损伤模型，谷氨酰胺通过降低NF-κB的活性和增加PPAR-γ活性的机制，对肠缺血再灌注损伤起到保护作用。
[Abstract]:Aim: to investigate the protective effect and molecular mechanism of glutamine on intestinal mucosal morphology in rats with intestinal ischemia-reperfusion injury, and to observe the effects of different administration methods and administration time on glutamine. Method 1: 1. The model of intestinal ischemia-reperfusion injury in rats was established by surgical method. 2. Enzyme-linked immunosorbent assay (ELISA) was used to detect the content of TNF- 伪 in blood samples of rats. The effects of glutamine on the morphology of intestinal ischemia-reperfusion injury in rats were observed by paraffin embedding, sectioning and HE staining. 4. The expression of mRNA and protein of NF- 魏 B and PPAR- 纬 genes in intestinal tissues of rats with intestinal ischemia-reperfusion injury was detected by RT-PCR and Western-blot. The result is 1: 1. Model of intestinal ischemia in rats: normal feeding group (NG group), caudal intravenous injection of glutamine group (GIG group), glutamine gastric perfusion group (GOG group), intraoperative superior mesenteric vein glutamine group (GIOG group), intraoperative lumen administration of glutamyl glutamate. Amines group (GIEC group), the model was successfully constructed. 2. The content of TNF- 伪 in serum of five groups of rats (NG,GIG,GOG,GIOG,GIEC) was detected by ELISA method. We found that the content of TNF- 伪 in serum of the NG,GIEC group was significantly higher than that of the GIG,GOG group (P < 0. 05) and was higher than that of the GIOG group after 1 hour and 12 hours of NG,GIEC (P < 0. 05), and was also higher than that of the GIOG group (P < 0. 05). The effects of glutamine on the morphology of intestinal mucosal ischemia reperfusion injury in rats were observed by HE staining. The results showed that the morphological damage of intestinal mucosa in NG,GIEC group was very serious at 1 h and 12 h. However, the degree of intestinal mucosal damage in GOG,GIG group was more severe than that in NG,GIEC group at 1 and 12 hours compared with that in NG,GIEC group. 4. We used reverse transcriptase polymerase chain reaction (RT-PCR) and Western blot (Western-blot) to evaluate the expression of mRNA and protein of NF- 魏 B and PPAR- 纬 genes in the intestinal mucosa of rats in each group (NG,GIG,GOG,GIOG,GIEC). The mRNA and protein expression of NF- 魏 B and PPAR- 纬 genes were the highest in NGG IEC group. There was no difference between the two groups (P > 0. 05). The mRNA and protein expression of NF- 魏 B and PPAR- 纬 gene were the lowest in GIGGG group, and the mRNA and protein expression of NF- 魏 B and PPAR- 纬 gene in NG,GIEC group were significantly different (P < 0. 05). Conclusion 1. For the model of intestinal ischemia-reperfusion injury in rats, glutamine was given 4 days in advance. Both intravenous and oral groups, glutamine could protect intestinal mucosa and decrease serum TNF- 伪 level. And when given during intestinal ischemia, because intravenous injection can quickly make the cells rich in glutamine, so it can play a protective role, but intraluminal administration of drugs, because the presence of ischemia-reperfusion injury can not be absorbed, So it has no protective effect on intestinal mucosa, nor can it reduce the level of serum TNF- 伪. 2 for intestinal ischemia-reperfusion injury model, glutamine reduces the activity of NF- 魏 B and increases the activity of PPAR- 纬. It has protective effect on intestinal ischemia reperfusion injury.
【学位授予单位】：福建医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R965

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