植酸对二甲肼诱导的结直肠癌大鼠模型肠粘膜机械屏障的影响

发布时间：2018-02-26 11:10

本文关键词： 植酸结直肠癌机械屏障　出处：《青岛大学》2017年硕士论文　论文类型：学位论文

【摘要】：目的:通过建立二甲肼(DMH)诱导的结直肠癌大鼠模型,观察不同剂量植酸干预对结直肠癌大鼠肠粘膜机械屏障粘液层MUC2和TFF3蛋白及三种紧密连接蛋白的影响,探讨植酸对结直肠癌大鼠模型肠粘膜机械屏障的作用机制。方法:60只Wistar雄性大鼠按体重随机区组分为正常对照组(NC)、模型组(MG)、植酸低剂量组(PL)、植酸中剂量组(PM)和植酸高剂量组(PH)。各植酸作用组每天分别给予0.25g/kg、0.5g/kg和1g/kg的植酸钠灌胃,NC和MG组灌胃等体积生理盐水。实验第5周起,除NC组外,其余各组大鼠颈背部注射剂量为30mg/kg的DMH持续18周,期间NC组注射等体积生理盐水。继续植酸钠灌胃至实验39周,处死大鼠。取NC组结肠组织及其他组结直肠肿瘤,免疫组化的方法检测其zo-1和occludin蛋白的表达情况;Western Blot方法检测MUC2、TFF3和claudin-1蛋白的表达情况;实时定量PCR方法检测MUC2、TFF3和claudin-1 m RNA的表达情况;酶联免疫吸附法(ELISA)检测血清中D-乳酸(D-LA)和内毒素(LPS)的表达情况。结果:实验结束时,模型组和植酸各剂量组均发生结直肠肿瘤,其中,模型组与植酸低剂量组肿瘤发生率均为100%,植酸中、高剂量组肿瘤发生率下降,分别为88.9%和66.7%;与对照组比较,模型组zo-1和occludin蛋白的表达明显降低(P0.05),植酸干预后,结直肠肿瘤中zo-1和occludin蛋白的表达显著升高(P0.05);DMH诱导使得MUC2和TFF3蛋白表达明显降低(P0.05),claudin-1蛋白表达明显升高(P0.05),经植酸作用,各植酸组MUC2蛋白表达均明显升高(P0.05),植酸中、高剂量组TFF3的蛋白表达明显升高(P0.05),且植酸中、高剂量组claudin-1蛋白的表达显著降低(P0.05);MUC2和claudin-1 m RNA表达水平的变化趋势与其对应的蛋白变化一致,而各植酸作用组TFF3 m RNA的表达水平与模型组相比均明显升高(P0.05)。与对照组相比,模型组大鼠血清中D-LA和LPS的表达明显升高(P0.05),各植酸作用组显著降低D-LA和LPS的表达(P0.05)。结论:(1).植酸可以改善DMH诱导的肠粘膜损伤,保护肠粘膜机械屏障,减少DMH诱导的大鼠结直肠肿瘤的发生率。(2)植酸通过上调肠粘膜粘液层中MUC2和TFF3蛋白及m RNA的表达水平来改善粘液层的屏障功能;植酸可以上调肠粘膜紧密连接蛋白zo-1、occludin的水平,并下调claudin-1的水平,进而调节肠上皮细胞的连接功能;植酸下调了血清中D-LA和LPS的水平,减少机体遭受侵袭及移位的可能。
[Abstract]:Aim: to establish a rat model of colorectal cancer induced by dimethylhydrazide (DMH), and to observe the effects of different doses of phytic acid on the mucus layer MUC2, TFF3 and three kinds of tight junction proteins in the mucous layer of intestinal mucosal barrier in rats with colorectal cancer. To investigate the mechanism of phytic acid on intestinal mucosal mechanical barrier in colorectal cancer rat model. Methods 60 Wistar male rats were randomly divided into normal control group, model group, low dose group, phytic acid medium dose group and middle dose group. Each phytic acid group was given 0.25 g / kg of 0.5 g / kg and 1 g / kg of sodium phytate by intragastric administration of normal saline of the same volume as NC group and MG group respectively from the 5th week of the experiment. Except NC group, the other groups were injected with 30 mg / kg DMH for 18 weeks, during which the NC group was injected with normal saline of the same volume. The rats were sacrificed after oral administration of sodium phytate for 39 weeks. The colon tissue of NC group and other groups of colorectal tumor were taken out. The expression of zo-1 and occludin protein was detected by immunohistochemical method, the expression of MUC2tFF3 and claudin-1 protein was detected by Western Blot, the expression of MUC2tF3 and claudin-1 m RNA was detected by real-time quantitative PCR method. Results: at the end of the experiment, colorectal neoplasms were found in the model group and each dose group of phytic acid, in which, the expression of D- lactic acid D-LAA and endotoxin LPS) in serum was detected by ELISA.Results: at the end of the experiment, colorectal neoplasms were found in the model group and the phytic acid group. The incidence of tumor in the model group and the low dose group was 100. In the phytic acid group, the tumor incidence rate in the high dose group was 88.9% and 66.7, respectively. Compared with the control group, the expression of zo-1 and occludin protein in the model group was significantly lower than that in the control group, and after the intervention of phytic acid, the expression of zo-1 and occludin protein in the model group was significantly lower than that in the control group. The expression of zo-1 and occludin in colorectal neoplasms was significantly increased. The expression of MUC2 and TFF3 protein decreased significantly in colorectal neoplasms. After phytic acid treatment, the expression of MUC2 protein in each phytic acid group was significantly increased, and in phytic acid, the expression of MUC2 protein was significantly increased in phytic acid. The protein expression of TFF3 increased significantly in high dose group, and in phytic acid, the expression of claudin-1 protein in high dose group decreased significantly the changes of MUC2 and claudin-1 m RNA levels in high dose group were consistent with the corresponding protein changes. The expression of TFF3 m RNA in each phytic acid group was significantly higher than that in the model group. The expression of D-LA and LPS in serum of the model group was significantly increased, and the expression of D-LA and LPS was significantly decreased in each phytic acid group. Conclusion the expression of D-LA and LPS was significantly decreased in the model group. Phytic acid can improve the intestinal mucosal injury induced by DMH and protect the intestinal mucosal mechanical barrier. To reduce the incidence of colorectal neoplasms induced by DMH) phytic acid can improve the barrier function of mucous layer by up-regulating the expression of MUC2 and TFF3 protein and m RNA in mucous layer of intestine, and phytic acid can up-regulate the level of zo-1occludin in intestinal mucosal mucous layer. Phytic acid down-regulated the levels of D-LA and LPS in serum and reduced the possibility of invasion and translocation.
【学位授予单位】：青岛大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R735.3;R-332

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