TLR4及相关炎症因子在糖尿病大鼠心脏、肝脏和肾脏中的表达

发布时间：2018-05-19 18:47

本文选题：糖尿病 + 糖尿病并发症　；参考：《河北医科大学》2015年硕士论文

【摘要】：目的:糖尿病(diabete mellitus,DM)是一种常见病,其常见的并发症主要发生在微血管和大血管,如糖尿病肾病、视网膜病、冠心病等。是糖尿病致伤、致残的主要原因。目前对并发症发病机制的研究较多,但近些年慢性炎症反应在并发症中的作用受关注。其中TLR4(Toll1ike receptor4,TLR4)这一普遍存在于多种免疫细胞的模式识别受体介导的炎症激活可能与DM并发症中的炎症反应有关。已知TLR4表达于巨噬细胞等固有性免疫细胞表面,糖尿病发生时,其内源性配体如热休克蛋白60(HSP60)、高迁移率族蛋白1(HMGBI)增加,可激活TLR4,诱导炎症反应。且我们前期临床和实验研究中均证实高糖状态下多种细胞TLR4的表达显著升高,与其相关的炎症因子IL-1β和TNF-α等的表达也升高,但是TLR4在其它脏器的表达及其相关炎症因子的作用尚未完全清楚。因此,本课题拟制备DM大鼠模型,测定不同组织如肝脏、心脏和肾脏的TLR4及其相关炎症因子IL-1β和TNF-α的表达,以及大鼠各组织炎症浸润的状态,全面了解DM状态下多脏器的炎症反应状态,进一步探讨DM的并发症的发生机制,为DM并发症的抗炎治疗提供实验依据。方法:1制备糖尿病大鼠模型:SD雄性大鼠腹腔注射STZ(链脲佐菌素),72h后取尾静脉血测定血糖,两次检测浓度均大于16.7mmol/L时为造模成功。糖尿病实验组大鼠和正常对照组大鼠各10只。2实验方法2.1检测各器官TLR4及其相关炎症因子m RNA的表达:造模12周后取大鼠的肾脏、心脏、肝脏组织,Real-time PCR方法检测TLR-4、TNF-α和IL-1β表达水平。2.2检测各组织器官炎症浸润状态:制备肾脏、心脏、肝脏的组织切片,HE染色检测炎症各组织的炎症细胞。3统计学方法:采用Excel和SPSS13.0统计学软件进行数据处理分析,数据以平均值±标准差表示,均数用独立样本t检验来比较组间差异性,P0.05为有统计学意义。结果:1成功建立了Ⅱ型糖尿病大鼠模型。2 TLR4 m RNA的表达水平Real-time PCR结果显示,大鼠心脏、肝脏和肾脏的TLR4表达如下,心脏内TLR4m RNA表达水平,在健康对照组和DM组中分别为1.104±0.189和1.079±0.503,健康对照组与DM组之间无统计学差异;肝脏内TLR4 m RNA的表达水平,健康对照组和DM组分别为0.933±0.275和8.899±1.769,TLR4 m RNA的表达水平明显高于对照组;肾脏内TLR4 m RNA的表达水平,健康对照组和DM组分别为1.032±0.125和2.766±0.663,DM组的TLR4 m RNA水平相比健康对照组明显升高。3 IL-1βm RNA的表达水平Real-time PCR结果显示,心脏的IL-1βm RNA表达健康对照组和DM组分别为1.025±0.263和1.120±0.311,健康对照组与DM组之间无显著差异;肝脏的IL-1βm RNA表达,健康对照组和DM组分别为1.175±0.275和3.940±0.958,DM组显著高于健康对照组;肾脏的IL-1βm RNA健康对照组和DM组分别为1.125±0.222和6.900±1.700,DM组显著高于健康对照组。4 TNF-αm RNA的表达水平心脏的TNF-αm RNA表达在健康对照组和DM组分别为1.075±0.359和1.080±0.449,健康对照组与DM组之间无显著差异;肝脏的TNF-αm RNA表达在健康对照组和DM组分别为1.150±0.311和4.060±1.097,DM组显著高于健康对照组;肾脏的TNF-αm RNA表达在健康对照组和DM组分别为1.050±0.208和10.38±1.753,DM组显著高于健康对照组。5 HE染色结果心脏、肝脏、肾脏组织HE染色结果显示:12w时,与健康对照组相比,DM组心脏心肌纤维肥大,间质水肿,血管周有炎细胞;DM组肝脏脂肪炎症细胞浸润多于对照组;DM组肾脏间质大量炎症细胞浸润。结论:1 TLR4 m RNA在DM大鼠肝脏、肾脏中有高表达,而TLR4 m RNA在心脏中的表达,DM组和对照组间不存在统计学差异。2 DM组大鼠在肝脏和肾脏中的IL-1βm RNA、TNF-αm RNA表达均显著升高,而在心脏无显著变化。3 DM组心脏、肝脏和肾脏均出现炎症细胞浸润。
[Abstract]:Objective: diabete mellitus (DM) is a common disease. The common complications are mainly in microvascular and large blood vessels, such as diabetic nephropathy, retinopathy, coronary heart disease and so on. It is the main cause of diabetes injury and disability. There are many studies on the pathogenesis of complications, but the chronic inflammatory reaction in recent years is in the complications. It is concerned. TLR4 (Toll1ike receptor4, TLR4), a pattern recognition receptor commonly found in many immune cells, may be associated with inflammatory responses in DM complications. The known TLR4 is expressed on the surface of a solid immune cell, such as macrophages, and its endogenous ligand such as heat shock protein 60 (HSP60) occurs when diabetes occurs. The high mobility group protein 1 (HMGBI) increased and activated TLR4 to induce the inflammatory response. In our previous clinical and experimental studies, the expression of TLR4 in a variety of cells in high glucose state was significantly increased, and the expression of the related inflammatory factors, IL-1 beta and TNF- a, was also increased, but it was the expression of TLR4 in other organs and related inflammatory factors. The role of DM rat is not completely clear. Therefore, we prepare a rat model to determine the expression of TLR4 and its related inflammatory factors IL-1 beta and TNF- alpha in different tissues such as liver, heart and kidney, as well as the state of inflammatory infiltration of various tissues in rats, to understand the inflammatory reaction state of multiple organs in DM state, and to further explore the occurrence of the complications of DM. Mechanism to provide experimental basis for the anti inflammatory treatment of DM complications. Methods: 1 the diabetic rat model was prepared: SD male rats were intraperitoneally injected with STZ (streptozotocin), 72h after 72h was used to measure blood sugar, and the concentration was greater than 16.7mmol/L. The experimental group of diabetic test group and normal control group of rats each had 10.2 experimental methods 2 .1 was used to detect the expression of TLR4 and related inflammatory factors m RNA in each organ: after 12 weeks of modeling, the kidney, heart, liver tissue, TLR-4, TNF- A and IL-1 beta expression level.2.2 were used to detect the inflammatory infiltration of tissues and organs by Real-time PCR method: the tissue sections of the kidneys, heart and liver were prepared, and the inflammation of the tissues was detected by HE staining. .3 statistical method: using Excel and SPSS13.0 statistics software for data processing and analysis, the data were expressed as mean standard deviation, and the average number was compared with the independent sample t test. The P0.05 was statistically significant. Results: 1 the expression level of.2 TLR4 m RNA in type II diabetic rat model was successfully established by Real-time PCR results. The expression of TLR4 in the heart, liver and kidney of the rat was as follows. The expression level of TLR4m RNA in the heart was 1.104 + 0.189 and 1.079 + 0.503 in the healthy control group and the DM group respectively. There was no statistical difference between the healthy control group and the DM group. The expression level of TLR4 m RNA in the liver was 0.933 + 0.275 and 8.899 + 1.769 and TLR4 m in the healthy group and the DM group, respectively. The expression level of RNA was significantly higher than that in the control group, and the expression level of TLR4 m RNA in the kidney was 1.032 + 0.125 and 2.766 + 0.663 in the healthy control group and DM group respectively. The TLR4 m RNA level in the DM group was significantly higher than that in the healthy control group, and the expression level of.3 IL-1 beta m was significantly higher than that of the healthy control group. There was no significant difference between 1.025 + 0.263 and 1.120 + 0.311 in the healthy control group and the DM group. The expression of IL-1 beta m RNA in the liver was 1.175 + 0.275 and 3.940 + 0.958 in the healthy control group and DM group, and the DM group was significantly higher than the healthy control group; the IL-1 beta m RNA healthy control group and the DM component of the kidney were 1.125 + 0.222 and 6.900 + 1.700, and the DM group was significant Gao Yujian The expression of TNF- alpha m RNA in the expression level of.4 TNF- alpha m RNA in the control group was 1.075 + 0.359 and 1.080 + 0.449 in the healthy control group and DM group respectively. There was no significant difference between the healthy control group and the DM group. The TNF- alpha m RNA expression in the liver was 1.150 + 0.311 and 4.060 + 1.097 respectively in the healthy control group and the healthy control group, and the group was significantly higher than the healthy control group. The expression of TNF- alpha m RNA in the kidney was 1.050 + 0.208 and 10.38 + 1.753 in the healthy control group and the DM group respectively. The DM group was significantly higher than the healthy control group with.5 HE staining results. The liver and renal tissue HE staining showed that when 12W, the cardiac muscle fiber was large, interstitial edema, vascular peripheral inflammatory cells, and DM group liver fat was compared with the healthy control group. The infiltration of inflammatory cells in the renal interstitium in DM group was more than that in the control group. Conclusion: 1 TLR4 m RNA is highly expressed in the liver of DM rats, and the expression of TLR4 m RNA in the heart, there is no statistical difference between the DM group and the control group, and the IL-1 beta in the liver and kidney of the.2 DM group is significantly elevated. There was no significant change in heart. Inflammatory cells infiltrated in heart, liver and kidneys of.3 DM group.
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R587.1

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