主动脉旁脂肪组织释放IL-6增加LDL受体缺陷小鼠动脉僵硬度的研究

发布时间：2018-08-23 10:15

【摘要】：慢性心血管疾病是一种常见的慢性疾病，其发病率和死亡率在疾病谱中均排在第一位。据报道，到2030年全球范围将有2330万人死于心血管疾病。心血管疾病导致了巨额的医疗保健支出，这种现象在中、低收入的发展中国家尤为突出。预计我国2005-2015年在心血管疾病的防治方面投入大约5580亿美元。因此，寻找预防心血管疾病的发生、发展的方法具有重要的社会意义。动脉僵硬度与心血管事件密切相关。因此，研究动脉僵硬度的发生机制及影响因素对于防治心血管疾病具有深刻的意义。由于高脂血症常合并动脉僵硬度的增加，从而使心血管事件发生率升高，早期干预高脂血症患者的动脉僵硬度对延缓和控制心血管疾病的进程有重大意义。研究表明，高脂血症可增加动脉僵硬度，但是高脂血症是否通过影响动脉壁细胞外基质蛋白的表达而促进动脉僵硬度的增加不得而知。另外，以往的研究发现，主动脉旁脂肪组织（PAT）可释放包括白介素-6（IL-6）在内的多种炎症介质和细胞因子，而IL-6是增加动脉僵硬度的重要原因。因此，我们推测高脂血症时，PAT通过释放IL-6而增加动脉僵硬度。 LDL受体缺陷小鼠（LDLr-/-小鼠）是典型高脂血症模型小鼠，本研究以LDLr-/-小鼠为主要研究对象，旨在探讨高脂血症对动脉僵硬度的影响及机制。首先，为了明确高脂血症通过影响动脉壁细胞外基质蛋白的表达而增加动脉僵硬度，本研究选取LDLr-/-小鼠为研究对象，并以野生型（WT）小鼠做对照，观察并比较两组小鼠的一般指标（体重、心脏重量、脂肪组织比重、瘦肉组织比重以及收缩压水平）、血脂、动脉脉搏波速度（aPWV）、主动脉弹性模量及动脉僵硬度相关蛋白（胶原蛋白、糖基化终末产物、弹性蛋白）的表达水平。其次，为了明确LDLr-/-小鼠通过PAT促进主动脉僵硬度的增加，我们分离LDLr-/-小鼠和WT小鼠的主动脉（带或不带PAT）然后分为以下四组进行体外培养：①WT组；②WT+PAT组；③LDLr-/-组；④LDLr-/-+PAT组。检测并比较四组小鼠的主动脉弹性模量，主动脉胶原蛋白、弹性蛋白、糖基化终末产物的表达水平。第三，我们分离LDLr-/-小鼠和WT小鼠的PAT进行体外培养，然后应用这两种不同的条件培养基孵育WT小鼠分离的主动脉段，分为以下两组：①WT组；②LDLr-/-组。检测并比较两组主动脉弹性模量的变化。然后，直接分离LDLr-/-小鼠和WT小鼠的PAT进行体外培养，分为以下两组：①WT组；②LDLr-/-组。检测这两组条件培养基中IL-6的表达水平。接下来，应用含或不含IL-6的培养液孵育WT小鼠分离的主动脉段，分为以下两组：①Control组；②IL-6组。检测并比较孵育后主动脉弹性模量差异。最后，分离LDLr-/-小鼠带有或不带有PAT的主动脉段，加用IL-6抗体干预，分为以下四组：①Control组；②IL-6Ab组；③PAT+control组；④PAT+IL-6Ab组。检测并比较各组主动脉段弹性模量的变化。结果显示：（1）LDLr-/-小鼠与WT小鼠的体重、心脏重量、脂肪组织比重、瘦肉组织比重以及收缩压水平未见明显差异（P0.05）；（2）LDLr-/-小鼠的总胆固醇、甘油三酯水平均明显高于WT小鼠，差异具有显著意义（P0.05）；（3）LDLr-/-小鼠主动脉僵硬度（aPWV、弹性模量）明显高于WT小鼠，动脉壁与动脉僵硬度相关的细胞外基质蛋白表达增加，差异具有显著意义（P0.05）；（4）与WT组相比，LDLr-/-组小鼠主动脉僵硬度明显增加，并且LDLr-/-+PAT组小鼠主动脉僵硬度进一步增加，差异具有统计学意义（P0.05）；（5）与WT小鼠相比，来自LDLr-/-小鼠的PAT条件培养基显著增加WT小鼠的主动脉僵硬度，差异具有统计学意义（P0.05）；（6）带有PAT培养的LDLr-/-小鼠主动脉壁中膜、外膜胶原蛋白I表达增加，外膜AGEs表达增加，，弹性蛋白表达减少，差异具有统计学意义（P0.05）；（7）LDLr-/-小鼠PAT细胞的大小明显大于WT小鼠，差异具有统计学意义（P0.05）；（8）LDLr-/-小鼠PAT的条件培养基对主动脉僵硬度的影响高于皮下脂肪组织、附睾旁脂肪组织的条件培养基；（9）LDLr-/-小鼠PAT较WT小鼠PAT分泌更多的IL-6，并且IL-6促进主动脉僵硬度的增加，IL-6抗体可降低PAT引起的动脉僵硬度的增加。以上结果表明：（1）LDLr-/-小鼠这一高脂血症模型小鼠动脉壁细胞外基质蛋白的表达失衡，主动脉僵硬程度明显高于WT小鼠；（2）LDLr-/-小鼠PAT促进主动脉僵硬度增加；（3）LDLr-/-小鼠PAT对主动脉僵硬度的影响要强于皮下脂肪组织、附睾旁脂肪组织及WT小鼠PAT。（4）LDLr-/-小鼠PAT通过释放IL-6促进动脉僵硬度增加。
[Abstract]:Chronic cardiovascular disease is a common chronic disease with the highest morbidity and mortality in the disease spectrum. It is reported that 23.3 million people will die of cardiovascular diseases worldwide by 2030. Cardiovascular diseases lead to huge medical and health expenditures, especially in low-income developing countries. From 2005 to 2015, China has invested about 558 billion US dollars in the prevention and treatment of cardiovascular diseases. Therefore, it is of great social significance to find a way to prevent the occurrence and development of cardiovascular diseases. Hyperlipidemia is often associated with increased arterial stiffness, which increases the incidence of cardiovascular events. Early intervention of arterial stiffness in patients with hyperlipidemia is of great significance in delaying and controlling the progression of cardiovascular disease. Studies have shown that hyperlipidemia can increase arterial stiffness, but whether hyperlipidemia passes through. In addition, previous studies have found that para-aortic adipose tissue (PAT) releases a variety of inflammatory mediators and cytokines, including interleukin-6 (IL-6), and IL-6 is an important cause of increased arterial stiffness. During the disease, PAT increased arterial stiffness by releasing IL-6.
LDL receptor deficient mice (LDLr-/-mice) are typical hyperlipidemia model mice. The purpose of this study was to investigate the effect of hyperlipidemia on arterial stiffness and its mechanism. Firstly, in order to clarify that hyperlipidemia increases arterial stiffness by affecting the expression of extracellular matrix protein in arterial wall, this study selected LDLr-/-mice as the main research object. LDLr - / - mice were selected as the research object, and wild type (WT) mice were used as the control group. The general indexes (body weight, heart weight, adipose tissue weight, lean tissue weight and systolic blood pressure level), blood lipid, arterial pulse wave velocity (aPWV), aortic elastic modulus and arterial stiffness related protein (collagen, glycosylation) were observed and compared between the two groups. Secondly, in order to clarify the effect of LDLr - / - on aortic stiffness, we isolated the aortas of LDLr - / - mice and WT mice (with or without PAT) and divided them into the following four groups: WT group; WT + PAT group; LDLr - / - group; LDLr - / - group; LDLr - / -+ PAT group. Compared with the four groups of mice, the aortic elastic modulus, aortic collagen, elastin and the expression level of glycation end products were measured. The changes of aortic elastic modulus were detected and compared between the two groups. Then, the PATs of LDLr - / - mice and WT mice were separated and cultured in vitro. The PATs were divided into the following two groups: WT group and LDLr - / - group. The arterial segments were divided into the following two groups: control group, IL-6 group, PAT + control group, PAT + IL-6Ab group, and PAT + IL-6Ab group. The change of elastic modulus of aortic segments.
The results showed that: (1) There were no significant differences in body weight, heart weight, adipose tissue weight, lean tissue weight and systolic blood pressure between LDLr - / - mice and WT mice (P 0.05); (2) Total cholesterol and triglyceride levels in LDLr - / - mice were significantly higher than those in WT mice (P 0.05); (3) aortic stiffness of LDLr - / - mice (aPWV, aPWV, triglyceride level). Compared with WT group, the aortic stiffness of LDLr-/- group was significantly increased, and the aortic stiffness of LDLr-/-+PAT group was further increased, the difference was statistically significant (P 0.05). (5) Compared with WT mice, PAT conditioned medium from LDLr - / - mice significantly increased aortic stiffness of WT mice, the difference was statistically significant (P 0.05); (6) In LDLr - / - mice aortic wall media with PAT culture, the expression of collagen I in adventitia increased, the expression of AGEs in adventitia increased, and the expression of elastin decreased, the difference was statistically significant (P 0.05). Significance (P 0.05); (7) The size of PAT cells in LDLr - / - mice was significantly larger than that in WT mice, and the difference was statistically significant (P 0.05); (8) The effect of conditioned medium of LDLr - / - mice PAT on aortic stiffness was higher than that in subcutaneous adipose tissue, paraepididymal adipose tissue conditioned medium; (9) LDLr - / - mice PAT secreted more IL-6 than WT mice PAT, and Moreover, IL-6 increased aortic stiffness. IL-6 antibody could reduce the increase of arterial stiffness caused by PAT.
The results showed that: (1) the expression of extracellular matrix protein in arterial wall of LDLr - / - mice was unbalanced, and the degree of aortic stiffness was significantly higher than that of WT mice; (2) LDLr - / - mice PAT promoted the increase of aortic stiffness; (3) LDLr - / - mice PAT had stronger effect on aortic stiffness than subcutaneous adipose tissue, paraepididymal fat. The PAT of PAT. (4) LDLr-/- mice in fat tissue and WT mice increased the arterial stiffness through releasing IL-6.
【学位授予单位】：吉林大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R543

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