隔药饼灸对高脂血症合并AS兔胆固醇逆转运SR-B1介导途径及炎症反应的影响

发布时间：2018-01-31 05:19

本文关键词： 隔药饼灸高脂血症动脉粥样硬化胆固醇逆转运炎症因子　出处：《湖南中医药大学》2016年博士论文　论文类型：学位论文

【摘要】：目的:1、观察隔药饼灸对动脉粥样硬化兔血脂代谢变化、组织结构变化和血清相关炎性因子的影响;2、观察隔药饼灸对动脉粥样硬化兔胆固醇逆转运SR-B1介导途径中PPARγ、SR-B1表达的影响,探讨隔药饼灸抗AS的作用机制。方法:健康纯种雄性新西兰大耳白兔40只随机分为5组:空白对照组,模型组,隔药饼灸组,直接灸组,辛伐他汀组。采用高脂饲料喂养方式制备高脂血症合并AS兔模型,分别给予不同干预手段进行治疗,干预结束后,禁食,经耳缘静脉麻醉后采集血液样本,处死动物,取主动脉及肝脏样本。各组兔主动脉、肝脏标本HE染色,观察组织变化情况;酶法测定血清中总胆固醇(TC)、甘油三酯(TG)、低密度脂蛋白胆固醇(LDL-C)、高密度脂蛋白胆固醇(HDL-C)水平,测定胆固醇含量;酶联免疫吸附法(ELISA)测定血清中肿瘤坏死因子α(TNF-a)、干扰素γ(IFN-γ),白介素6(IL-6)白介素10(IL-10)水平;蛋白印迹法(WB)测定肝脏PPARγ、SR-B1蛋白表达量;实时定量聚合酵链式反应(real-time邋PCR)测定肝脏PPARγ、SR-B1 mRNA表达量。结果:1、正常组主动脉血管壁内膜表面光滑,各层结构清晰,内皮细胞完整连续,平滑肌细胞排列整齐紧密有序,无脂质、斑块等沉积;模型组主动脉血管壁明显增厚,内皮层脱落,管壁满布脂质、斑块,大量泡沫细胞出现;隔药饼灸组和直接灸组内膜轻度增厚,平滑肌细胞厚度均匀、排列整齐,少见泡沫细胞;辛伐他汀组内膜增厚,各层结构相比于模型组清晰,有泡沫细胞形成,内膜下少量脂质沉积,未见明显斑块。正常组肝组织结构明显,肝细胞核显示清楚,肝窦空虚、无瘀血,无脂质变性,排列规整,呈多边形分布,胞浆均匀;模型组肝组织结构不清晰,部分肝窦变窄或缺失,内皮细胞不明显,肝细胞肿胀、胞浆透明,部分肝细胞呈脂肪变性,部分出现脂滴融合,脂肪空泡弥漫充布,间质组织有淋巴细胞和浆细胞的浸润;隔药灸组和直接灸组肝脏组织结构比较清晰,肝细胞轻度脂肪变性,肝脏组织的脂滴沉积减少,轻度炎性反应;辛伐他汀组肝细胞脂肪变性程度较模型组减轻,少数有脂滴空泡,肝组织结构有改变,少量肝细胞呈现水样变性,大部分肝细胞核位置未发生改变,形态相对正常。2、与正常组比较,模型组兔的血清TC、TG、LDL-C水平较高,差异有统计学意义(p0.01);模型组兔的血清hdl-c水平较低,差异有统计学意义(p0.01)。与模型组比较,隔药饼灸组、直接灸组、辛伐他汀组血清tc、tg、ldl-c降低明显,hdl-c水平升高,差异有统计学意义。与正常组比较,模型组主动脉、肝组织、脂肪组织的胆固醇含量较高,差异有统计学意义(p0.01)。与模型组比较,隔药饼灸组、直接灸组、辛伐他汀组主动脉、肝组织、脂肪组织的胆固醇含量明显降低,差异有统计学意义(p0.01)。3、与正常组比较,模型组、直接灸组、辛伐他汀组血清il-6、tnf-α、ifnγ表达升高,有显著性差异(p0.01或p0.05),隔药灸组与正常组比较,il-6、ifnγ无显著性差异,tnf-α表达升高与正常组有显著差异;与模型组相比,隔药灸组、直接灸组、辛伐他汀组血清il-6、tnf-α、ifnγ表达均有明显降低,差异有统计学意义(p0.01或p0.05)。与正常组比较,模型组、辛伐他汀组血清il-10表达明显减少,差异有统计学意义(p0.05);与模型组相比,隔药灸组、直接灸组、辛伐他汀组血清il-10表达均有明显升高,差异有统计学意义(p0.01或p0.05)。4、与正常组比较,模型组pparγ、sr-b1蛋白表达降低,差异有统计学意义(p0.01或p0.05),隔药灸组肝脏pparγ、sr-b1蛋白表达较高,差异有统计学意义(p0.01或p0.05),直接灸组兔肝脏pparγ蛋白表达较高,差异有统计学意义(p0.01或p0.05),sr-b1蛋白表达与正常组比较无统计学差异;与模型组相比较,隔药灸组、直接灸组、辛伐他汀组肝脏的pparγ、sr-b1蛋白表达均呈升高趋势,差异有统计学意义(p0.01或p0.05);与隔药灸组相比较,直接灸组、辛伐他汀组肝脏的pparγ、sr-b1蛋白表达均呈降低趋势,差异有统计学意义(p0.01或p0.05)。与正常组比较,模型组肝脏pparγ、sr-b1mrna表达降低,差异有统计学意义(p0.05),隔药灸组、直接灸组肝脏pparγ、sr-b1mrna表达较高,差异有统计学意义(p0.05),辛伐他汀组肝脏pparγmrna表达较高,差异有统计学意义(p0.05),sr-b1mrna表达与正常组比较无统计学差异;与模型组相比较,隔药灸组、直接灸组、辛伐他汀组肝脏的pparγ、sr-b1mrna表达均升高,差异有统计学意义(p0.01或p0.05);与隔药灸组相比较,直接灸组、辛伐他汀组肝脏的pparγ、sr-b1mrna表达均呈降低趋势,差异有统计学意义(p0.01或p0.05)。结论:1、隔药饼灸对动脉粥样硬化兔血脂代谢变化具有良性调节作用,对动脉内膜、肝脏等组织病理变化有修复作用。2、隔药饼灸抗动脉粥样硬化作用可能与其对动脉粥样硬化兔血清相关炎性因子的影响有关。3、隔药饼灸影响胆固醇逆转运SR-B1介导途径中PPARγ、SR-B1蛋白及基因表达,激活PPARγ,诱导SR-B1的表达,可以促进胆固醇逆转运,抑制或减缓动脉粥样硬化,可能是隔药饼灸抗动脉粥样硬化的作用机制之一。
[Abstract]:Objective: To observe the change of blood lipid metabolism in 1 rabbits with atherosclerosis moxibustion effect, the changes of organizational structure and the related serum inflammatory factors; 2, the observation of cake separated moxibustion on cholesterol reverse transport of atherosclerotic rabbit SR-B1 mediated pathway in PPAR gamma, SR-B1 expression, and explore mechanisms of moxibustion on anti AS methods: male purebred New Zealand 40 rabbits were randomly divided into 5 groups: control group, model group, moxibustion group, direct moxibustion group and simvastatin group. The rabbit model of hyperlipidemia and AS by feeding high fat diet, respectively given different interventions for treatment, intervention after fasting, intravenously after anesthesia, blood samples were taken, animal, aorta and liver samples. The Rabbits Aorta, liver specimens HE staining, observe the changes of tissue; Determination of total serum bile cholesterol enzyme (TC), triglyceride (TG), Low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C) levels, determination of cholesterol content; enzyme linked immunosorbent assay (ELISA) determination of tumor necrosis factor alpha in serum (TNF-a) and interferon gamma (IFN- gamma), interleukin 6 (IL-6) and interleukin 10 (IL-10) protein level; Western blot (WB) were detected by PPAR gamma, SR-B1 protein expression; real-time quantitative polymerase chain reaction (real-time cream yeast PCR) determination of liver PPAR SR-B1 gamma, the expression of mRNA. Results: 1, normal aortic wall intimal smooth surface, each layer of the structure is clear, endothelial cells complete and continuous, smooth muscle cells arranged in neat rows closely, no lipid plaque deposition; model group aortic wall thickening, endothelial layer off, the wall covered with lipid plaque, a large number of foam cells; moxibustion group and direct moxibustion group in membrane mild thickening, smooth muscle cells arranged in neat, uniform thickness, rare bubble Foam cell; simvastatin group intimal thickening, compared the structure of each layer in the model group, with the formation of foam cells and a small amount of lipid deposition under intima, no obvious plaque. Normal liver tissue structure significantly, liver nuclei showed clear liver sinus empty, no blood, no change of lipid, ordered, polygonal cell distribution. Pulp evenly; model group liver tissue structure is clear, part of the liver sinus narrowing or lack of endothelial cells is not obvious, liver cell swelling, clear cytoplasm, part of the liver cells showed fatty degeneration, some lipid droplets fused, fat vacuoles diffuse filling cloth, interstitial tissue infiltration of lymphocytes and plasma cells; isolation medicine moxibustion group and liver tissue in direct moxibustion group structure is relatively clear, mild liver cell fatty degeneration, lipid deposition in liver tissue decreased, mild inflammatory reaction; simvastatin group the degree of liver steatosis reduced compared with model group, there are a few lipid droplet vacuoles, The liver structure changes, a small amount of liver cells showed hydropic degeneration, most liver nuclei did not change the position, form relatively normal.2, compared with the normal group, serum TC, TG in the model group, the LDL-C level is higher, the difference was statistically significant (P0.01); the serum level of HDL-C in the model group was statistically lower significant differences (P0.01). Compared with the model group, moxibustion group, direct moxibustion group, simvastatin group, serum TC, TG, LDL-C decreased, HDL-C level increased, the difference was statistically significant. Compared with the normal group, model group, liver aorta, cholesterol content of adipose tissue is higher, the difference was statistically significant (P0.01). Compared with the model group, moxibustion group, direct moxibustion group, simvastatin group, liver aorta, cholesterol content of adipose tissue decreased significantly, the difference was statistically significant (.3, P0.01) compared with the normal group, model group, direct moxibustion group, Essien In the statin group serum IL-6, tnf- alpha, IFN expression increased significantly (P0.01 or P0.05), moxibustion group compared with normal group, no significant difference in IL-6, IFN gamma, tnf- expression increased significantly compared with normal group; compared with the model group, moxibustion group, direct moxibustion group, simvastatin group, serum IL-6, tnf- alpha, IFN expression decreased significantly, the difference was statistically significant (P0.01 or P0.05). Compared with the normal group, model group, simvastatin group serum IL-10 expression was significantly reduced, the difference was statistically significant (P0.05); compared with the model group, moxibustion group, direct moxibustion the expression of serum IL-10 group, simvastatin group were significantly increased, the difference was statistically significant (P0.01 or P0.05).4, compared with the normal group, model group, PPAR gamma, SR-B1 protein expression decreased, the difference was statistically significant (P0.01 or P0.05), moxibustion group liver PPAR gamma, SR-B1 protein expression is higher, the difference was significant Yi (P0.01 or P0.05), direct moxibustion group of rabbit liver PPAR gamma protein expression was higher, the difference was statistically significant (P0.01 or P0.05), the expression of SR-B1 protein showed no significant difference compared with the normal group; compared with the model group, moxibustion group, direct moxibustion group, simvastatin group PPAR gamma liver, the expression of SR-B1 protein increased, the difference was statistically significant (P0.01 or P0.05); compared with moxibustion group, direct moxibustion group, simvastatin group PPAR gamma liver, SR-B1 protein expression were decreased, the difference was statistically significant (P0.01 or P0.05). Compared with the normal group, model group, liver PPAR, sr-b1mrna the expression is reduced, the difference was statistically significant (P0.05), moxibustion group, direct moxibustion group liver PPAR, sr-b1mrna expression was higher, the difference was statistically significant (P0.05), simvastatin group liver PPAR gamma mRNA expression was higher, the difference was statistically significant (P0.05), and the expression of sr-b1mrna in normal group There was no significant difference; compared with the model group, moxibustion group, direct moxibustion group, simvastatin group PPAR gamma liver, the expression of sr-b1mrna was increased, the difference was statistically significant (P0.01 or P0.05); compared with moxibustion group, direct moxibustion group, simvastatin group liver PPAR gamma, sr-b1mrna expression decreased, the difference was statistically significant (P0.01 or P0.05). Conclusion: 1. Moxibustion has a good regulating effect on blood lipid metabolism in rabbits with atherosclerosis changes on arterial intima, the liver tissue pathological changes of repair.2, medicine separated moxibustion anti atherosclerosis effect may be related to atherosclerosis in rabbit serum inflammatory factors related to.3, moxibustion effect of reverse cholesterol transport mediated by SR-B1 pathway in PPAR gamma, SR-B1 protein expression and gene activation, PPAR gamma, induce the expression of SR-B1, can promote the reverse cholesterol transport, inhibit or slow down Atherosclerosis may be one of the mechanisms of anti atherosclerotic action of cake separated moxibustion.

【学位授予单位】：湖南中医药大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R245

【参考文献】