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目的探讨visfatin蛋白在动脉粥样硬化(AS)病变形成中的作用,以及丹参酮ⅡA抗载脂蛋白E基因缺陷(apo E-/-)小鼠AS病变的可能机制。方法将C57BL/6J背景的apo E-/-小鼠随机分为4组:模型组,丹参酮ⅡA高、低剂量组,阳性对照组(辛伐他汀组),每组8只。并以8只C57BL/6J小鼠作为空白对照组。apo E-/-小鼠给予高脂饮食,并按分组情况给予丹参酮ⅡA磺酸钠(20,10 mg·kg-1)或辛伐他汀(5 mg·kg-1)干预。第14周后全部处死,HE染色观察动脉斑块形成程度,测定斑块面积与管腔面积之比。酶法检测血清脂质含量。ELISA方法检测血清高敏C反应蛋白(hs-CRP)、肿瘤坏死因子-琢(TNF-琢)、金属基质蛋白酶-9(MMP-9)含量。采用western blotting法检测主动脉壁visfatin表达。结果 (1)空白对照组主动脉未见动脉粥样硬化改变,模型组有较明显的主动脉内膜增厚及粥样斑块的形成。模型组血清总胆固醇(TC)、甘油三酯(TG)、低密度脂蛋白胆固醇(LDL-C)水平及hs-CRP、TNF-α、MMP-9表达、动脉壁visfatin表达显著高于空白对照组,而高密度脂蛋白胆固醇(HDL-C)水平则显著低于空白对照组(均P<0.05)。(2)与模型组比较,丹参酮ⅡA高、低剂量组、辛伐他汀组均可显著减少斑块面积与管腔面积比值,降低血脂中TC、TG、LDL-C水平(P<0.05),升高HDL-C水平(P<0.05);明显降低血清细胞因子hs-CRP、TNF-α、MMP-9水平,降低小鼠主动脉壁visfatin表达。(3)在减少斑块面积、改善血脂水平、抑制炎症、抑制visfatin表达方面,丹参酮ⅡA均具有显著的剂量依赖性。丹参酮ⅡA高剂量在减少斑块面积、改善血脂水平方面与辛伐他汀差异无显著性意义,但在降低hs-CRP、TNF-α、MMP-9水平,以及下调主动脉壁visfatin表达方面显著优于辛伐他汀组。(4)apo E-/-小鼠动脉壁组织visfatin表达水平与血清hs-CRP、TNF-α、MMP-9水平具有显著的正相关关系。结论 visfatin参与了AS的发生发展,其参与机制可能与炎症机制有关。丹参酮ⅡA可能通过调节apo E-/-小鼠血脂代谢、抑制炎症细胞因子hs-CRP、TNF-α、MMP-9的表达,及下调主动脉壁visfatin表达,进而抑制apo E-/-小鼠AS病变的形成发展。
Objective To investigate the role of visfatin in the development of atherosclerosis (AS) and the possible mechanism of tanshinone ⅡA in the pathogenesis of AS in apolipoprotein E deficient mice. Methods Apo E - / - mice on C57BL / 6J background were randomly divided into 4 groups: model group, tanshinone Ⅱ A high and low dose group and positive control group (simvastatin group), with 8 mice in each group. Eight C57BL / 6J mice were used as blank control group. Apo E - / - mice were given a high fat diet and were given tanshinone Ⅱ A sulfonate (20, 10 mg · kg -1) or simvastatin (5 mg · kg -1) by grouping. After 14 weeks, all of them were sacrificed, the degree of arterial plaque formation was observed by HE staining, and the ratio of plaque area to luminal area was determined. Enzymatic detection of serum lipid content. Serum high-sensitivity C-reactive protein (hs-CRP), tumor necrosis factor-α (TNF-α) and MMP-9 were detected by ELISA. The expression of visfatin in the aortic wall was detected by western blotting. Results (1) There was no change of atherosclerosis in the aorta of control group. The aortic intima thickening and plaque formation were obvious in model group. The levels of total cholesterol (TC), triglyceride (TG), low density lipoprotein cholesterol (LDL-C) and the expression of hs-CRP, TNF-α and MMP-9 and visfatin in arterial wall in model group were significantly higher than those in blank control Group, while the level of HDL-C was significantly lower than that of the blank control group (all P <0.05). (2) Compared with the model group, tanshinone ⅡA high, low dose and simvastatin groups could significantly reduce the ratio of plaque area and luminal area, and lower the levels of TC, TG and LDL-C in blood lipids (P <0.05) (P <0.05). The levels of serum hs-CRP, TNF-α and MMP-9 were significantly decreased and the visfatin expression in the aorta was decreased. (3) Tanshinone IIA has a significant dose-dependent manner in reducing plaque area, improving blood lipid levels, inhibiting inflammation and inhibiting visfatin expression. Tanshinone Ⅱ A high dose in the reduction of plaque area, improve blood lipid levels and simvastatin difference was not significant, but in reducing hs-CRP, TNF-α, MMP-9 levels, and down regulation of aortic wall visfatin expression was significantly better In simvastatin group. (4) There was a significant positive correlation between the expression of visfatin and the levels of hs-CRP, TNF-α and MMP-9 in the apo E - / - mice arterial wall. Conclusion Visfatin is involved in the pathogenesis of AS, and its mechanism may be related to the mechanism of inflammation. Tanshinone IIA may inhibit the apo E - / - mice by regulating blood lipid metabolism, inhibiting the expression of inflammatory cytokines hs-CRP, TNF-α and MMP-9 and ameliorating visfatin The formation and development of AS lesions.