缺氧诱导因子1α在异丙肾上腺素诱导的大鼠心房纤维化中的表达及意义

Expression and its significance of hypoxia-inducing factor-1α in rats with isoproterenol-induced atrial fibrosis

目的探讨缺氧诱导因子1α(HIF-1α)在盐酸异丙肾上腺素(ISO)诱导的大鼠心房纤维化中的表达及其可能机制。方法健康雄性Wistar大鼠30只,随机均分为空白对照组、ISO组、ISO+西罗莫司(Rapa)干预组(Rapa组)。于实验15d时处死大鼠取心肌组织,放射免疫法检测血管紧张素Ⅱ(AngⅡ)的含量,HE和Masson染色法观察纤维化程度即胶原容积分数(CVF),免疫组织化学法、Western blotting检测HIF-1α、转化生长因子β1(TGF-β1)和基质金属蛋白酶9(MMP-9)在大鼠心房纤维化组织中的表达,并分析HIF-1α、TGF-β1、MMP-9蛋白表达量与心房纤维化指标CVF的相关性,以及HIF-1α、TGF-β1、MMP-9蛋白表达量之间的相关性。结果 ISO组、Rapa组AngⅡ含量较空白对照组明显升高(P<0.01)。空白对照组CVF为15.482%±0.837%,无心房纤维化,而Rapa组、ISO组CVF分别为16.730%±1.052%、86.704%±1.928%,Rapa组较ISO组的心房纤维化程度明显减弱(P<0.01)。免疫组化及Western blotting检测显示,与空白对照组相比,ISO组HIF-1α、TGF-β1和MMP-9蛋白表达量均明显增加(P<0.01),与ISO组比较,Rapa组中HIF-1α、TGF-β1和MMP-9蛋白表达量明显降低(P<0.01)。HIF-1α、TGF-β1和MMP-9蛋白表达量均与心房纤维化程度(以CVF为评价指标)呈正相关(r=0.987,r=0.988,r=0.917,P<0.01);HIF-1α与TGF-β1、MMP-9蛋白表达量呈正相关(r=0.976,r=0.901,P<0.01),MMP-9与TGF-β1蛋白表达量亦呈正相关(r=0.912,P<0.01)。结论在异丙肾上腺素诱导的大鼠心房纤维化过程中,AngⅡ、HIF-1α、TGF-β1、MMP-9发挥着重要作用。

Objective To explore the expression ofhypoxia-inducing factor-1 α （HIF-1α） and its significance in the rats with isoproterenol-induced atrial fibrosis. Methods Thirty male Wistar rats were randomly divided into control group, isoproterenol [Iso, 5mg/（kg.d） for 7 days] group, and Iso ＋ sirolimus （Rapa） group. The atrial fibrosis rat model was reproduced by subcutaneous injection of high doses of Iso in Iso group and Rapa group. Rats were treated with isoproterenol plus sirolimus in Iso-si group. All the animals were sacrificed 15 days after treatment. The contents of angiotensin ]I （Ang I] ） in rat myocardial tissue were determined by radioimmunoassay. HE staining and Masson staining were performed for the measurement of the degree of atrial fibrosis in terms of collagen volume fraction （CVF）. Immunohistochemistry and Western blotting were performed to detect the expressions of HIF-1α, TGF-β1 and MMP-9. The correlations of HIF-1α, TGF-β1 and MMP-9 to CVF and expressions of HIF-1α, TGF-β1 and MMP-9 were analyzed. Results The Ang Ⅱ content was significantly higher in Iso group and Rapa group than that in control group （P〈0.01）. No atrial fibrosis occurred in control group （CVF was 15.482% ± 0.837%）, and the atrial fibrosis was significantly milder in Rapa group （CVF was 16.730% ± 1.052%） than in Iso group （CVF was 86.704% ± 1.928%, P〈0.01）. Immunohistochemistry and Western blotting revealed that expressions of HIF-1α, TGF-β1 and MMP-9 were significantly increasedin Iso group compared with that of control group （P〈0.01）, but the expressions were significantly decreased in Rapa group compared with that in Iso group （P〈0.01）. The expressions of HIF-1α, TGF-β1 and MMP-9 were positively correlated with the fibrosis degree （P〈0.01）; the HIF-1α content was positively correlated with the contents of TGF-β1 and MMP-9 （P〈0.01）; the MMP-9 content was positively correlated with the TGF-β1 content （P〈0.01）. Conclusion Ang Ⅱ, HIF-1α, TGF-β1 and MMP-9 play an important role during the development of isoproterenol-induced atrial fibrosis in rats.