槲皮素通过抑制蛋白酶体活性减轻心肌细胞肥大

目的:探讨槲皮素对血管紧张素Ⅱ(angiotensinⅡ,AngⅡ)诱导的心肌细胞肥大的防治作用及机制。方法:分别在原代新生大鼠心肌细胞和培养的H9c2心肌细胞,用100 nmol/L的AngⅡ诱导心肌细胞肥大模型,并给予3种不同浓度的槲皮素(10μmol/L,20μmol/L,40μmol/L)处理,利用免疫荧光检测原代和H9c2心肌细胞表面积的改变,采用荧光底物法检测H9c2心肌细胞蛋白酶体的活性变化,以及用Western blot检测H9c2心肌细胞糖原合酶激酶(GSK)-3α/β、Akt及其磷酸化情况。结果:与对照组相比,模型组原代心肌细胞和H9c2心肌细胞表面积均显著增大,槲皮素处理组2种心肌细胞表面积较模型组均明显减小,而20μmol/L槲皮素组减小更明显(P0.05)。在H9c2细胞实验中发现模型组蛋白酶体的糜蛋白酶样、半胱天冬酶样和胰蛋白酶样活性明显升高,20μmol/L和40μmol/L槲皮素组半胱天冬酶样和胰蛋白酶样活性较模型组均明显降低,而糜蛋白酶样活性只有在槲皮素20μmol/L时有显著差异(P0.05);Western blot检测结果显示,模型组磷酸化(p)-GSK-3α、p-GSK-3β和p-Akt水平较对照组均明显增加,而20μmol/L和40μmol/L槲皮素处理均使p-GSK-3α、p-GSK-3β和p-Akt水平明显下降(P0.05)。结论:槲皮素可明显抑制蛋白酶体活性从而减轻心肌细胞肥大,其机制可能是通过下调Akt活性而升高GSK-3α/β活性实现的。

Quercetin attenuates cardiomyocyte hypertrophy through proteasome inhibition

AIM: To investigate the protective effect of quercetin on angiotensin Ⅱ (AngⅡ)-induced cardiomyocyte hypertrophy and its possible mechanism. METHODS: Cardiomyocyte hypertrophy was induced by AngⅡ (100 nmol/L) in primary neonatal cardiomyocytes and H9c2 cells. The cells were treated with different concentration of quercetin (10 μmol/L, 20 μmol/L and 40 μmol/L) for 48 h and then the cardiomyocyte surface areas were measured by immunofluorescence. Proteasome activity was detected by fluorescent peptide substrate. The phosphorylated levels of GSK-3α/β and Akt in H9c2 cells were determined by Western blot. RESULTS: Compared with control group, the cardiomyocyte surface areas were both increased in primary cultured neonatal cardiomyocytes and H9c2 cells, while the surface areas were significantly decreased by quercetin, especially at concentration of 20 μmol/L compared with Ang Ⅱ group (P<0.05). Compared with control group, the chymotrypsin-like, trypsin-like and caspase-like activities of proteasome were all increased in H9c2 cells (P<0.05). The trypsin-like and caspase-like activities of proteasome were inhibited by 20 μmol/L and 40 μmol/L quercetin, while chymotrypsin-like activity was inhibited only at 20 μmol/L of quercetin compared with Ang Ⅱ group (P<0.05). In addition, phosphorylated levels of GSK-3α-Ser21, GSK-3β-Ser9 and Akt-Ser473 in Ang Ⅱ group were all increased compared with control group, which were obviously inhibited by in 20 μmol/L and 40 μmol/L quercetin (P<0.05). CONCLUSION: Quercetin decreases cardiomyocyte hypertrophy through proteasome inhibition, which may be related to the inhibition of Akt and therefore increasing activation of GSK-3α/β in H9c2 cells.