抑制鞘氨醇激酶-2活性加重血管紧张素Ⅱ诱导的心肌细胞肥大

Pharmacological inhibition of sphingosine kinase-2 aggravates angiotensin Ⅱ-induced cardiomyocyte hypertrophy

目的本实验旨在明确鞘氨醇激酶(sphingosine kinase,Sph K)2在血管紧张素(angiotensin,Ang)Ⅱ诱导的心肌细胞肥大中的作用。方法分离并体外培养SD乳鼠心肌细胞。给予Ang Ⅱ(10μmol/L)处理24h诱导心肌细胞肥大。Ang Ⅱ刺激时分别给予溶媒或Sph K2特异性抑制剂ABC294640(1μmol/L)共处理。采用蛋白免疫印迹法检测心肌细胞Sph K2蛋白表达。采用酶联免疫吸附试验(enzyme-linked immunosorbant assay,ELISA)检测心肌细胞细胞核1-磷酸鞘氨醇(sphingosine 1-phosphate,S1P)水平。采用结晶紫染色观察Ang Ⅱ诱导的心肌细胞肥大程度。采用实时定量聚合酶链式反应(real time-polymerase chain reaction,RT-PCR)检测心肌细胞肥大标志基因心房钠尿肽(atrial natriuretic peptide,ANP)、脑钠尿肽(brain natriuretic peptide,BNP)和β-肌球蛋白重链(β-myosin heavy chain,β-MHC)mRNA表达水平。结果与对照组比较,Ang Ⅱ处理上调心肌细胞Sph K2蛋白表达和S1P浓度(均P<0.05),并增加心肌细胞横截面积与ANP、BNP和β-MHC mRNA表达水平(均P<0.05)。与溶媒组比较,ABC294640处理显著降低了心肌细胞核S1P浓度,并进一步增加了Ang Ⅱ处理后的心肌细胞横截面积和肥大标志基因mRNA表达水平(均P<0.05)。结论抑制Sph K2活性加重Ang Ⅱ诱导的心肌细胞肥大。Sph K2有可能成为治疗病理性心肌肥大的新靶点。

AIM The present study aimed to determine the role of sphingosine kinase-2 （SphK2） in angiotensin-II （AngII）-induced cardiomyocyte hypertrophy. METHODS Cardiomyocytes were isolated from Sprague Dawley rats and were cultured in vitro. Then, cardiomyocytes were exposed to AngII （10 μmol/L） for 24 hours. The expression of SphK2 was determined by Western blot and nuclear S1P abundance was analyzed by enzyme-linked immunosorbant assay （ELISA）. After vehicle or ABC294640 （1 μmol/L） co-treatment, AngII-induced cardiomyocyte hypertrophy was assayed by crystal violet staining. The mRNA expression levels of hypertrophy marker genes, including artial natriuretic peptide （ANP）, brain natriuretic peptide （BNP）, and β-myosin heavy chain （β-MHC）, were determined by real-time polymerase chain reaction （RT-PCR）. RESULTS The AngII treatment promoted cardiomyocyte hypertrophy and upregulated expression of hypertrophy marker genes. ABC294640 treatment significantly reduced nuclear S1P abundance in the cardiomyocyte. Notably, compared with the vehicle group, ABC294640 aggravated AngII-induced cardiomyocyte hypertrophy, as evidenced by increased cardiomyocyte size and expression of hypertrophy marker genes. CONCLUSION Inhibition of SphK2 exacerbates AngII-induced cardiomyocyte hypertrophy. SphK2 may be a potential therapeutic target for the prevention and treatment of pathological cardiac hypertrophy.