PDE5抑制剂对异丙肾上腺素诱导乳鼠心肌细胞肥大的作用

Effect of PDE5 inhibitor on isoproterenol-induced cardiomyocyte hypertrophy in neonatal rats

目的探讨磷酸二酯酶(PDE)5抑制剂对异丙肾上腺素(Iso)所致的乳鼠心肌细胞肥大的保护作用。方法分离大鼠乳鼠心肌细胞,分为对照(Con)组, Iso组及异丙肾上腺素+西地那非(Iso+Sil)组,通过检测各组细胞活力及乳酸脱氢酶(LDH)含量,确定Sil的后续实验浓度,通过RT-PCR检测心肌肥大指标心房钠尿肽(ANP)和β-肌球蛋白重链(β-MHC) mRNA含量、流式细胞计数检测凋亡情况及Western blot检测内质网应激相关蛋白葡萄糖调节蛋白(GRP)78和CCAAT增强子结合蛋白同源蛋白(CHOP)水平。结果与Con组相比,Iso组细胞活力降低(P<0. 01),LDH的释放增加(P<0. 05)。与Iso组相比,一定浓度Sil预处理可以提高细胞活力及减少LDH的释放(P<0. 05),在5μmol/L Sil预处理时达到最大效应。与Con组相比,Iso组增加ANP和β-MHC mRNA的表达、上调凋亡比率以及增加GRP78和CHOP的蛋白水平。与Iso组相比,Sil预处理可以降低ANP和β-MHC mRNA的表达,下调凋亡比率以及抑制GRP78和CHOP的蛋白表达。结论 PDE5抑制剂Sil可以有效抑制Iso诱导的乳鼠心肌细胞肥大,其机制可能与凋亡和内质网应激的下调有关。

AIM To investigate the protective effects of a phosphodiesterase 5(PDE5) inhibitor against isoproterenol(Iso)-induced cardiomyocyte hypertrophy in neonatal rats. METHODS Rat cardiomyocytes were isolated and divided into a control(Con) group, an isoproterenol(Iso) group and an isoproterenol+sildenafil(Iso+Sil) group. The cell viability and lactate dehydrogenation(LDH) were detected by each group. The concentration of Sil in subsequent experimens was determined. Measurement of cardiac atrial natriuretic peptide(ANP) and β-myosin heavy chain(β-MHC) mRNA content by RT-PCR, flow cytometry and Western blot were used to detect apoptosis or endoplasmic reticulum stress-related protein glucose-regulated protein 78(GRP78) and CCAAT enhancer-binding protein homologous protein(CHOP) levels. RESULTS Compared with the Con group, the cell viability of the Iso group was decreased(P<0.01), and the release of LDH was increased(P<0.05). Compared with the Iso group, a certain concentration of Sil pretreatment increased cell viability and reduced LDH(P<0.05). The release achieved maximum effect at 5 μmol/L Sil. Compared with the Con group, the Iso group increased the expression of ANP and β-MHC mRNA, up-regulated the apoptosis rate, and increased the protein levels of GRP78 and CHOP. Compared with the Iso group, Sil pretreatment reduced the expression of ANP and β-MHC mRNA, down-regulation of apoptosis rate and inhibition of GRP78 and CHOP protein expression. CONCLUSION The PDE5 inhibitor Sil significantly inhibited Iso-induced cardiomyocyte hypertrophy in neonatal rats and its mechanism may be related to down-regulation of apoptosis and endoplasmic reticulum stress.