脂多糖刺激多能干细胞诱导人源心肌细胞与大鼠心肌细胞的体外模型比较

目的比较脂多糖(lipopolysaccharides,LPS)对多能干细胞诱导的人源心肌细胞(human induced pluripotent stem cell-derived cardiomyocytes,hiPS-CMs)及原代新生大鼠心肌细胞(cardiomyocytes,CMs)的影响。方法不同浓度LPS处理hiPS-CMs和原代新生大鼠CMs 24~48 h,通过实时无标记细胞分析技术(xCELLigence Real-Time Cell Analyser Cardio system RTCA)观察hiPS-CMs和原代新生大鼠CMs的电生理变化;qRT-PCR方法检测NPPB mRNA表达水平和qPCR阵列法检测炎性基因表达水平。结果不同浓度LPS刺激后,原代新生大鼠CMs的电生理变化为搏动频率增加和搏动幅度减少(P<0.01),NPPB mRNA表达水平增加(P<0.01)。在hiPS-CMs中,相对应的LPS浓度刺激未能引起搏动幅度和搏动频率显著变化(P>0.05),NPPB mRNA表达水平未显著增加(P>0.05)。进一步增加浓度(2.5μg/mL~40μg/mL),hiPS-CMs的搏动幅度和搏动频率仍未发生明显变化(P>0.05),NPPB mRNA在高浓度LPS(5μg/mL~40μg/mL)发生差异性改变(P<0.01)。最后,在炎症相关基因表达方面,原代新生大鼠CMs表现为C3、Gpnmb、Atf3、Il6r和Ly96基因水平上调至1.5倍,hiPS-CMs表现为AK4、TOLLIP、SPP1、FABP1、IL6R、LY96和C3基因水平上调至1.5倍。结论与原代新生大鼠CMs相比,hiPS-CMs受到LPS的损伤明显减轻,表现出不同的炎症基因表达模式。

Human induced pluripotent stem cell-derived cardiomyocytes as an in vitro model for lipopolysaccharide-induced cardiomyopathy comparison with primary neonatal rat cardiomyocytes

Objective:To investigate the effect of lipopolysaccharide (LPS) on primary neonatal rat cardiomyocytes (CMs) and human induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs).Methods:The hiPS-CMs and primary neonatal rat CMs were treated with different concentrations of LPS for 24 to 48 h. Then the cellular viability was analyzed by the xCELLigence RTCA Cardio system. The measurement of NPPB gene was studied by qRT-PCR and the gene expression analysis was performed by the qPCR array, in order to evaluate the cardiac inflammation effect induced by LPS.Results:The LPS exposure led to dysfunction in the primary neonatal rat CMs, which shown as an increase in beating rate and a decrease in contraction amplitude ( P<0.01), accompanied by an increased NPPB mRNA level ( P<0.01). There was no significant alteration in beating rate and the contraction amplitude in the corresponding concentration of the primary neonatal rat CMs ( P>0.05), as well as the NPPB mRNA level ( P>0.05). However, the expression of NPPB mRNA in hiPS-CMs was significantly different at a higher concentration of LPS (5 μg/mL~40 μg/mL) ( P<0.01), but the beating rate and the contraction amplitude showed no significant change, even the concentration of LPS up to 40 μg/mL ( P>0.05). Finally, the genes of C3, Gpnmb, Atf3, Il6r and Ly96 upregulated to 1.5 folds in the primary neonatal rat CMs. In comparison with primary neonatal rat CMs, the AK4, TOLLIP, SPP1, FABP1, IL6R, LY96 and C3 were over expression to 1.5 folds in the hiPS-CMs. Conclusions:In comparison with primary neonatal rat CMs, hiPS-CMs are markedly less injured by LPS and show a different pattern of inflammation gene expression.