RGFP966通过抑制YAP通路活化减轻长时程低温保存诱导的心功能损伤

RGFP966 inactivation of the YAP pathway attenuates cardiac dysfunction induced by prolonged hypothermic preservation

目的:观察组蛋白脱乙酰化酶3(HDAC3)抑制剂RGFP966是否能够促进低温保存心脏复灌期心功能的恢复,并探讨Hippo-YAP信号通路是否参与了RGFP966的心肌保护作用。创新点:首次证实了在Celsior保存液中添加RGFP966可以减轻长时程低温保存引起的心脏功能障碍。该研究结果为临床实践中的心脏移植提供有价值且可行的策略,为患者获得远距供体心脏提供了可能。方法:将大鼠离体心脏置于含有或不含有RGFP966的Celsior保存液中低温保存12 h后,于Langendorff灌流系统中复灌60 min,测定复灌期各项心功能指标。用Western blotting法分析Mst1和YAP蛋白表达和磷酸化水平。用TUNEL法测定心肌细胞凋亡情况。结论:RGFP966可以促进低温保存心脏复灌期心功能的恢复,其作用机制可能是通过抑制Hippo-YAP信号传导通路的激活,增强心肌抗氧化能力,抑制心肌细胞凋亡而实现的。

Oxidative stress and apoptosis are the key factors that limit the hypothermic preservation time of donor hearts to within 4–6 h.The aim of this study was to investigate whether the histone deacetylase 3(HDAC3)inhibitor RmGodFyPn9 a6 m6 i cc opualrda mpreotteercst daugrianing srt ecpaerrdfiuasci oinn juwreyr ei nedvuacleuda tebyd.pTrholeo nexgperde shsyipoont haenrdm pich opsrpehsoerryvlaattiioonn.leRvaet lsh eoaf rtms awmemrea lihayn-??STE20-like kinase-1(Mst1)and Yes-associated protein(YAP)were determined by western blotting.Cell apoptosis was measured by the terminal deoxynucleotidyl-transferase(TdT)-mediated dUTP nick-end labeling(TUNEL)method.Addition of RGFP966 in Celsior solution significantly inhibited cardiac dysfunction induced by hypothermic preservation.RGFP966 inhibited the hypothermic preservation-induced increase of the phosphorylated(p)-Mst1/Mst1 and p-YAP/YAP ratios,prevented a reduction in total YAP protein expression,and increased the nuclear YAP protein level.Verteporfin(VP),a small molecular inhibitor of YAP–transcriptional enhanced associate domain(TEAD)interaction,partially abolished the protective effect of RGFP966 on cardiac function,and reduced lactate dehydrogenase activity and malondialdehyde content.RGFP966 increased superoxide dismutase,catalase,and glutathione peroxidase gene and protein expression,which was abolished by VP.RGFP966 inhibited hypothermic preservation-induced overexpression of B-cell lymphoma protein 2(Bcl-2)-associated X(Bax)and cleaved caspase-3,increased Bcl-2 mRNA and protein expression,and reduced cardiomyocyte apoptosis.The antioxidant and anti-apoptotic effects of RGFP966 were cancelled by VP.The results suggest that supplementation of Celsior solution with RGFP966 attenuated prolonged hypothermic preservation-induced cardiac dysfunction.The mechanism may involve inhibition of oxidative stress and apoptosis via inactivation of the YAP pathway.