289Pharmacologic rejuvenation of senescent human cardiac stem cells enhances myocardial repair
289Pharmacologic rejuvenation of senescent human cardiac stem cells enhances myocardial repair
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289 衰老人心脏干细胞的药理复兴可增强心肌修复
DOI：
10.1093/cvr/cvu087.3
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发表时间：
2014
影响因子：
10.8
通讯作者：
Avolio E
中科院分区：
文献类型：
--
作者：
Avolio E
目的：与衰老和疾病相关的细胞衰老阻碍了体外人c-Kit+心脏干细胞（CSC）的功能特性。本研究的目的是确定与CSC衰老相关的分子机制，并对后者进行快速调节，目的是提高CSC在vivo.Methods中的治疗潜力：从终末期衰竭心脏（E-CSC，n=20）和供移植心脏（D-CSC，n=14）的心房标本中分离c-Kit+ CSC。在体外比较D-和E-CSC的衰老标记物和分泌组的表达。本文分析了CSC分泌体对体外培养的成年大鼠心肌细胞（CM）模拟缺血-再灌注损伤的保护作用。研究了3条与细胞衰老相关的分子信号通路。用10 nM雷帕霉素和0.5μM白藜芦醇的混合物进行3天的药理学处理，以在体外逆转E-CSC衰老。移植后14天，比较了用载体（n=17）、D-CSC（n=17）、E-CSC（n=18）或药物调节的E-CSC（n=18）（300，000个细胞/心脏）注射的SCID-Beige小鼠梗死心脏的功能恢复。IL-1β的中和恢复了E-CSC的分泌组保护作用。在分子水平上，E-CSC的特征在于经典NFB途径和Caspase 1的超活化，TORC 1复合物的活性增加，自噬通量的损害以及AMPK、Akt和CREB活化的减少。所有这些改变可以成功地恢复采用雷帕霉素和白藜芦醇的鸡尾酒。药物治疗降低了体外衰老E-CSC的比例和IL-1β的分泌量，从而恢复了对大鼠CM的保护作用。最后，D-CSC而不是E-CSC能够增强心肌愈合时，注射在小鼠梗死心脏。然而，离体预处理的E-CSC与雷帕霉素和白藜芦醇，在体内transplantation之前，恢复其修复潜力的D-CSC level.Conclusions：我们首先证明，衰老的c-Kit+ CSC居民在人类衰竭的心脏显示受损的体内修复能力，重要的是，衰老的CSC可以在体外恢复活力，一个短的药理学条件，最后促进体内心脏再生。这些发现为改善心力衰竭的自体CSC治疗开辟了新途径。
Purpose:Cellular senescence associated with ageing and disease hampers the functional properties of human c-Kit+ Cardiac Stem Cells (CSC) in vitro. Aim of the study was to identify molecular mechanisms associated with CSC senescence and to pharmacologically modulate this latter, at the aim to improve the therapeutic potential of CSC in vivo.Methods:c-Kit+ CSC were isolated from atrial specimens of end-stage failing hearts (E-CSC, n=20) and hearts donated for transplantation (D-CSC, n=14). D- and E-CSC were compared in vitro for the expression of senescence markers and secretome. The ability of CSC's secretome to protect rat adult cardiomyocytes (CM) in vitro from a simulated ischemia-reperfusion injury was analysed. 3 molecular signalling pathways associated with cell senescence were investigated. A 3-days pharmacologic treatment with a cocktail of 10nM Rapamycin and 0.5μM Resveratrol was employed to revert E-CSC senescence in vitro. The functional recovery of SCID-Beige mice infarcted hearts injected with Vehicle (n=17), D-CSC (n=17), E-CSC (n=18) or drug-conditioned-E-CSC (n=18) (300,000 cells/heart) was compared 14 days post-transplantation.Results:with respect to D-CSC, E-CSC show, in vitro, higher levels of senescence markers (p16, γH2AX), a blunted proliferation and a secretome strongly enriched in the pro-inflammatory IL-1β, unable to protect rat CM from apoptosis and senescence in vitro. Neutralisation of IL-1β restored E-CSC's secretome protective effect. At the molecular level, E-CSC are characterized by an hyper-activation of the canonical NFB pathway and of Caspase1, an increased activity of the TORC1 complex, an impairment of the authophagic flux and a reduction of AMPK, Akt and CREB activation. All these alterations could be successfully reverted employing a cocktail of Rapamycin and Resveratrol. The drug treatment reduced the fraction of senescent E-CSC and the amount of IL-1β secreted in vitro, thus restoring the protective effect on rat CM. Last, D-CSC but not E-CSC were able to enhance myocardial healing when injected in a mouse infarcted heart. However, the ex vivo preconditioning of E-CSC with Rapamycin and Resveratrol, prior to the in vivo transplantation, restored their reparative potential to D-CSC levels.Conclusions:we first demonstrate that senescent c-Kit+ CSC resident in human failing hearts display an impaired in vivo reparative ability; importantly, senescent CSC can be rejuvenated in vitro with a short pharmacologic conditioning, finally boosting the in vivo cardiac regeneration. These findings open new avenues to improve autologous CSC therapy in heart failure.