Intrinsic activation of cardiosphere-derived cells enhances myocardial repair
Intrinsic activation of cardiosphere-derived cells enhances myocardial repair
复制标题
心脏源性细胞的内在激活增强心肌修复
DOI：
10.1016/j.jtcvs.2020.05.040
复制
发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Sano Shunji
中科院分区：
文献类型：
--
作者：
Sano Toshikazu;Ito Tatsuo;Ishigami Shuta;Bandaru Srinivas;Sano Shunji
心肌梗死后心肌细胞的永久性丧失会导致心脏功能不可逆的损害。本研究旨在通过内源性激活心肌特异性分化因子，提高心球源性细胞（CDCs）向心肌细胞分化的效率，使其向大群体心肌细胞分化（Gata4，Mef2c，Nkx2-5，Hand2，和Tnnt 2）通过CRISPR/dCas 9辅助的转录增强系统。通过与转录激活因子VP 64融合的CRISPR/dCas 9系统（CRISPR-dCas 9-VP 64），将负责靶基因的转录激活的CRISPR/dCas 9（增强子和启动子）与靶基因的转录激活相结合。在大鼠急性心肌梗死模型中，我们比较了有或没有转录activation.ResultsWe的再生潜力和功能的好处CDC确定了一组特定的CRISPR RNA靶向增强子和启动子，表现出显着较高的表达分化因子ofGata 4，手2，和Tnnt 2。具有转录激活因子VP 64的CDC组（CDC与VP 64）显示左心室射血分数显著改善（61.9%vs52.5%and44.1%in CDC without transcriptional activation group and control）和减少心脏瘢痕面积。通过CRISPR/dCas 9基因转录系统辅助的特异性分化因子。CRISPR/dCas 9系统可以提供一种有效的调节干细胞内Tnnt 2基因激活的方法。因此，该系统可用于增强移植的CDCs在缺血心肌内的分化潜能，以更好地治疗缺血性心脏病患者。
ObjectivePermanent loss of cardiomyocytes after myocardial infarction results in irreversible damage to cardiac function. The present study aims to enhance the cardiomyogenic efficiency of cardiosphere-derived cells (CDCs) to develop into large populations of cardiomyocytes by intrinsic activation of cardio-specific differentiation factors (Gata4,Mef2c,Nkx2-5,Hand2, andTnnt2) by a CRISPR/dCas9 assisted transcriptional enhancement system.MethodsExhaustive screening was performed to identify the specific sequences in endogenous regulatory regions (enhancers and promoters) responsible for transcriptional activation of the target genes, via a CRISPR/dCas9 system fused with transcriptional activator VP64 (CRISPR-dCas9-VP64). In a rat model of acute myocardial infarction, we compared the regenerative potential and functional benefits of CDCs with or without transcriptional activation.ResultsWe identified a panel of specific CRISPR RNA targeting the enhancers and promoters, which demonstrated significantly higher expression of differentiation factors ofGata4,Hand2, andTnnt2. The group of CDCs with transcriptional activator VP64 (CDC with VP64) showed significant improvement in the left ventricular ejection fraction (61.9% vs 52.5% and 44.1% in the CDC without transcriptional activation group and control) and decreased scar area in the heart.ConclusionsWe have identified endogenous regulatory regions responsible for an intrinsic activation of cardio-specific differentiation factors assisted via a CRISPR/dCas9 gene transcriptional system. The CRISPR/dCas9 system may provide an efficient and effective means of regulatingTnnt2gene activation within stem cells. Subsequently, this system can be used to enhance transplanted CDCs differentiation potential within ischemic myocardia to better therapeutic outcomes of patients with ischemic heart disease.