多肽P-EKKE的生物学特征分析及其在心脏再生中的功能与机制研究

The heart of neonatal mouse has a certain ability to regenerate, and born after seven days will lose this ability. P-EKKE is an endogenous peptides identified by mass spectrometry without any function report. It is highly expressed in newborn mouse myocardium, and highly conserved among species. In previous studies we found that P-EKKE located in the nucleus and was stable in vivo; P-EKKE could significantly promote the H9c2 cells proliferation and cycle related gene expression ; Vivo study in mice showed that P-EKKE tail vein injection can promote heart regeneration after myocardial injury; pull-down experiments found P-EKKE can interact with Sufu, Sufu is one of the key proteins in the hedgehog signaling pathway, and hedgehog signaling pathway was reported to be closely related to cardiac regeneration. So we speculate that P-EKKE may play an important role in cardiac regeneration. In this study we intend to: ① Further clarify the spatiotemporal expression pattern of P-EKKE in mice before and after birth, before and after myocardial infarction; ②Separate cultured primary myocardial cells, to observe the effect of P-EKKE on proliferation and hypoxia tolerance of primary cardiocytes;③ Establishe animal models of apical resection and myocardial infarction, to evaluate the role of P-EKKE in heart regeneration, and elucidate the specific mechanism from the point of Sufu-Hedgehog pathway systematically. This study can provide a new opportunity for the treatment of myocardial infarction.

新生小鼠心脏具有一定的再生能力，而出生七天后将基本丧失这种能力。P-EKKE是我们通过质谱分析筛选出的一条在新生小鼠心肌中高表达、功能未知的多肽。前期发现P-EKKE定位于细胞核内；显著促进H9c2细胞增殖及周期相关基因表达，在体研究表明P-EKKE促进小鼠损伤后的心脏修复；Pull-Down实验提示P-EKKE与Sufu相互作用，Sufu是Hedgehog信号通路中关键蛋白，而后者与心脏再生密切相关。因此我们推测P-EKKE在心脏再生中可能发挥重要作用。本研究拟：①阐明P-EKKE在小鼠出生前后、心肌梗死前后的时空表达特征。②培养原代心肌细胞，观察P-EKKE对原代心肌细胞的增殖、周期、缺氧耐受的影响。③建立小鼠心尖切除与心肌梗死模型，评估P-EKKE在心脏再生中的作用；以Sufu-Hedgehog信号通路为切入点，系统论证其促进心脏再生的具体机制。本研究可为心肌梗死的治疗提供新契机。

背景：心脏再生是目前心血管科学领域的研究热点。新生小鼠的心脏具有一定的再生能力，而出生七天后将基本丧失这种能力。P-EKKE是我们通过质谱分析筛选出的一条在新生小鼠心肌中高表达、功能未知的内源性多肽，其在物种间高度保守。前期研究发现P-EKKE定位于细胞核内，在体内能够稳定存在；P-EKKE能够显著促进H9c2细胞的增殖以及周期相关基因的表达。Pull-Down实验提示P-EKKE能够与Sufu相互作用，Sufu是Hedgehog信号通路中的关键蛋白之一，而Hedgehog信号通路与心脏再生密切相关。.研究内容：阐明P-EKKE在小鼠出生前后、心肌梗死前后的时空表达特征；分离培养原代心肌细胞，观察P-EKKE对原代心肌细胞周期、增殖分裂、缺氧耐受的影响；建立心尖切除与心肌梗死两种动物模型，综合评估P-EKKE在心脏再生中的作用，同时以Sufu-Hedgehog信号通路为切入点，系统论证P-EKKE促进心脏再生的具体机制。.研究结果：P-EKKE能够促进原代心肌细胞由G1期进入S期，调节周期相关基因的表达，从而促进细胞增殖；P-EKEK能够抑制缺氧环境诱导的细胞凋亡；P-EKKE能够刺激损伤后的心肌再生，促进心脏修复；P-EKKE可促进心肌梗死后心功能恢复，减少心肌梗死面积，促进梗死后心肌细胞的增殖、分裂，抑制氧化应激引起的细胞凋亡；P-EKKE能够调节Hedgehog信号通路中关键基因与蛋白的表达。.科学意义：迄今国内外并无P-EKKE在心脏再生中功能与机制的报道，因此本研究具有源头创新性，有助于进一步理解心脏再生的的分子机制，并可能为心肌梗死的治疗提供新的契机。