β Adrenergic Receptor Kinase C-Terminal Peptide Gene-Therapy Improves β2-Adrenergic Receptor-Dependent Neoangiogenesis after Hindlimb Ischemia
β Adrenergic Receptor Kinase C-Terminal Peptide Gene-Therapy Improves β2-Adrenergic Receptor-Dependent Neoangiogenesis after Hindlimb Ischemia
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DOI：
10.1124/jpet.115.228411
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发表时间：
2016-02-01
影响因子：
3.5
通讯作者：
Rengo, Giuseppe
中科院分区：
文献类型：
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作者：
Cannavo, Alessandro;Liccardo, Daniela;Rengo, Giuseppe
后肢缺血（HI）后，缺血肌肉内儿茶酚胺水平升高可引起β(2)-肾上腺素能受体（β (2)AR）信号失调，导致血运重建减少。事实上，在大鼠HI模型中，通过基因治疗体内β (2)AR过表达可增强血管生成。G蛋白偶联受体激酶2 （GRK2）是β - AR信号的关键调节因子，而β肾上腺素能受体激酶c末端肽（β ARKct）是GRK2的肽抑制剂，已被证明可以阻止β AR下调，并通过恢复β (2)AR保护信号（即蛋白激酶B/内皮型一氧化氮合酶）来保护心肌细胞和干细胞免受缺血损伤。在此，我们测试了腺病毒介导的β - ARKct基因转移在HI实验模型中的潜在治疗作用，以及它对β - AR信号传导和内皮细胞（EC）功能的影响。因此，在本研究中，我们通过股动脉切除术（FAR）手术诱导大鼠HI。15天的缺血导致β - AR显著下调，与此同时，缺血肌肉中GRK2水平增加约2倍。重要的是，与对照组相比，FAR时大鼠后肢β - ARKct的体内基因转移导致后肢灌注明显改善，缺血肌肉的毛细血管和β - AR密度增加。在体外培养的内皮细胞中也评估了β - ARKct表达的影响。有趣的是，表达β ARKct非诺特罗（β (2)AR激动剂）的内皮细胞诱导β (2)AR促血管生成信号增强和EC功能增强。我们的研究结果表明，β - ARKct基因治疗和随后的GRK2抑制通过阻止缺血诱导的β (2)AR下调来促进HI模型中的血管生成。
After hindlimb ischemia (HI), increased catecholamine levels within the ischemic muscle can cause dysregulation of beta(2)-adrenergic receptor (beta(2)AR) signaling, leading to reduced revascularization. Indeed, in vivo beta(2)AR overexpression via gene therapy enhances angiogenesis in a rat model of HI. G protein-coupled receptor kinase 2 (GRK2) is a key regulator of beta AR signaling, and beta adrenergic receptor kinase C-terminal peptide (beta ARKct), a peptide inhibitor of GRK2, has been shown to prevent beta AR down-regulation and to protect cardiac myocytes and stem cells from ischemic injury through restoration of beta(2)AR protective signaling (i.e., protein kinase B/endothelial nitric oxide synthase). Herein, we tested the potential therapeutic effects of adenoviral-mediated beta ARKct gene transfer in an experimental model of HI and its effects on beta AR signaling and on endothelial cell (EC) function in vitro. Accordingly, in this study, we surgically induced HI in rats by femoral artery resection (FAR). Fifteen days of ischemia resulted in significant beta AR down-regulation that was paralleled by an approximately 2-fold increase in GRK2 levels in the ischemic muscle. Importantly, in vivo gene transfer of the beta ARKct in the hindlimb of rats at the time of FAR resulted in a marked improvement of hindlimb perfusion, with increased capillary and beta AR density in the ischemic muscle, compared with control groups. The effect of beta ARKct expression was also assessed in vitro in cultured ECs. Interestingly, ECs expressing the beta ARKct fenoterol, a beta(2)AR-agonist, induced enhanced beta(2)AR proangiogenic signaling and increased EC function. Our results suggest that beta ARKct gene therapy and subsequent GRK2 inhibition promotes angiogenesis in a model of HI by preventing ischemia-induced beta(2)AR down-regulation.