Department of Cardiovascular Surgery, The First Affiliated Hospital of Nanchang University, Nanchang 330006, Jiangxi, China.
Department of Cardiovascular Surgery, The First Affiliated Hospital of Nanchang University, Nanchang 330006, Jiangxi, China.
Heart Lung Circ. 2014 Feb;23(2):152-8. doi: 10.1016/j.hlc.2013.07.004. Epub 2013 Aug 13.
Recent studies have demonstrated that Notch signalling pathway is an important mediator of cardiac repair and regeneration after myocardial infarction. However, the mechanism by which Notch signalling pathway is mediating cardioprotection after ischaemic postconditioning (IPost) is still not understood thoroughly. The aim of the present study was to investigate the mechanism by which Notch signalling pathway mediated the cardioprotection effect after IPost.
Rat heart-derived H9c2 cells were randomly divided into six groups as follows: Control group, hypoxia/reoxygenation group (H/R), H/R+N1ICD group, H-post group, H-post+Notch-1miRNA group, and Mock group. We used pcDNA3.1-Myc-His plasmid and RNA interference (RNAi) to activate/inhibit the expression of Notch-1 in H9c2 cell lines. The Bcl-2, Bax genes and proteins were assessed by quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) and Western blot analysis. The effects of Notch 1 signalling on cell survival, proliferation and apoptosis were detected by 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide (MTT) and flow cytometry analysis, respectively. Furthermore, Notch 1 signalling induced the disruption of mitochondrial membrane potential, thus leading to the activation of caspase-9/-3 measured using the colorimetric activity assay.
We found Notch 1 signalling reduced cardiomyocyte apoptosis in IPost through regulating the expression of Bcl-2, Bax and activation of caspase-9 and -3. We found that after transfected with pcDNA3.1-Myc-His plasmid, activation of the Notch 1 gene effectively promoted cell proliferation and inhibited apoptosis. The Notch 1 upregulation was accompanied by an upregulation of Bcl-2 and a downregulation of Bax. In addition, a paralled increase in caspase-9/-3 activities was observed. These effects were blunted by transfected with Notch-1 miRNA in the H9c2 cells.
Notch 1 signalling has a cardioprotection effect, which may result from cardiomyocyte apoptosis, by means of regulating the expression of cell apoptosis inhibiting proteins Bcl-2, Bax and the activation of caspase-9 and -3.
最近的研究表明，Notch 信号通路是心肌梗死后心脏修复和再生的重要介质。然而，Notch 信号通路在缺血后处理（IPost）介导心脏保护的确切机制仍不清楚。本研究旨在探讨 Notch 信号通路介导 IPost 后心脏保护作用的机制。
将大鼠心脏源性 H9c2 细胞随机分为 6 组：对照组、缺氧/复氧组（H/R）、H/R+N1ICD 组、H-post 组、H-post+Notch-1miRNA 组和 Mock 组。我们使用 pcDNA3.1-Myc-His 质粒和 RNA 干扰（RNAi）激活/抑制 H9c2 细胞系中 Notch-1 的表达。通过实时定量聚合酶链反应（qRT-PCR）和 Western blot 分析评估 Bcl-2、Bax 基因和蛋白。通过 3-（4,5）-二甲基噻唑（-z-y1）-3,5-二苯基四唑溴盐（MTT）和流式细胞术分析分别检测 Notch 1 信号对细胞存活、增殖和凋亡的影响。此外，Notch 1 信号诱导线粒体膜电位破坏，从而导致 caspase-9/-3 的激活，通过比色活性测定法进行测量。
我们发现，通过调节 Bcl-2、Bax 的表达和 caspase-9/-3 的激活，IPost 中的 Notch 1 信号可减少心肌细胞凋亡。我们发现，转染 pcDNA3.1-Myc-His 质粒后，Notch 1 基因的激活可有效促进细胞增殖并抑制凋亡。Notch 1 的上调伴随着 Bcl-2 的上调和 Bax 的下调。此外，还观察到 caspase-9/-3 活性的平行增加。在 H9c2 细胞中转染 Notch-1 miRNA 可减弱这些作用。
Notch 1 信号具有心脏保护作用，可能通过调节细胞凋亡抑制蛋白 Bcl-2、Bax 的表达和 caspase-9/-3 的激活来实现心肌细胞凋亡。
