Purpose

Previous study indicated the protective role of cilostazol in ischemia–reperfusion (I/R) injury. Here, we aimed to explore the function of cilostazol in myocardial I/R injury and the underlying mechanism.

Methods

The myocardial I/R injury rat model was constructed, and the expression levels of vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), basic fibroblast growth factor (bFGF), platelet-derived growth factor receptor b (PDGF-B) and the number of new blood vessels were measured by qRT-PCR and immunohistochemistry. VSMC and HUVEC cells were treated with hypoxia to induce in vivo I/R injury model. The protein expression of AKT, endothelial nitric oxide synthase (eNOS) and apoptosis-related protein levels were detected by western blotting. Besides, the positive staining rate and cell viability were tested by 5-bromo-2-deoxyuridine (Brdu)/4′,6-diamidino-2-phenylindole (DAPI) or DAPI/TdT-mediated dUTP Nick-End Labeling (TUNEL) staining and MTT assay.

Results

Cilostazol promoted angiogenesis by increasing the number of new blood vessels and up-regulating the expression of VEGF, HGF, bFGF and PDGF-B in myocardial I/R-injury rat model. The in vitro experiments showed that cilostazol increased the level of eNOS and AKT, and also enhanced cell proliferation in hypoxia-treated VSMC and HUVEC cells. Furthermore, after 8-Br-cAMP treatment, VEGF, HGF, bFGF, PDGF-B, p-AKT and p-eNOS expression were up-regulated, while cleaved-caspase 3 and cleaved-PARP expression were down-regulated. In addition, the effects of cilostazol on cell viability and apoptosis were aggravated by 8-Br-cAMP and attenuated after KT-5720 treatment.

Conclusion

Cilostazol could promote angiogenesis, increase cell viability and inhibit cell apoptosis, consequently protecting myocardial tissues against I/R-injury through activating cAMP.

中文翻译：

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西洛他唑通过cAMP依赖性机制促进心肌缺血/再灌注损伤后的血管生成并增加细胞增殖。

目的

先前的研究表明西洛他唑在缺血/再灌注（I / R）损伤中的保护作用。在这里，我们旨在探讨西洛他唑在心肌I / R损伤中的功能及其潜在机制。

方法

建立了心肌I / R损伤大鼠模型，并表达了血管内皮生长因子（VEGF），肝细胞生长因子（HGF），碱性成纤维细胞生长因子（bFGF），血小板衍生生长因子受体b（PDGF-B）的表达水平），并通过qRT-PCR和免疫组织化学测量新血管的数量。用缺氧处理VSMC和HUVEC细胞以诱导体内I / R损伤模型。Western blot检测AKT蛋白表达，内皮型一氧化氮合酶（eNOS）和凋亡相关蛋白水平。此外，通过5-溴-2-脱氧尿苷（Brdu）/ 4'，6-二mid基-2-苯基吲哚（DAPI）或DAPI / TdT介导的dUTP缺口末端标记（TUNEL）检测阳性染色率和细胞活力。染色和MTT测定。

结果

西洛他唑通过增加新血管数量并上调心肌I / R损伤大鼠模型中VEGF，HGF，bFGF和PDGF-B的表达来促进血管生成。的体外实验表明，西洛他唑增加eNOS和AKT的水平，并且也增强了在缺氧处理的VSMC和HUVEC细胞的细胞增殖。此外，在8-Br-cAMP处理后，VEGF，HGF，bFGF，PDGF-B，p-AKT和p-eNOS的表达上调，而裂解的半胱天冬酶3和裂解的PARP表达下调。另外，西洛他唑对细胞活力和细胞凋亡的影响被8-Br-cAMP加重，并在KT-5720处理后减弱。

结论

西洛他唑可以促进血管生成，增加细胞活力并抑制细胞凋亡，从而通过激活cAMP保护心肌组织免受I / R损伤。