PEP-1-MsrA ameliorates inflammation and reduces atherosclerosis in apolipoprotein E deficient mice.
PEP-1-MsrA ameliorates inflammation and reduces atherosclerosis in apolipoprotein E deficient mice.
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DOI：
10.1186/s12967-015-0677-8
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发表时间：
2015-09-26
影响因子：
7.4
通讯作者：
Yu H
中科院分区：
文献类型：
--
作者：
Wu Y;Xie G;Xu Y;Ma L;Tong C;Fan D;Du F;Yu H
蛋氨酸亚砜还原酶A （MsrA）是一种有效的细胞内氧化还原酶，是保护细胞免受氧化损伤的重要因素。然而，外源性MsrA在氧化应激诱导疾病中的治疗应用是有限的，因为它不能进入细胞。本研究的目的是研究带有PEP-1（一种细胞穿透肽）融合到其n端的MsrA是否可以保护巨噬细胞免受氧化应激，并可以减轻载脂蛋白E缺陷（apoE−/−）小鼠的动脉粥样硬化。利用pET28a表达系统对MsrA和融合蛋白PEP-1-MsrA进行表达和纯化。Western blot和免疫荧光染色证实融合蛋白转染巨噬细胞。流式细胞术检测细胞内活性氧（ROS）和细胞凋亡水平。在体内研究中，将MsrA或PEP-1-MsrA蛋白腹腔注射到喂食西方饮食的apoE−/−小鼠中12周。评估血脂水平、炎症基因表达、对氧磷酶-1 （PON1）和超氧化物歧化酶（SOD）活性。采用油红O染色和免疫组织化学分析动脉粥样硬化病变。PEP-1-MsrA能穿透细胞，显著降低h2o2处理巨噬细胞的细胞内ROS水平和细胞凋亡。降低脂多糖处理巨噬细胞TNFα和IL-1β mRNA水平，提高IL-10 mRNA水平。在体内研究中，与注射对照或MsrA相比，PEP-1-MsrA显著提高了血浆PON1和SOD活性，降低了血浆单核细胞趋化蛋白1 （MCP-1）水平。注射PEP-1-MsrA后，小鼠肝脏PON1水平升高，肝脏TNFα和IL-6 mRNA表达受到抑制。虽然血浆总胆固醇和甘油三酯水平没有改变，但PEP-1-MsrA治疗小鼠的主动脉粥样硬化明显减少。这伴随着病变中巨噬细胞总数和凋亡的减少。我们的研究提供了证据，证明PEP-1-MsrA可能是动脉粥样硬化相关心血管疾病的潜在治疗剂。本文的在线版本（doi:10.1186/s12967-015-0677-8）包含补充材料，可供授权用户使用。
Methionine sulfoxide reductase A (MsrA) is a potent intracellular oxidoreductase and serves as an essential factor that protects cells against oxidative damage. However, therapeutic use of exogenous MsrA in oxidative stress-induced diseases is limited, because it cannot enter the cells. The aim of this study is to investigate whether MsrA with PEP-1, a cell penetrating peptide, fused to its N-terminus can protect against oxidative stress in macrophages and can attenuate atherosclerosis in apolipoprotein E deficient (apoE−/−) mice. MsrA and the fusion protein PEP-1-MsrA were expressed and purified using a pET28a expression system. Transduction of the fusion protein into macrophages was confirmed by Western blot and immunofluorescence staining. Intracellular reactive oxygen species (ROS) and apoptosis levels were measured by flow cytometry. In in vivo study, MsrA or PEP-1-MsrA proteins were intraperitoneally injected into apoE−/− mice fed a Western diet for 12 weeks. Plasma lipids levels, inflammatory gene expression, and paraoxonase-1 (PON1) and superoxide dismutase (SOD) activities were assessed. Atherosclerotic lesions were analyzed by Oil Red O staining and immunohistochemistry. PEP-1-MsrA could penetrate the cells and significantly reduced intracellular ROS levels and apoptosis in H2O2-treated macrophages. It also decreased TNFα and IL-1β mRNA levels and increased the IL-10 mRNA level in lipopolysaccharide-treated macrophages. In in vivo study, PEP-1-MsrA injection significantly increased plasma PON1 and SOD activities and decreased plasma monocyte chemoattractant protein 1 (MCP-1) level compared to the injection of vehicle control or MsrA. In PEP-1-MsrA injected mice, hepatic PON1 levels were increased, while the expression of TNFα and IL-6 mRNA in the liver was suppressed. Although plasma total cholesterol and triglyceride levels did not change, the aortic atherosclerosis in PEP-1-MsrA treated mice was significantly reduced. This was accompanied by a reduction of total and apoptotic macrophages in the lesions. Our study provides evidence that PEP-1-MsrA may be a potential therapeutic agent for atherosclerosis-related cardiovascular diseases. The online version of this article (doi:10.1186/s12967-015-0677-8) contains supplementary material, which is available to authorized users.