Proteomic Analysis of Exosomes Secreted from Human Alpha-1 Antitrypsin Overexpressing Mesenchymal Stromal Cells.
Proteomic Analysis of Exosomes Secreted from Human Alpha-1 Antitrypsin Overexpressing Mesenchymal Stromal Cells.
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DOI：
10.3390/biology11010009
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发表时间：
2021-12-21
期刊：
影响因子：
4.2
通讯作者：
Wang H
中科院分区：
文献类型：
--
作者：
Wei H;Green E;Ball L;Fan H;Lee J;Strange C;Wang H
人α-1抗胰蛋白酶（hAAT）在间充质基质细胞（MSC）中的过表达改善了其内在特性，在先前的1型糖尿病小鼠模型中具有增强的保护作用。本研究比较了骨髓来源对照或hAAT-MSC的细胞外囊泡（EV）的蛋白质谱。来自两种细胞类型的EV在大小和外泌体标志物表达方面具有共同特征。通过使用基因本体（GO）分析比较来自三个供体细胞系的EV中的共同蛋白，我们发现来自所有供体的共同EV蛋白对细胞粘附和细胞外基质组织是重要的。来自hAAT-MSC和MSC的差异表达的蛋白质参与免疫系统的细胞因子信号传导、干细胞分化和碳水化合物代谢。该研究表明，与对照MSC相比，hAAT-MSC具有不同的旁分泌效应外泌体蛋白谱。细胞外囊泡（EV）在细胞治疗期间介导干细胞的许多治疗作用。制造骨髓来源的间充质基质细胞（BM-MSC）以过表达人抗蛋白酶α-1抗胰蛋白酶（hAAT），并研究以比较与慢病毒处理的对照MSC相比的EV产生。本研究的目的是使用无偏的高分辨率液相色谱和质谱法比较对照和hAAT-MSC的EV/外泌体中的蛋白质谱以探索差异。纳米颗粒跟踪分析（NTA）显示来自对照MSC或hAAT-MSC的EV的颗粒尺寸在30至200 nm范围内。MSC和hAAT-MSC均表达外泌体相关蛋白，包括CD 63、CD 81和CD 9。hAAT-MSC也表达高水平的hAAT。接下来，我们对来自三个健康供体细胞系的EV进行了蛋白质组学分析。通过GO分析对从细胞上清液收集的外泌体进行分类，GO分析显示了对细胞粘附和细胞外基质组织重要的蛋白质。然而，来自对照MSC和hAAT-MSC的外来体在免疫系统的细胞因子信号传导、干细胞分化和碳水化合物代谢方面存在差异（p < 0.05）。这些结果表明，hAAT-MSC外泌体含有不同谱的旁分泌效应物，其具有改变的免疫功能，对MSC干细胞性、分化和细胞凋亡和存活的预防的影响，这可能有助于改善治疗功能。
The overexpression of human alpha-1 antitrypsin (hAAT) in mesenchymal stromal cells (MSCs) improved their intrinsic properties with enhanced protective effects in previous murine models of type 1 diabetes. This study compared the protein profiles of extracellular vesicles (EVs) from bone marrow-derived control or hAAT-MSCs. EVs from both cell types share common features in size and exosome marker expression. By comparing common proteins in EVs from three donor cell lines using Gene Ontology (GO) analysis, we found that common EV proteins from all donors are important to cell adhesion and extracellular matrix organization. Differentially expressed proteins from hAAT-MSCs and MSCs are involved in cytokine signaling of the immune system, stem cell differentiation, and carbohydrate metabolism. This study shows that hAAT-MSCs have different profiles of paracrine effector exosomal proteins compared to control MSCs. Extracellular vesicles (EVs) mediate many therapeutic effects of stem cells during cellular therapies. Bone marrow-derived mesenchymal stromal cells (BM-MSCs) were manufactured to overexpress the human antiprotease alpha-1 antitrypsin (hAAT) and studied to compare the EV production compared to lentivirus treated control MSCs. The goal of this study was to compare protein profiles in the EVs/exosomes of control and hAAT-MSCs using unbiased, high resolution liquid chromatography and mass spectrometry to explore differences. Nanoparticle tracking analysis (NTA) showed that the particle size of the EVs from control MSCs or hAAT-MSCs ranged from 30 to 200 nm. Both MSCs and hAAT-MSCs expressed exosome-associated proteins, including CD63, CD81, and CD9. hAAT-MSCs also expressed high levels of hAAT. We next performed proteomic analysis of EVs from three healthy donor cell lines. Exosomes collected from cell supernatant were classified by GO analysis which showed proteins important to cell adhesion and extracellular matrix organization. However, there were differences between exosomes from control MSCs and hAAT-MSCs in cytokine signaling of the immune system, stem cell differentiation, and carbohydrate metabolism (p < 0.05). These results show that hAAT-MSC exosomes contain a different profile of paracrine effectors with altered immune function, impacts on MSC stemness, differentiation, and prevention of cell apoptosis and survival that could contribute to improved therapeutic functions.
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