Department of Nutrition and Health Sciences, University of Nebraska-Lincoln, 230 Filley Hall, Lincoln, NE, 68583, USA.
Department of Food Science and Technology, University of Nebraska-Lincoln, 260 Food Innovation Center, Lincoln, NE, 68588, USA.
J Nanobiotechnology. 2023 Sep 28;21(1):352. doi: 10.1186/s12951-023-02105-9.
Macrophages are highly plastic innate immune cells that play key roles in host defense, tissue repair, and homeostasis maintenance. In response to divergent stimuli, macrophages rapidly alter their functions and manifest a wide polarization spectrum with two extremes: M1 or classical activation and M2 or alternative activation. Extracellular vesicles (EVs) secreted from differentially activated macrophages have been shown to have diverse functions, which are primarily attributed to their microRNA cargos. The role of protein cargos in these EVs remains largely unexplored. Therefore, in this study, we focused on the protein cargos in macrophage-derived EVs.
Naïve murine bone marrow-derived macrophages were treated with lipopolysaccharide or interlukin-4 to induce M1 or M2 macrophages, respectively. The proteins of EVs and their parental macrophages were subjected to quantitative proteomics analyses, followed by bioinformatic analyses. The enriched proteins of M1-EVs were involved in proinflammatory pathways and those of M2-EVs were associated with immunomodulation and tissue remodeling. The signature proteins of EVs shared a limited subset of the proteins of their respective progenitor macrophages, but they covered many of the typical pathways and functions of their parental cells, suggesting their respective M1-like and M2-like phenotypes and functions. Experimental examination validated that protein cargos in M1- or M2-EVs induced M1 or M2 polarization, respectively. More importantly, proteins in M1-EVs promoted viability, proliferation, and activation of T lymphocytes, whereas proteins in M2-EVs potently protected the tight junction structure and barrier integrity of epithelial cells from disruption. Intravenous administration of M2-EVs in colitis mice led to their accumulation in the colon, alleviation of colonic inflammation, promotion of M2 macrophage polarization, and improvement of gut barrier functions. Protein cargos in M2-EVs played a key role in their protective function in colitis.
This study has yielded a comprehensive unbiased dataset of protein cargos in macrophage-derived EVs, provided a systemic view of their potential functions, and highlighted the important engagement of protein cargos in the pathophysiological functions of these EVs.
巨噬细胞是具有高度可塑性的先天免疫细胞，在宿主防御、组织修复和维持内环境稳定方面发挥着关键作用。巨噬细胞在应对不同的刺激时，会迅速改变其功能，并表现出广泛的极化谱，其中两个极端是 M1 或经典激活和 M2 或替代激活。已证实，来自不同激活的巨噬细胞分泌的细胞外囊泡（EVs）具有多种功能，这些功能主要归因于它们的 microRNA cargos。这些 EVs 中的蛋白质 cargos 的作用在很大程度上仍未得到探索。因此，在这项研究中，我们专注于巨噬细胞衍生的 EVs 中的蛋白质 cargos。
用脂多糖或白细胞介素-4 分别处理幼稚的鼠骨髓来源的巨噬细胞，以诱导 M1 或 M2 巨噬细胞。EVs 和其亲本巨噬细胞的蛋白质进行定量蛋白质组学分析，并进行生物信息学分析。M1-EVs 的富集蛋白参与促炎途径，而 M2-EVs 的富集蛋白与免疫调节和组织重塑有关。EVs 的特征蛋白与各自亲本巨噬细胞的蛋白质有有限的子集重叠，但它们涵盖了其亲本细胞的许多典型途径和功能，表明它们各自的 M1 样和 M2 样表型和功能。实验验证表明，M1-或 M2-EVs 中的蛋白质 cargos 分别诱导 M1 或 M2 极化。更重要的是，M1-EVs 中的蛋白质促进 T 淋巴细胞的活力、增殖和激活，而 M2-EVs 中的蛋白质则有效地保护上皮细胞的紧密连接结构和屏障完整性免受破坏。在结肠炎小鼠中静脉注射 M2-EVs 导致其在结肠中积累，缓解结肠炎症，促进 M2 巨噬细胞极化，并改善肠道屏障功能。M2-EVs 中的蛋白质 cargos 在其对结肠炎的保护功能中发挥了关键作用。
本研究提供了一个全面的、无偏倚的巨噬细胞衍生 EVs 中蛋白质 cargos 的数据集，系统地观察了它们的潜在功能，并强调了蛋白质 cargos 在这些 EVs 的病理生理功能中的重要作用。
