【解放军总医院】LPS预处理的MSCs通过exo调节巨噬细胞极化

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【解放军总医院】LPS预处理的MSCs通过外泌体传递let-7b调节炎症反应中巨噬细胞的极化

来自解放军总医院生命科学院分子免疫学研究室主任韩卫东教授今日在Journal of translational medicine杂志上发表文章，揭示了LPS预处理的MSCs通过外泌体传递let-7b调节炎症反应中巨噬细胞的极化。

最近几年的研究发现，LPS预处理的MSCs旁分泌作用显著增强，包括营养支持的增强和再生与修复能力的提高。MSCs可能分泌大量的外泌体用于细胞与细胞之间的交流并为组织损伤修复维持一个动态和自我平衡的内环境。该研究评价了LPS预处理的MSC来源的外泌体对慢性炎症和伤口修复的治疗效果与机制。

本研究的方法：首先利用梯度离心提取LPS预处理的MSCs的外泌体（LPS pre-Exo）。在体外，利用THP-1细胞高糖处理作为炎症反应模型，并用LPS pre-Exo处理48小时，利用real-time RT-PCR检测炎症相关的因子表达情况，利用免疫荧光检测巨噬细胞亚型的分布。利用miRNA芯片检测LPS pre-Exo中的miRNA表达谱。在体内，利用链唑霉素诱导的糖尿病大鼠建立皮肤损伤模型，将LPSpre-Exo弥散注射到伤口边缘，观察到LPS pre-Exo在炎症反应和伤口修复上的治疗作用。

本研究的结果：LPS pre-Exo可上调抗炎因子的表达和促进M2型巨噬细胞的激活。miRNA芯片筛选结果显示LPS pre-Exo中具有独特的let-7b的表达，而let-7b/TLR4通路与巨噬细胞极化和炎症消除有关。进一步研究结果显示，TLR4/NF-kappaB/STAT3/AKT调节通路在LPS pre-Exo调节炎症反应中巨噬细胞的极化上发挥着重要作用。

本研究的结论：LPS预处理的MSCs通过外泌体传递let-7b调节炎症反应中巨噬细胞的极化，这些外泌体具有对伤口修复提供免疫治疗的潜能。

Ti, D., et al. (2015). "LPS-preconditioned mesenchymal stromal cellsmodify macrophage polarization for resolution of chronic inflammationvia exosome-shuttled let-7b." J Transl Med 13(1): 308.

BACKGROUND: Within the last few years,it has become evident that LPS-preconditioned mesenchymal stromal cells (LPSpre-MSCs) show enhanced paracrine effects, including increased trophic supportand improved regenerative and repair properties. MSCs may release large amountsof exosomes for cell-to-cell communication and maintain a dynamic andhomeostatic microenvironment for tissue repair. The present study assesses thetherapeutic efficacy and mechanisms of LPS-preconditioned MSC-derived exosomes(LPS pre-Exo) for chronic inflammation and wound healing.

METHODS: We extracted exosomes from thesupernatant of LPS pre-MSCs using a gradient centrifugation method. In vitro,THP-1 cells were cultured with high glucose (HG, 30 mM) as an inflammatorymodel and treated with LPS pre-Exo for 48 h. The expression ofinflammation-related cytokines was detected by real-time RT-PCR, and thedistribution of macrophage subtype was measured by immunofluorescence. Next,the miRNA expression profiles of LPS pre-Exo were evaluated using miRNAmicroarray analysis. The molecular signaling pathway responsible for theregenerative potential was identified by western blotting. In vivo, weestablished a cutaneous wound model in streptozotocin-induced diabetic rats,and LPS pre-Exo were injected dispersively into the wound edge. The curativeeffects of LPS pre-Exo on inflammation and wound healing were observed andevaluated.

RESULTS: LPS pre-Exo have a betterability than untreated MSC-derived exosomes (un-Exo) to modulate the balance ofmacrophages due to their upregulation of the expression of anti-inflammatorycytokines and promotion of M2 macrophage activation. Microarray analysis of LPSpre-Exo identified the unique expression of let-7b compared with un-Exo, andthe let-7b/TLR4 pathway served as potential contributor to macrophagepolarization and inflammatory ablation. Further investigation of the mechanismsthat control let-7b expression demonstrated that a TLR4/NF-kappaB/STAT3/AKTregulatory signaling pathway plays a critical role in the regulation ofmacrophage plasticity. Knockdown of AKT in THP-1 cells similarly abolished theimmunomodulatory effect of LPS pre-Exo. In vivo, LPS pre-Exo greatly alleviatedinflammation and enhanced diabetic cutaneous wound healing.

CONCLUSION: LPS pre-Exo may haveimproved regulatory abilities for macrophage polarization and resolution ofchronic inflammation by shuttling let-7b, and these exosomes carry muchimmunotherapeutic potential for wound healing.