Office of Clinical Trial of Drug, Guangdong Provincial Key Laboratory of Bone and Joint Degeneration Diseases, The Third Affiliated Hospital of Southern Medical University, China.
Guangdong Provincial Key Laboratory of Bone and Joint Degeneration Diseases, The Third Affiliated Hospital of Southern Medical University, China.
Life Sci. 2020 Apr 1;246:117401. doi: 10.1016/j.lfs.2020.117401. Epub 2020 Feb 6.
The management of acute liver failure (ALF) is a major challenge worldwide. The current study aimed to determine the therapeutic potential of TNF-α pretreatment of umbilical cord mesenchymal stem cell-derived exosomes (T-Exo) in ALF.
Here, we enriched T-Exo and untreated exosomes (Exo), them were measured by nanoparticle tracking analysis (NTA) for particle size detection and identified surface marker by Western blot and flow cytometry. Then the cell proliferation was detected by CCK-8 and the effect of T-Exo on the expression levels of pro-inflammatory cytokines was tested by ELISA. ALF mouse models were induced by LPS and D-GalN. H&E staining, immunohistochemistry, and Western blot were used to detect the effect of T-Exo on the levels of NLRP3 and other inflammation-related pathway proteins. qPCR was used to detect the expression level of microRNA-299-3p in T-Exo and its transfer to macrophages. Laser confocal microscopy was used to detect colocalization of exosomes,Golgi and NLRP3 in macrophages.
Our study shows that T-Exo can reduce serum ALT, AST and proinflammatory cytokines level and inhibit activation of NLRP3 inflammation-associated pathway proteins. T-Exo treatment reduces pathological liver damage caused by ALF. Anti-inflammatory-related miRNA-299-3p is up-regulated in TNF-α-stimulated MSCs and selectively packaged into exosomes for role in exosomal treatment. And conducted preliminary exploration and hypothesis on the specific mechanism of this effect.
These in vitro and in vivo studies indicate that T-Exo attenuates inflammatory damage caused by ALF and promotes liver tissue repair by inhibiting the activation of the NLRP3 pathway.
急性肝衰竭（ALF）的治疗是全球性的重大挑战。本研究旨在确定 TNF-α预处理脐带间充质干细胞衍生的细胞外囊泡（T-Exo）在 ALF 中的治疗潜力。
在这里，我们富集了 T-Exo 和未经处理的细胞外囊泡（Exo），并用纳米颗粒跟踪分析（NTA）测量粒径，并通过 Western blot 和流式细胞术鉴定表面标志物。然后通过 CCK-8 检测细胞增殖，通过 ELISA 检测 T-Exo 对促炎细胞因子表达水平的影响。通过 LPS 和 D-GalN 诱导 ALF 小鼠模型。使用 H&E 染色、免疫组织化学和 Western blot 检测 T-Exo 对 NLRP3 及其他炎症相关途径蛋白水平的影响。qPCR 用于检测 T-Exo 中 microRNA-299-3p 的表达水平及其向巨噬细胞的转移。激光共聚焦显微镜用于检测巨噬细胞中外泌体、高尔基体和 NLRP3 的共定位。
我们的研究表明，T-Exo 可降低血清 ALT、AST 和促炎细胞因子水平，并抑制 NLRP3 炎症相关途径蛋白的激活。T-Exo 治疗可减轻 ALF 引起的病理性肝损伤。TNF-α 刺激的 MSC 中抗炎相关的 microRNA-299-3p 上调，并选择性包装到细胞外囊泡中，在细胞外囊泡治疗中发挥作用。并对这种作用的具体机制进行了初步探索和假设。
这些体内外研究表明，T-Exo 通过抑制 NLRP3 途径的激活，减轻 ALF 引起的炎症损伤，促进肝组织修复。
