title: LPS-preconditioned mesenchymal stromal cells modify macrophage polarization for resolution of chronic inflammation via exosome-shuttled let-7b.
abstract: BACKGROUND
  Within the last few years, it has become evident that LPS-preconditioned mesenchymal stromal cells (LPS pre-MSCs) show enhanced paracrine effects, including increased trophic support and improved regenerative and repair properties. MSCs may release large amounts of exosomes for cell-to-cell communication and maintain a dynamic and homeostatic microenvironment for tissue repair. The present study assesses the therapeutic efficacy and mechanisms of LPS-preconditioned MSC-derived exosomes (LPS pre-Exo) for chronic inflammation and wound healing.
  
  
  METHODS
  We extracted exosomes from the supernatant of LPS pre-MSCs using a gradient centrifugation method. In vitro, THP-1 cells were cultured with high glucose (HG, 30 mM) as an inflammatory model and treated with LPS pre-Exo for 48 h. The expression of inflammation-related cytokines was detected by real-time RT-PCR, and the distribution of macrophage subtype was measured by immunofluorescence. Next, the miRNA expression profiles of LPS pre-Exo were evaluated using miRNA microarray analysis. The molecular signaling pathway responsible for the regenerative potential was identified by western blotting. In vivo, we established a cutaneous wound model in streptozotocin-induced diabetic rats, and LPS pre-Exo were injected dispersively into the wound edge. The curative effects of LPS pre-Exo on inflammation and wound healing were observed and evaluated.
  
  
  RESULTS
  LPS pre-Exo have a better ability than untreated MSC-derived exosomes (un-Exo) to modulate the balance of macrophages due to their upregulation of the expression of anti-inflammatory cytokines and promotion of M2 macrophage activation. Microarray analysis of LPS pre-Exo identified the unique expression of let-7b compared with un-Exo, and the let-7b/TLR4 pathway served as potential contributor to macrophage polarization and inflammatory ablation. Further investigation of the mechanisms that control let-7b expression demonstrated that a TLR4/NF-κB/STAT3/AKT regulatory signaling pathway plays a critical role in the regulation of macrophage plasticity. Knockdown of AKT in THP-1 cells similarly abolished the immunomodulatory effect of LPS pre-Exo. In vivo, LPS pre-Exo greatly alleviated inflammation and enhanced diabetic cutaneous wound healing.
  
  
  CONCLUSIONS
  LPS pre-Exo may have improved regulatory abilities for macrophage polarization and resolution of chronic inflammation by shuttling let-7b, and these exosomes carry much immunotherapeutic potential for wound healing.
authors.affiliation: Institute of Basic Medicine Science, College of Life Science, Chinese PLA General Hospital, No. 28 Fuxing Road, Beijing, 100853, China. tddee@163.com.
authors.forename: Dongdong
authors.initials: D
authors.lastname: Ti
authors.affiliation: Institute of Basic Medicine Science, College of Life Science, Chinese PLA General Hospital, No. 28 Fuxing Road, Beijing, 100853, China. haojieh@163.com.
authors.forename: Haojie
authors.initials: H
authors.lastname: Hao
authors.affiliation: Institute of Basic Medicine Science, College of Life Science, Chinese PLA General Hospital, No. 28 Fuxing Road, Beijing, 100853, China. tc6636@sina.com.
authors.forename: Chuan
authors.initials: C
authors.lastname: Tong
authors.affiliation: Institute of Basic Medicine Science, College of Life Science, Chinese PLA General Hospital, No. 28 Fuxing Road, Beijing, 100853, China. ljj0558@126.com.
authors.forename: Jiejie
authors.initials: J
authors.lastname: Liu
authors.affiliation: Institute of Basic Medicine Science, College of Life Science, Chinese PLA General Hospital, No. 28 Fuxing Road, Beijing, 100853, China. dl505504@qq.com.
authors.forename: Liang
authors.initials: L
authors.lastname: Dong
authors.affiliation: Institute of Basic Medicine Science, College of Life Science, Chinese PLA General Hospital, No. 28 Fuxing Road, Beijing, 100853, China. zhengjxi@163.com.
authors.forename: Jingxi
authors.initials: J
authors.lastname: Zheng
authors.affiliation: Central Laboratory, Hainan Branch of Chinese PLA General Hospital, Sanya, 572013, China. zhaoyl301@163.com.
authors.forename: Yali
authors.initials: Y
authors.lastname: Zhao
authors.affiliation: Institute of Basic Medicine Science, College of Life Science, Chinese PLA General Hospital, No. 28 Fuxing Road, Beijing, 100853, China. liu_hui_ling@sohu.com.
authors.forename: Huiling
authors.initials: H
authors.lastname: Liu
authors.affiliation: Institute of Basic Medicine Science, College of Life Science, Chinese PLA General Hospital, No. 28 Fuxing Road, Beijing, 100853, China. fuxiaobing@vip.sina.com.
authors.forename: Xiaobing
authors.initials: X
authors.lastname: Fu
authors.affiliation: Institute of Basic Medicine Science, College of Life Science, Chinese PLA General Hospital, No. 28 Fuxing Road, Beijing, 100853, China. hanwdrsw69@yahoo.com.
authors.forename: Weidong
authors.initials: W
authors.lastname: Han
mesh_terms: D002460:Cell Line; D055354:Exosomes / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D007249:Inflammation / Q000473:pathology / Q000628:therapy*; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008264:Macrophages / Q000276:immunology*; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000187:drug effects*; D035683:MicroRNAs / Q000378:metabolism*
chemical_list: D008070:Lipopolysaccharides; D035683:MicroRNAs; C487931:mirnlet7 microRNA, human