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M2 Microglia-derived Exosomes Promote Spinal Cord Injury Recovery in Mice by Alleviating A1 Astrocyte Activation

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Abstract

M2 microglia transplantation has previously demonstrated beneficial effects on spinal cord injury (SCI) by regulating neuroinflammation and enhancing neuronal survival. Exosomes (EXOs), secreted by almost all cell types, embody partial functions and properties of their parent cells. However, the effect of M2 microglia-derived EXOs (M2-EXOs) on SCI recovery and the underlying molecular mechanisms remain unclear. In this study, we isolated M2-EXOs and intravenously introduced them into mice with SCI. Considering the reciprocal communication between microglia and astroglia in both healthy and injured central nervous systems (CNSs), we subsequently focused on the influence of M2-EXOs on astrocyte phenotype regulation. Our findings indicated that M2-EXOs promoted neuron survival and axon preservation, reduced the lesion area, inhibited A1 astrocyte activation, and improved motor function recovery in SCI mice. Moreover, they inhibited the nuclear translocation of p65 and the activation of the NF-κB signalling pathway in A1 astrocytes. Therefore, our research suggests that M2-EXOs mitigate the activation of neurotoxic A1 astrocytes by inhibiting the NF-κB signalling pathway, thereby improving spinal tissue preservation and motor function recovery following SCI. This positions M2-EXOs as a promising therapeutic strategy for SCI.

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Data Availability

The data of this study are available from the corresponding author upon reasonable request.

Abbreviations

GFAP :

Glial fibrillary acidic protein

IL-4 :

Interleukin 4

EXOs :

Exosomes

M2-EXO :

M2 microglia-derived exosomes

PDTC :

Pyrrolidine dithiocarbamate

C3 :

Component 3

SCI :

Spinal cord injury

PBS :

Phosphate-buffered saline

DMEM :

Dulbecco’s Modified Eagle medium

FBS :

Foetal bovine serum

PFA :

Paraformaldehyde

BMS :

Basso Mouse Scale scoring

DAPI :

4, 6-Diamidino-2-phenylindole

CNS :

Central nervous system

SEM :

Standard error of mean

References

  1. Ahuja CS, Wilson JR, Nori S, Kotter MRN, Druschel C, Curt A et al (2017) Traumatic spinal cord injury. Nat Rev Dis Primers 3:17018

    Article  PubMed  Google Scholar 

  2. Lu X, Xu G, Lin Z, Zou F, Liu S, Zhang Y et al (2023) Engineered exosomes enriched in netrin-1 modRNA promote axonal growth in spinal cord injury by attenuating inflammation and pyroptosis. Biomater Res 27:3

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Assinck P, Duncan GJ, Hilton BJ, Plemel JR, Tetzlaff W (2017) Cell transplantation therapy for spinal cord injury. Nat Neurosci 20:637–647

    Article  CAS  PubMed  Google Scholar 

  4. Kobashi S, Terashima T, Katagi M, Nakae Y, Okano J, Suzuki Y et al (2020) Transplantation of M2-deviated microglia promotes recovery of motor function after spinal cord injury in mice. Mol Ther 28:254–265

    Article  CAS  PubMed  Google Scholar 

  5. Gao ZS, Zhang CJ, Xia N, Tian H, Li DY, Lin JQ et al (2021) Berberine-loaded M2 macrophage-derived exosomes for spinal cord injury therapy. Acta Biomater 126:211–223

    Article  CAS  PubMed  Google Scholar 

  6. Zhang C, Li D, Hu H, Wang Z, An J, Gao Z et al (2021) Engineered extracellular vesicles derived from primary M2 macrophages with anti-inflammatory and neuroprotective properties for the treatment of spinal cord injury. J Nanobiotechnol 19:373

    Article  CAS  Google Scholar 

  7. Chen X, Chen C, Fan S, Wu S, Yang F, Fang Z et al (2018) Omega-3 polyunsaturated fatty acid attenuates the inflammatory response by modulating microglia polarization through SIRT1-mediated deacetylation of the HMGB1/NF-κB pathway following experimental traumatic brain injury. J Neuroinflammation 15:116

    Article  PubMed  PubMed Central  Google Scholar 

  8. Ajami M, Eghtesadi S, Razaz JM, Kalantari N, Habibey R, Nilforoushzadeh MA et al (2011) Expression of Bcl-2 and Bax after hippocampal ischemia in DHA + EPA treated rats. Neurol Sci 32:811–818

    Article  PubMed  Google Scholar 

  9. Ajami M, Davoodi SH, Habibey R, Namazi N, Soleimani M, Pazoki-Toroudi H (2013) Effect of DHA+EPA on oxidative stress and apoptosis induced by ischemia-reperfusion in rat kidneys. Fundam Clin Pharmacol 27:593–602

    Article  CAS  PubMed  Google Scholar 

  10. Kigerl KA, Gensel JC, Ankeny DP, Alexander JK, Donnelly DJ, Popovich PG (2009) Identification of two distinct macrophage subsets with divergent effects causing either neurotoxicity or regeneration in the injured mouse spinal cord. J Neurosci 29:13435–13444

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Kalluri R, LeBleu VS (2020) The biology, function, and biomedical applications of exosomes. Science 367:eaau6977

  12. Zhang J, Shi W, Qu D, Yu T, Qi C, Fu H (2022) Extracellular vesicle therapy for traumatic central nervous system disorders. Stem Cell Res Ther 13:442

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Zhang R, Mao W, Niu L, Bao W, Wang Y, Wang Y et al (2023) NSC-derived exosomes enhance therapeutic effects of NSC transplantation on cerebral ischemia in mice. Elife 12:e84493

  14. Sheng X, Zhao J, Li M, Xu Y, Zhou Y, Xu J et al (2021) Bone marrow mesenchymal stem cell-derived exosomes accelerate functional recovery after spinal cord injury by promoting the phagocytosis of macrophages to clean myelin debris. Front Cell Dev Biol 9:772205

    Article  PubMed  PubMed Central  Google Scholar 

  15. Han M, Yang H, Lu X, Li Y, Liu Z, Li F et al (2022) Three-dimensional-cultured MSC-derived exosome-hydrogel hybrid microneedle array patch for spinal cord repair. Nano Lett 22:6391–6401

    Article  ADS  CAS  PubMed  Google Scholar 

  16. Song Y, Li Z, He T, Qu M, Jiang L, Li W et al (2019) M2 microglia-derived exosomes protect the mouse brain from ischemia-reperfusion injury via exosomal miR-124. Theranostics 9:2910–2923

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Huang S, Ge X, Yu J, Han Z, Yin Z, Li Y et al (2018) Increased miR-124-3p in microglial exosomes following traumatic brain injury inhibits neuronal inflammation and contributes to neurite outgrowth via their transfer into neurons. Faseb J 32:512–528

    Article  CAS  PubMed  Google Scholar 

  18. Fu H, Zhao Y, Hu D, Wang S, Yu T, Zhang L (2020) Depletion of microglia exacerbates injury and impairs function recovery after spinal cord injury in mice. Cell Death Dis 11:528

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Sofroniew MV, Vinters HV (2010) Astrocytes: biology and pathology. Acta Neuropathol 119:7–35

    Article  PubMed  Google Scholar 

  20. Endo F, Kasai A, Soto JS, Yu X, Qu Z, Hashimoto H et al (2022) Molecular basis of astrocyte diversity and morphology across the CNS in health and disease. Science 378:9020

    Article  Google Scholar 

  21. Liddelow SA, Barres BA (2017) Reactive astrocytes: production, function, and therapeutic potential. Immunity 46:957–967

    Article  CAS  PubMed  Google Scholar 

  22. Escartin C, Galea E, Lakatos A, O’Callaghan JP, Petzold GC, Serrano-Pozo A et al (2021) Reactive astrocyte nomenclature, definitions, and future directions. Nat Neurosci 24:312–325

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Anderson MA, Burda JE, Ren Y, Ao Y, O’Shea TM, Kawaguchi R et al (2016) Astrocyte scar formation aids central nervous system axon regeneration. Nature 532:195–200

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  24. Liddelow SA, Guttenplan KA, Clarke LE, Bennett FC, Bohlen CJ, Schirmer L et al (2017) Neurotoxic reactive astrocytes are induced by activated microglia. Nature 541:481–487

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  25. D’Ambrosi N, Apolloni S (2020) Fibrotic scar in neurodegenerative diseases. Front Immunol 11:1394

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Jha MK, Jo M, Kim JH, Suk K (2019) Microglia-astrocyte crosstalk: an intimate molecular conversation. Neuroscientist 25:227–240

    Article  CAS  PubMed  Google Scholar 

  27. Budnik V, Ruiz-Cañada C, Wendler F (2016) Extracellular vesicles round off communication in the nervous system. Nat Rev Neurosci 17:160–172

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Liu K, Lu Y, Lee JK, Samara R, Willenberg R, Sears-Kraxberger I et al (2010) PTEN deletion enhances the regenerative ability of adult corticospinal neurons. Nat Neurosci 13:1075–1081

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Zukor K, Belin S, Wang C, Keelan N, Wang X, He Z (2013) Short hairpin RNA against PTEN enhances regenerative growth of corticospinal tract axons after spinal cord injury. J Neurosci 33:15350–15361

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Brommer B, He M, Zhang Z, Yang Z, Page JC, Su J et al (2021) Improving hindlimb locomotor function by non-invasive AAV-mediated manipulations of propriospinal neurons in mice with complete spinal cord injury. Nat Commun 12:781

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  31. Periyasamy P, Liao K, Kook YH, Niu F, Callen SE, Guo ML et al (2018) Cocaine-mediated downregulation of miR-124 activates microglia by targeting KLF4 and TLR4 signaling. Mol Neurobiol 55:3196–3210

    Article  CAS  PubMed  Google Scholar 

  32. Yao X, Liu S, Ding W, Yue P, Jiang Q, Zhao M et al (2017) TLR4 signal ablation attenuated neurological deficits by regulating microglial M1/M2 phenotype after traumatic brain injury in mice. J Neuroimmunol 310:38–45

    Article  CAS  PubMed  Google Scholar 

  33. Lai JJ, Chau ZL, Chen SY, Hill JJ, Korpany KV, Liang NW et al (2022) Exosome processing and characterization approaches for research and technology development. Adv Sci (Weinh) 9:e2103222

    Article  PubMed  Google Scholar 

  34. Kimiz-Gebologlu I, Oncel SS (2022) Exosomes: Large-scale production, isolation, drug loading efficiency, and biodistribution and uptake. J Control Release 347:533–543

    Article  CAS  PubMed  Google Scholar 

  35. Hara M, Kobayakawa K, Ohkawa Y, Kumamaru H, Yokota K, Saito T et al (2017) Interaction of reactive astrocytes with type I collagen induces astrocytic scar formation through the integrin-N-cadherin pathway after spinal cord injury. Nat Med 23:818–828

    Article  CAS  PubMed  Google Scholar 

  36. Zhang Y, Meng T, Chen J, Zhang Y, Kang J, Li X et al (2021) miR-21a-5p promotes inflammation following traumatic spinal cord injury through upregulation of neurotoxic reactive astrocyte (A1) Polarization by Inhibiting the CNTF/STAT3/Nkrf Pathway. Int J Biol Sci 17:2795–2810

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Sacristán C (2020) Microglia and Astrocyte Crosstalk in Immunity. Trends Immunol 41:747–748

    Article  PubMed  Google Scholar 

  38. Han RT, Kim RD, Molofsky AV, Liddelow SA (2021) Astrocyte-immune cell interactions in physiology and pathology. Immunity 54:211–224

    Article  CAS  PubMed  Google Scholar 

  39. Wheeler MA, Clark IC, Lee HG, Li Z, Linnerbauer M, Rone JM et al (2023) Droplet-based forward genetic screening of astrocyte-microglia cross-talk. Science 379:1023–1030

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  40. Dresselhaus EC, Meffert MK (2019) Cellular specificity of NF-κB function in the nervous system. Front Immunol 10:1043

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Lawrence T (2011) Macrophages and NF-κB in cancer. Curr Top Microbiol Immunol 349:171–184

    CAS  PubMed  Google Scholar 

  42. Lian H, Yang L, Cole A, Sun L, Chiang AC, Fowler SW et al (2015) NFκB-activated astroglial release of complement C3 compromises neuronal morphology and function associated with Alzheimer′s disease. Neuron 85:101–115

    Article  CAS  PubMed  Google Scholar 

  43. Lu T, Zhang Z, Zhang J, Pan X, Zhu X, Wang X et al (2022) CD73 in small extracellular vesicles derived from HNSCC defines tumour-associated immunosuppression mediated by macrophages in the microenvironment. J Extracell Vesicles 11:e12218

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Ren J, Zhu B, Gu G, Zhang W, Li J, Wang H et al (2023) Schwann cell-derived exosomes containing MFG-E8 modify macrophage/microglial polarization for attenuating inflammation via the SOCS3/STAT3 pathway after spinal cord injury. Cell Death Dis 14:70

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Xiong W, Li C, Kong G, Zeng Q, Wang S, Yin G et al (2022) Treg cell-derived exosomes miR-709 attenuates microglia pyroptosis and promotes motor function recovery after spinal cord injury. J Nanobiotechnol 20:529

    Article  CAS  Google Scholar 

  46. Li L, Zhang Y, Mu J, Chen J, Zhang C, Cao H et al (2020) Transplantation of human mesenchymal stem-cell-derived exosomes immobilized in an adhesive hydrogel for effective treatment of spinal cord injury. Nano Lett 20:4298–4305

    Article  ADS  CAS  PubMed  Google Scholar 

  47. Li Y, He X, Kawaguchi R, Zhang Y, Wang Q, Monavarfeshani A et al (2020) Microglia-organized scar-free spinal cord repair in neonatal mice. Nature 587:613–618

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  48. Szalay G, Martinecz B, Lénárt N, Környei Z, Orsolits B, Judák L et al (2016) Microglia protect against brain injury and their selective elimination dysregulates neuronal network activity after stroke. Nat Commun 7:11499

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  49. Yang J, Zhang X, Chen X, Wang L, Yang G (2017) Exosome mediated delivery of miR-124 promotes neurogenesis after ischemia. Mol Ther Nucleic Acids 7:278–287

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Stirling DP, Cummins K, Mishra M, Teo W, Yong VW, Stys P (2014) Toll-like receptor 2-mediated alternative activation of microglia is protective after spinal cord injury. Brain 137:707–723

    Article  PubMed  Google Scholar 

  51. Ma D, Zhao Y, Huang L, Xiao Z, Chen B, Shi Y et al (2020) A novel hydrogel-based treatment for complete transection spinal cord injury repair is driven by microglia/macrophages repopulation. Biomaterials 237:119830

    Article  CAS  PubMed  Google Scholar 

  52. Lu Y, Belin S, He Z (2014) Signaling regulations of neuronal regenerative ability. Curr Opin Neurobiol 27:135–142

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Zamanian JL, Xu L, Foo LC, Nouri N, Zhou L, Giffard RG et al (2012) Genomic analysis of reactive astrogliosis. J Neurosci 32:6391–6410

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  54. Brambilla R, Bracchi-Ricard V, Hu WH, Frydel B, Bramwell A, Karmally S et al (2005) Inhibition of astroglial nuclear factor kappaB reduces inflammation and improves functional recovery after spinal cord injury. J Exp Med 202:145–156

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Ding Y, Chen Q (2023) The NF-κB pathway: a focus on inflammatory responses in spinal cord injury. Mol Neurobiol 60:5292–5308

  56. Karin M, Greten FR (2005) NF-kappaB: linking inflammation and immunity to cancer development and progression. Nat Rev Immunol 5:749–759

    Article  CAS  PubMed  Google Scholar 

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Funding

This work was supported by grants from the National Natural Science Foundation of China (82102558), the Shandong Province Natural Science Foundation (ZR2020QH117), and the China Postdoctoral Science Foundation (2021M691690).

Author information

Author notes

  1. Jing Zhang and Die Hu contributed equally to this work.

Authors and Affiliations

  1. Department of Sports Medicine, The Affiliated Hospital of Qingdao University, Qingdao, 266003, China

    Jing Zhang, Di Qu, Weipeng Shi, Qi Jiang, Haifeng Li, Chao Qi & Haitao Fu

  2. Medical Department of, Qingdao University, 308 Ningxia Road, Qingdao, 266071, China

    Jing Zhang, Di Qu, Weipeng Shi, Lei Xie & Qi Jiang

  3. Eye Institute of Shandong First Medical University, Qingdao Eye Hospital of Shandong First Medical University, Qingdao, 266071, China

    Die Hu

  4. Department of Bone Surgery, Qingdao Central Hospital, Qingdao, 266000, China

    Liping Li

  5. Department of Orthopedic Surgery, Qingdao Hospital, University of Health and Rehabilitation Sciences (Qingdao Municipal Hospital), Qingdao, 266071, China

    Lei Xie & Tengbo Yu

  6. Institute of Sports Medicine and Health, Qingdao University, Qingdao, 266000, China

    Tengbo Yu

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  1. Jing Zhang
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Contributions

J.Z and D.H performed the experiments and drafted the manuscript. H.F and C.Q conceived the project and designed the experiments. L.L contributed to the behavioural tests. D.Q contributed to the mRNA analysis. W.S and L.X contributed to immunostaining. Q.J and H.L contributed to the establishment and intervention of animal models. T.Y helped to interpret the data. All authors have read and approved the final version of the manuscript.

Corresponding authors

Correspondence to Chao Qi or Haitao Fu.

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The experimental procedures described in this work were approved by Medical Ethics Committee of The Affiliated Hospital of Qingdao University.

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Zhang, J., Hu, D., Li, L. et al. M2 Microglia-derived Exosomes Promote Spinal Cord Injury Recovery in Mice by Alleviating A1 Astrocyte Activation. Mol Neurobiol (2024). https://doi.org/10.1007/s12035-024-04026-6

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