The secretome of macrophages has a differential impact on spinal cord injury recovery according to the polarization protocol

José Lentilhas-Graça; Diogo J Santos; João Afonso; Andreia Monteiro; Andreia G Pinho; Vera M Mendes; Marta S Dias; Eduardo D Gomes; Rui Lima; Luís S Fernandes; Fernando Fernandes-Amorim; Inês M Pereira; Nídia de Sousa; Jorge R Cibrão; Aline M Fernandes; Sofia C Serra; Luís A Rocha; Jonas Campos; Tiffany S Pinho; Susana Monteiro; Bruno Manadas; António J Salgado; Ramiro D Almeida; Nuno A Silva

doi:10.3389/fimmu.2024.1354479

The secretome of macrophages has a differential impact on spinal cord injury recovery according to the polarization protocol

Front Immunol. 2024 Feb 20:15:1354479. doi: 10.3389/fimmu.2024.1354479. eCollection 2024.

Authors

José Lentilhas-Graça^{1

2

3}, Diogo J Santos^#^{1

2}, João Afonso^#^{1

2}, Andreia Monteiro^{1

2}, Andreia G Pinho^{1

2}, Vera M Mendes³, Marta S Dias^{3

4}, Eduardo D Gomes^{1

2}, Rui Lima^{1

2}, Luís S Fernandes^{1

2}, Fernando Fernandes-Amorim^{1

2}, Inês M Pereira^{1

2}, Nídia de Sousa^{1

2}, Jorge R Cibrão^{1

2}, Aline M Fernandes^{1

2}, Sofia C Serra^{1

2}, Luís A Rocha^{1

2}, Jonas Campos^{1

2}, Tiffany S Pinho^{1

2}, Susana Monteiro^{1

2}, Bruno Manadas³, António J Salgado^{1

2}, Ramiro D Almeida^{3

4}, Nuno A Silva^{1

2}

Affiliations

¹ Life and Health Sciences Research Institute (ICVS), School of Medicine, University of Minho, Braga, Portugal.
² ICVS/3B's Associate Lab, PT Government Associated Lab, Braga, Portugal.
³ CNC-Center for Neuroscience and Cell Biology, University of Coimbra, Coimbra, Portugal.
⁴ iBiMED- Institute of Biomedicine, Department of Medical Sciences, University of Aveiro, Aveiro, Portugal.

^# Contributed equally.

Abstract

Introduction: The inflammatory response after spinal cord injury (SCI) is an important contributor to secondary damage. Infiltrating macrophages can acquire a spectrum of activation states, however, the microenvironment at the SCI site favors macrophage polarization into a pro-inflammatory phenotype, which is one of the reasons why macrophage transplantation has failed.

Methods: In this study, we investigated the therapeutic potential of the macrophage secretome for SCI recovery. We investigated the effect of the secretome in vitro using peripheral and CNS-derived neurons and human neural stem cells. Moreover, we perform a pre-clinical trial using a SCI compression mice model and analyzed the recovery of motor, sensory and autonomic functions. Instead of transplanting the cells, we injected the paracrine factors and extracellular vesicles that they secrete, avoiding the loss of the phenotype of the transplanted cells due to local environmental cues.

Results: We demonstrated that different macrophage phenotypes have a distinct effect on neuronal growth and survival, namely, the alternative activation with IL-10 and TGF-β1 (M(IL-10+TGF-β1)) promotes significant axonal regeneration. We also observed that systemic injection of soluble factors and extracellular vesicles derived from M(IL-10+TGF-β1) macrophages promotes significant functional recovery after compressive SCI and leads to higher survival of spinal cord neurons. Additionally, the M(IL-10+TGF-β1) secretome supported the recovery of bladder function and decreased microglial activation, astrogliosis and fibrotic scar in the spinal cord. Proteomic analysis of the M(IL-10+TGF-β1)-derived secretome identified clusters of proteins involved in axon extension, dendritic spine maintenance, cell polarity establishment, and regulation of astrocytic activation.

Discussion: Overall, our results demonstrated that macrophages-derived soluble factors and extracellular vesicles might be a promising therapy for SCI with possible clinical applications.

Keywords: macrophages; neuroimmunology; neuroregeneration; secretome; spinal cord injury.

Copyright © 2024 Lentilhas-Graça, Santos, Afonso, Monteiro, Pinho, Mendes, Dias, Gomes, Lima, Fernandes, Fernandes-Amorim, Pereira, de Sousa, Cibrão, Fernandes, Serra, Rocha, Campos, Pinho, Monteiro, Manadas, Salgado, Almeida and Silva.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Humans
Interleukin-10*
Mice
Proteomics
Secretome
Spinal Cord Injuries* / therapy
Transforming Growth Factor beta1

Substances

Interleukin-10
Transforming Growth Factor beta1

Grants and funding

The author(s) declare financial support was received for the research, authorship, and/or publication of this article. This work has been funded by National funds, through the Foundation for Science and Technology (FCT) - project UIDB/50026/2020 (DOI 10.54499/UIDB/50026/2020), UIDP/50026/2020 (DOI 10.54499/UIDP/50026/2020), LA/P/0050/2020 (DOI 10.54499/LA/P/0050/2020) and EXPL/MED-PAT/0931/2021. Financial support was also provided by Prémios Santa Casa Neurociências–Prize Melo e Castro for Spinal Cord Injury Research (MC-18-2021) and by Wings For Life Spinal Cord Research Foundation (WFL-PT-14/23). Imaging acquisition was performed in the LiM facilities of iBiMED and at the ICVS Scientific Microscopy Platform, both are members of the national infrastructure PPBI - Portuguese Platform of Bioimaging (PPBI-POCI-01-0145-FEDER-022122).