Microglial inflammation after chronic spinal cord injury is enhanced by reactive astrocytes via the fibronectin/β1 integrin pathway

Abstract

Background After spinal cord injury (SCI), glial scarring is mainly formed around the lesion and inhibits axon regeneration. Recently, we reported that anti-β1 integrin antibody (β1Ab) had a therapeutic effect on astrocytes by preventing the induction of glial scar formation. However, the cellular components within the glial scar are not only astrocytes but also microglia, and whether or not β1Ab treatment has any influence on microglia within the glial scar remains unclear. Methods To evaluate the effects of β1Ab treatment on microglia within the glial scar after SCI, we applied thoracic contusion SCI to C57BL/6N mice, administered β1Ab in the sub-acute phase, and analyzed the injured spinal cords with immunohistochemistry in the chronic phase. To examine the gene expression in microglia and glial scars, we selectively collected microglia with fluorescence-activated cell sorting and isolated the glial scars using laser-captured microdissection (LMD). To examine the interaction between microglia and astrocytes within the glial scar, we stimulated BV-2 microglia with conditioned medium of reactive astrocytes (RACM) in vitro, and the gene expression of TNFα (pro-inflammatory M1 marker) was analyzed via quantitative polymerase chain reaction. We also isolated both naïve astrocytes (NAs) and reactive astrocytes (RAs) with LMD and examined their expression of the ligands for β1 integrin receptors. Statistical analyses were performed using Wilcoxon’s rank-sum test. Results After performing β1Ab treatment, the microglia were scattered within the glial scar and the expression of TNFα in both the microglia and the glial scar were significantly suppressed after SCI. This in vivo alteration was attributed to fibronectin, a ligand of β1 integrin receptors. Furthermore, the microglial expression of TNFα was shown to be regulated by RACM as well as fibronectin in vitro. We also confirmed that fibronectin was secreted by RAs both in vitro and in vivo. These results highlighted the interaction mediated by fibronectin between RAs and microglia within the glial scar. Conclusion Microglial inflammation was enhanced by RAs via the fibronectin/β1 integrin pathway within the glial scar after SCI. Our results suggested that β1Ab administration had therapeutic potential for ameliorating both glial scar formation and persistent neuroinflammation in the chronic phase after SCI.

Fascin-1 is Highly Expressed Specifically in Microglia After Spinal Cord Injury and Regulates Microglial Migration

“…BV-2 is a commonly used microglial cell line that has been widely used in vitro experiments to explore inflammatory responses in many studies ( Riedl et al, 2009 ; Basso et al, 2017 ; Yoshizaki et al, 2021 ). The BV-2 microglial cell line was obtained from the American Type Culture Collection (CRL-3265, ATCC, Manassas, VA, United States) and cultured in Dulbecco’s modified Eagle’s medium (DMEM, HyClone, SH30021) supplemented with 10% fetal bovine serum (FBS, Gibco, 10,270,106), 100 U/ml penicillin and 100 g/ml streptomycin (Gibco, Grand Island, NY, United States).…”

Effect of ganglioside combined with Chip Jiaji electro-acupuncture on Nogo-NgR signal pathway in SCI rats

“…It can be inferred that the combined use of Jiaji EA and ganglioside ester may effectively reduce the inhibitory effect on nerve axon growth and further play a protective role on SCI by reducing the signal regulation of Nogo-A, NgR and LINGO-1 gene expression, especially the change of LINGO-1 expression. Considering that there is no effective clinical treatment currently ( Yates et al, 2021 , Yoshizaki et al, 2021 ), our study provided alternative therapeutic strategy for spinal cord injury.…”

“…SCI is a serious trauma to the central nervous system, causing serious disability ( Andrade et al, 2021 , Jogia et al, 2021 , Bilchak et al, 2021 ). There is no effective clinical treatment at present ( Yates et al, 2021 , Yoshizaki et al, 2021 ).…”

B Cells in Neuroinflammation: New Perspectives and Mechanistic Insights

“…Microglia, the CNS resident macrophages, can shift their functional states between a pro-inflammatory phenotype—characterized by release of inflammatory factors, antigen presentation, and expression of C1q and other markers—to a more neuroprotective phenotype—characterized by phagocytosis of debris and apoptotic cells and expression of S100B and other markers. Similarly, astrocytes display functional heterogeneity under both homeostatic and disease states that may be either neurotoxic or neuroprotective [ 47 , 48 , 49 , 50 ]. Together, microglia and astrocytes can adopt pro- or anti-inflammatory phenotypes to either prevent or exacerbate disease, and studies have shown that reactive microglia and astrocytes contribute to disease through upregulated transcriptional activity of pro-inflammatory genes [ 51 , 52 , 53 ].…”