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Apoptosis associated speck like protein containing a card (ASC), the key adaptor protein of the assembly and activation of canonical inflammasomes, has been found to play a significant role in neuroinflammation after spinal cord injury (SCI). The previous studies indicated that widely block or knockout ASC can ameliorate SCI. However, ASC is ubiquitously expressed in infiltrated macrophages and local microglia, so further exploration is needed on which type of cell playing the key role. In this study, using the LysM cre ;Asc flox/flox mice with macrophage-specifc ASC conditional knockout (CKO) and contusive SCI model, we focus on evaluating the specific role of ASC in lysozyme 2 (LysM) + myeloid cells (mainly infiltrated macrophages) in this pathology. The results revealed that macrophage-specifc Asc CKO exhibited the follow effects: (1) A significant reduction in the numbers of infiltrated macrophages in the all phases of SCI, and activated microglia in the acute and subacute phases. (2) A significant reduction in ASC, caspase-1, interleukin (IL)-1β, and IL-18 compared to control mice. (3) In the acute and subacute phases of SCI, M1 subset differentiation was inhibited, and M2 differentiation was increased. (4) Histology and hindlimb motor recoveries were improved. In conclusion, this study elucidates that macrophage-specific ASC CKO can improve nerve function recovery after SCI by regulating M1/M2 polarization through inhibiting ASC-dependent inflammasome signaling axis. This indicates that ASC in peripheral infiltrated macrophages may play an important role in SCI pathology, at least in mice, could be a potential target for treatment., Competing Interests: Declaration of competing interest The authors declare that there are no conflicts of interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Inflammation is one of the key injury factors for spinal cord injury (SCI). Exosomes (Exos) derived from M2 macrophages have been shown to inhibit inflammation and be beneficial in SCI animal models. However, lacking targetability restricts their application prospects. Considering that chemokine receptors increase dramatically after SCI, viral macrophage inflammatory protein II (vMIP-II) is a broad-spectrum chemokine receptor binding peptide, and lysosomal associated membrane protein 2b (Lamp2b) is the key membrane component of Exos, we speculated that vMIP-II-Lamp2b gene-modified M2 macrophage-derived Exos (vMIP-II-Lamp2b-M2-Exo) not only have anti-inflammatory properties, but also can target the injured area by vMIP-II. In this study, using a murine contusive SCI model, we revealed that vMIP-II-Lamp2b-M2-Exo could target the chemokine receptors which highly expressed in the injured spinal cords, inhibit some key chemokine receptor signaling pathways (such as MAPK and Akt), further inhibit proinflammatory factors (such as IL-1β, IL-6, IL-17, IL-18, TNF-α, and iNOS), and promote anti-inflammatory factors (such as IL-4 and Arg1) productions, and the transformation of microglia/macrophages from M1 into M2. Moreover, the improved histological and functional recoveries were also found. Collectively, our results suggest that vMIP-II-Lamp2b-M2-Exo may provide neuroprotection by targeting the injured spinal cord, inhibiting some chemokine signals, reducing proinflammatory factor production and modulating microglia/macrophage polarization., Competing Interests: Declaration of competing interest The authors declare that there are no conflicts of interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Objective To explore the effect of shionone(SHI)on motor function in the mouse model of spinal cord injury(SCI)and probe into the underlying molecular mechanism.Methods C57BL/6 mice were treated to induce the SCI model and then assigned into a model group(SCI group),a SCI+SHI group,and a sham surgery(control)group.The Basso mouse scale(BMS)score was determined to evaluate the recovery of motor function in SCI mice.Hematoxylin-eosin(HE)staining,Nissl staining,and immunofluorescence staining were employed to examine the fibrosis,morphological changes of neurons,and neuron apoptosis in the spinal cord tissue of SCI mice,respectively.The mouse hippocampal neuronal cell line HT22 was cultured in vitro and then classified into tumor necrosis factor α(TNF-α)induction and SHI groups.Western blotting was employed to determine the expression of apoptosis-associated proteins.Network pharmacology,gene ontology annotation,and Kyoto Encyclopedia of Genes and Genomes pathway enrichment were employed to predict the possible molecular targets and signaling pathways of SHI in promoting functional recovery from SCI.Furthermore,the prediction results were verified by in vitro and in vivo experiments.Results Compared with the SCI group,the SCI+SHI group showed increased BMS score on days 21,28,35,and 42( P =0.003, P =0.004, P =0.023,and P =0.007,respectively),reduced area of spinal cord fibrosis( P =0.021),increased neurons survived( P =0.001),and down-regulated expression of cleaved cysteine aspastic acid-specific protease 3(cleaved Caspase-3)( P =0.017).Compared with the TNF-α group,the SHI group presented down-regulated expression levels of cleaved Caspase-3 and Bax( P =0.010, P =0.001)and up-regulated expression level of Bcl-2( P =0.001).The results of bioinformatics analysis showed that SHI might improve the motor function of SCI mice via the phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt)signaling pathway.The results of in vivo and in vitro experiments showed that SHI inhibited the phosphorylation of PI3K and Akt in SCI mice or HT22 cells exposed to TNF-α(all P <0.05).The number of apoptotic HT22 cells after treatment with insulin-like growth factor 1 was higher than that in the SHI group( P =0.003).Conclusion SHI may inhibit neuron apoptosis via the PI3K/Akt signaling pathway,thereby promoting the recovery of motor function in SCI mice.

Background: Following spinal cord injury (SCI), a large number of peripheral monocytes infiltrate into the lesion area and differentiate into macrophages (Mø). These monocyte-derived Mø are very difficult to distinguish from the local activated microglia (MG). Therefore, the term Mø/MG are often used to define the infiltrated Mø and/or activated MG. It has been recognized that pro-inflammatory M1-type Mø/MG play "bad" roles in the SCI pathology. Our recent research showed that local M1 cells are mainly CD45 -/low CD68 + CD11b + in the subacute stage of SCI. Thus, we speculated that the M1 cells in injured spinal cords mainly derived from MG rather than infiltrating Mø. So far, their dynamics following SCI are not yet entirely clear., Methods: Female C57BL/6 mice were used to establish SCI model, using an Infinite Horizon impactor with a 1.3 mm diameter rod and a 50 Kdynes force. Sham-operated (sham) mice only underwent laminectomy without contusion. Flow cytometry and immunohistofluorescence were combined to analyze the dynamic changes of polarized Mø and MG in the acute (1 day), subacute (3, 7 and 14 days) and chronic (21 and 28 days) phases of SCI., Results: The total Mø/MG gradually increased and peaked at 7 days post-injury (dpi), and maintained at high levels 14, 21 and 28 dpi. Most of the Mø/MG were activated, and the Mø increased significantly at 1 and 3 dpi. However, with the pathological process, activated MG increased nearly to 90% at 7, 14, 21 and 28 dpi. Both M1 and M2 Mø were increased significantly at 1 and 3 dpi. However, they decreased to very low levels from 7 to 28 dpi. On the contrary, the M2-type MG decreased significantly following SCI and maintained at a low level during the pathological process., Competing Interests: The authors declare that they have no competing interests., (© 2023 Li et al.)

Background: Inflammatory reactions induced by spinal cord injury (SCI) are essential for recovery after SCI. Atractylenolide III (ATL-III) is a natural monomeric herbal bioactive compound that is mainly derived in Atractylodes macrocephala Koidz and has anti-inflammatory and neuroprotective effects., Objective: Here, we speculated that ATL-III may ameliorate SCI by modulating microglial/macrophage polarization. In the present research, we focused on investigating the role of ATL-III on SCI in rats and explored the potential mechanism., Methods: The protective and anti-inflammatory effects of ATL-III on neuronal cells were examined in a rat SCI model and lipopolysaccharide (LPS)-stimulated BV2 microglial line. The spinal cord lesion area, myelin integrity, and surviving neurons were assessed by specific staining. Locomotor function was evaluated by the Basso, Beattie, and Bresnahan (BBB) scale, grid walk test, and footprint test. The activation and polarization of microglia/macrophages were assessed by immunohistofluorescence and flow cytometry. The expression of corresponding inflammatory factors from M1/M2 and the activation of relevant signaling pathways were assessed by Western blotting., Results: ATL-III effectively improved histological and functional recovery in SCI rats. Furthermore, ATL-III promoted the transformation of M1 into M2 and attenuated the activation of microglia/macrophages, further suppressing the expression of corresponding inflammatory mediators. This effect may be partly mediated by inhibition of neuroinflammation through the NF-κB, JNK MAPK, p38 MAPK, and Akt pathways., Conclusion: This study reveals a novel effect of ATL-III in the regulation of microglial/macrophage polarization and provides initial evidence that ATL-III has potential therapeutic benefits in SCI rats., (© 2022 The Authors. CNS Neuroscience & Therapeutics Published by John Wiley & Sons Ltd.)

We have previously reported that morroniside promoted motor activity after spinal cord injury (SCI) in rats. However, the mechanism by which morroniside induces recovery of injured spinal cord (SC) remains unknown. In the current study, RNA sequencing (RNA-seq) was employed to evaluate changes of gene expressions at the transcriptional level of the injured spinal cords in morroniside-administrated rats. Principal component analysis, analysis of enriched Gene Ontology (GO), enrichment analyses Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, and other bioinformatics analyses were executed to distinguish differentially expressed genes (DEGs). The results of RNA-seq confirmed the anti-inflammatory and anti-apoptotic effects of morroniside on injured SC tissues, and provided the basis for additional research of the mechanisms involving the protective effects of morroniside on SCI., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Spinal cord injury (SCI) is a disabling condition that often leads to permanent neurological deficits without an effective treatment. Reactive oxygen species (ROS) produced during oxidative stress play a vital role in the pathogenesis following SCI. The antioxidant morroniside is the main active component of the Chinese medicine Cornus officinalis. In recent years, it has been reported that morroniside has therapeutic effects on damage to multiple organs mediated by oxidative damage, but the effect of morroniside on SCI has not been reported. The purpose of this study was therefore to assess the therapeutic effect of morroniside on SCI, and to identify its underlying mechanism by direct intragastric administration immediately after SCI. Our study showed that morroniside treatment improved the functional recovery of rats following SCI. This behavioral improvement was associated with the higher survival in neurons and oligodendrocytes following SCI, which increased the capacity of injured spinal cord (SC) to form myelin and repair tissue, eventually contributing to improved neurological outcome. Furthermore, our study found that oxygen free radicals increased and antioxidant enzyme activity decreased in the injured SC. Interestingly, morroniside treatment decreased oxygen free radical levels and increased antioxidant enzyme activities. Together, our results suggested that morroniside may be an effective treatment for improving outcomes following SCI, and that its antioxidant activity may be one of the mechanisms by which morroniside exerts neuroprotective effects on SCI., (Copyright © 2021. Published by Elsevier Ltd.)

MicroRNAs (miRNAs) are involved in a series of pathology of spinal cord injury (SCI). Although, locally expressed miRNAs have advantages in studying the pathological mechanism, they cannot be used as biomarkers. The "free circulation" miRNAs can be used as biomarkers, but they have low concentration and poor stability in body fluids. Exosomal miRNAs in body fluids have many advantages comparing with free miRNAs. Therefore, we hypothesized that the specific miRNAs in the central nervous system might be transported to the peripheral circulation and concentrated in exosomes after injury. Using next-generation sequencing, miRNA profiles in serum exosomes of sham and subactue SCI rats were analyzed. The results showed that SCI can lead to changes of serum exosomal miRNAs. These changed miRNAs and their associated signaling pathways may explain the pathological mechanism of suacute SCI. More importantly, we found some valuable serum exosomal miRNAs for diagnosis and prognosis of SCI., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Background: After spinal cord injury (SCI), destructive immune cell subsets are dominant in the local microenvironment, which are the important mechanism of injury. Studies have shown that inflammasomes play an important role in the inflammation following SCI, and apoptosis-associated speck-like protein containing a card (ASC) is the adaptor protein shared by inflammasomes. Therefore, we speculated that inhibiting ASC may improve the local microenvironment of injured spinal cord. Here, CRID3, a blocker of ASC oligomerization, was used to study its effect on the local microenvironment and the possible role in neuroprotection following SCI., Methods: Murine SCI model was created using an Infinite Horizon impactor at T9 vertebral level with a force of 50 kdynes and CRID3 (50 mg/kg) was intraperitoneally injected following injury. ASC and its downstream molecules in inflammasome signaling pathway were measured by western blot. The immune cell subsets were detected by immunohistofluorescence (IHF) and flow cytometry (FCM). The spinal cord fibrosis area, neuron survival, myelin preservation, and functional recovery were assessed., Results: Following SCI, CRID3 administration inhibited inflammasome-related ASC and caspase-1, IL-1β, and IL-18 activation, which consequently suppressed M1 microglia, Th1 and Th1Th17 differentiation, and increased M2 microglia and Th2 differentiation. Accordingly, the improved histology and behavior have also been found., Conclusions: CRID3 may ameliorate murine SCI by inhibiting inflammasome activation, reducing proinflammatory factor production, restoring immune cell subset balance, and improving local immune microenvironment, and early administration may be a promising therapeutic strategy for SCI.

Previous studies have shown that caspase-1 plays an important role in the acute inflammatory response of spinal cord injury (SCI). VX‑765, a novel and irreversible caspase‑1 inhibitor, has been reported to effectively intervene in inflammation. However, the effect of VX‑765 on genome‑wide transcription in acutely injured spinal cords remains unknown. Therefore, in the present study, RNA‑sequencing (RNA‑Seq) was used to analyze the effect of VX‑765 on the local expression of gene transcription 8 h following injury. The differentially expressed genes (DEGs) underwent enrichment analysis of functions and pathways by Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses, respectively. Parallel analysis of western blot confirmed that VX‑765 can effectively inhibit the expression and activation of caspase‑1. RNA‑Seq showed that VX‑765 treatment resulted in 1,137 upregulated and 1,762 downregulated DEGs. These downregulated DEGs and their associated signaling pathways, such as focal adhesion, cytokine‑cytokine receptor interaction, leukocyte transendothelial migration, extracellular matrix‑receptor interaction, phosphatidylinositol 3‑kinase‑protein kinase B, Rap1 and hypoxia inducible factor‑1 signaling pathway, are mainly associated with inflammatory response, local hypoxia, macrophage differentiation, adhesion migration and apoptosis of local cells. This suggests that the application of VX‑765 in the acute phase can improve the local microenvironment of SCI by inhibiting caspase‑1. However, whether VX‑765 can be used as a therapeutic drug for SCI requires further exploration. The sequence data have been deposited into the Sequence Read Archive (https://www.ncbi.nlm.nih.gov/sra/PRJNA548970).

Ceruloplasmin (Cp), an enzyme containing six copper atoms, has important roles in iron homeostasis and antioxidant defense. After spinal cord injury (SCI), the cellular components in the local microenvironment are very complex and include functional changes of resident cells and the infiltration of leukocytes. It has been confirmed that Cp is elevated primarily in astrocytes and to a lesser extent in macrophages following SCI in mice. However, its expression in other cell types is still not very clear. In this manuscript, we provide a sensible extension of these findings by examining this system within a female Sprague-Dawley rat model and expanding the scope of inquiry to include additional cell types. Quantitative reverse transcription polymerase chain reaction and Western blot analysis revealed that the Cp mRNA and protein in SCI tissue homogenates were quite consistent with prior publications. However, we observed that Cp was expressed not only in GFAP + astrocytes (consistent with prior reports) but also in CD11b + microglia, CNPase + oligodendrocytes, NeuN + neurons, CD45 + leukocytes, and CD68 + activated microglia/macrophages. Quantitative analysis proved that infiltrated leukocytes, activated microglia/macrophages, and astrocytes should be the major sources of increased Cp., (© 2018 Wiley Periodicals, Inc.)

Interferon-induced transmembrane protein 1 (IFITM1) is a member of the IFITM family that is associated with some acute-phase cytokine-stimulated response. Recently, we demonstrated that IFITM1 was significantly upregulated in the injured spinal cords at the mRNA level. However, its expression and cellular localization at the protein level is still unclear. Here, a rat model of spinal cord injury (SCI) was performed to investigate the spatio-temporal expression of IFITM1 after SCI. IFITM1 mRNA and protein were assessed by quantitative reverse transcription-PCR and western blot, respectively. IHC was used to identify its cellular localization. We revealed that IFITM1 could be found in sham-opened spinal cords and gradually increased after SCI. It reached peak at 7 and 14 days postinjury (dpi) and still maintained at a relatively higher level at 28 dpi. IHC showed that IFITM1 expressed in GFAP + and APC + cells in sham-opened spinal cords. After SCI, in addition to the above-mentioned cells, it could also be found in CD45 + and CD68 + cells, and its expression in CD45 + , CD68 + , and GFAP + cells was increased significantly. These results demonstrate that IFITM1 is mainly expressed in astrocytes and oligodendroglia in normal spinal cords, and could rapidly increase in infiltrated leukocytes, activated microglia, and astrocytes after SCI.

Hexokinase-3 (HK3) is a member of hexokinase family, which can catalyze the first step of glucose metabolism. It can increase ATP levels, reduce the production of reactive oxygen species, increase mitochondrial biogenesis, protect mitochondrial membrane potential and play an antioxidant role. However, the change of its expression in spinal cord after injury is still unknown. In this study, we investigated the spatio-temporal expression of HK3 in the spinal cords by using a spinal cord injury (SCI) model in adult female Sprague-Dawley rats. Quantitative reverse transcription-PCR and western blot analysis revealed that HK3 could be detected in sham-opened spinal cords. After SCI, it gradually increased, reached a peak at 7 days post-injury (dpi), and then gradually decreased with the prolonging of injury time, but still maintained at a higher level for up to 28 dpi (the longest time evaluated in this study). Immunofluorescence staining showed that HK3 was found in GFAP + , β-tubulin III + and IBA-1 + cells in sham-opened spinal cords. After SCI, in addition to the above-mentioned cells, it could also be found in CD45 + and CD68 + cells. These results demonstrate that HK3 is mainly expressed in astrocytes, neurons and microglia in normal spinal cords, and could rapidly increase in infiltrated leukocytes, activated microglia/macrophages and astrocytes after SCI. These data suggest that HK3 may be involved in the pathologic process of SCI by promoting glucose metabolism., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

This study aims to explore the temporal changes of cytotoxic CD8 + CD28 + and regulatory CD8 + CD28 - T-cell subsets in the lesion microenvironment after spinal cord injury (SCI) in rats, by combination of immunohistochemistry (IHC) and flow cytometry (FCM). In the sham-opened spinal cord, few CD8 + T cells were found. After SCI, the CD8 + T cells were detected at one day post-injury (dpi), then markedly increased and were significantly higher at 3, 7, and 14 dpi compared with one dpi (p < 0.01), the highest being seven dpi. In CD8 + T cells, more than 90% were CD28 + , and there were only small part of CD28 - ( < 10%). After 14 days, the infiltrated CD8 + T cells were significantly decreased, and few could be found in good condition at 21 and 28 dpi. Annexin V and propidium iodide (PI) staining showed that the percentages of apoptotic/necrotic CD8 + cells at 14 dpi and 21 dpi were significantly higher than those of the other early time-points (p < 0.01). These results indicate that CD8 + T cells could rapidly infiltrate into the injured spinal cords and survive two weeks, however, cytotoxic CD8 + T cells were dominant. Therefore, two weeks after injury might be the "time window" for treating SCI by prolonging survival times and increasing the fraction of CD8 + regulatory T-cells. © 2016 Wiley Periodicals, Inc., (© 2016 Wiley Periodicals, Inc.)

Background: Spinal cord injury (SCI) results in fatal damage and currently has no effective treatment. The pathological mechanisms of SCI remain unclear. In this study, genome-wide transcriptional profiling of spinal cord samples from injured rats at different time points after SCI was performed by RNA-Sequencing (RNA-Seq). The transcriptomes were systematically characterized to identify the critical genes and pathways that are involved in SCI pathology., Results: RNA-Seq results were obtained from total RNA harvested from the spinal cords of sham control rats and rats in the acute, subacute, and chronic phases of SCI (1 day, 6 days and 28 days after injury, respectively; n = 3 in every group). Compared with the sham-control group, the number of differentially expressed genes was 1797 in the acute phase (1223 upregulated and 574 downregulated), 6590 in the subacute phase (3460 upregulated and 3130 downregulated), and 3499 in the chronic phase (1866 upregulated and 1633 downregulated), with an adjusted P-value <0.05 by DESeq. Gene ontology (GO) enrichment analysis showed that differentially expressed genes were most enriched in immune response, MHC protein complex, antigen processing and presentation, translation-related genes, structural constituent of ribosome, ion gated channel activity, small GTPase mediated signal transduction and cytokine and/or chemokine activity. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed that the most enriched pathways included ribosome, antigen processing and presentation, retrograde endocannabinoid signaling, axon guidance, dopaminergic synapses, glutamatergic synapses, GABAergic synapses, TNF, HIF-1, Toll-like receptor, NF-kappa B, NOD-like receptor, cAMP, calcium, oxytocin, Rap1, B cell receptor and chemokine signaling pathway., Conclusions: This study has not only characterized changes in global gene expression through various stages of SCI progression in rats, but has also systematically identified the critical genes and signaling pathways in SCI pathology. These results will expand our understanding of the complex molecular mechanisms involved in SCI and provide a foundation for future studies of spinal cord tissue damage and repair. The sequence data from this study have been deposited into Sequence Read Archive ( http://www.ncbi.nlm.nih.gov/sra ; accession number PRJNA318311).

Oligodendrocyte (OL) replacement is a promising treatment strategy for spinal cord injury (SCI). However, the poor survival of transplanted OLs or their precursors and inhibition of axonal regeneration are two major challenges with this approach. Our previous study showed that Schwann cells (SCs) promoted survival, proliferation, and migration of transplanted OL progenitor cells (OPCs) and neurological recovery. Remyelination is an important basis for functional recovery following spinal cord injury. It has been reported that myelin gene regulatory factor (MRF), a transcriptional regulator which specifically is expressed in postmitotic OLs within the CNS, is essential for OL maturation and CNS myelination. In the present study, we investigated whether co-transplantation of MRF-overexpressing OPCs (MRF-OPCs) and SCs could improve functional recovery in a rat model of contusional SCI. MRF overexpression had no effect on OPC survival or migration, but stimulated the differentiation of OPCs both in vitro and in vivo. Co-transplantation of MRF-OPCs and SCs increased myelination and tissue repair after SCI, leading to the recovery of neurological function. These results indicate that co-transplantation of MRF-OPCs and SCs may be an effective treatment strategy for SCI., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Myelin basic protein (MBP) activated T cells (MBP-T) play an important role in the damage and repair process of the central nervous system (CNS). However, whether these cells play a beneficial or detrimental role is still a matter of debate. Although some studies showed that MBP-T cells are mainly helper T (Th) cells, their subtypes are still not very clear. One possible explanation for MBP-T immunization leading to conflicting results may be the different subtypes of T cells are responsible for distinct effects. In this study, the Th1 and Th2 type MBP-T cells (MBP-Th1 and -Th2) were polarized in vitro, and their effects on the local immune microenvironment and tissue repair of spinal cord injury (SCI) after adoptive immunization were investigated. In MBP-Th1 cell transferred rats, the high levels of pro-inflammatory cells (Th1 cells and M1 macrophages) and cytokines (IFN-γ, TNF-α, -β, IL-1β) were detected in the injured spinal cord; however, the anti-inflammatory cells (Th2 cells, regulatory T cells, and M2 macrophages) and cytokines (IL-4, -10, and -13) were found in MBP-Th2 cell transferred animals. MBP-Th2 cell transfer resulted in decreased lesion volume, increased myelination of axons, and preservation of neurons. This was accompanied by significant locomotor improvement. These results indicate that MBP-Th2 adoptive transfer has beneficial effects on the injured spinal cord, in which the increased number of Th2 cells may alter the local microenvironment from one primarily populated by Th1 and M1 cells to another dominated by Th2, Treg, and M2 cells and is conducive for SCI repair., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Local activated macrophages derived from infiltrating monocytes play an important role in the damage and repair process of spinal cord injury (SCI). The present study investigates the dynamic change of classically activated proinflammatory (M1) and alternatively activated anti-inflammatory (M2) cells in a rat model with contusive SCI by flow cytometry (FCM) and immunohistochemistry. The macrophage subsets were immunophenotyped by using antibodies against cluster of differentiation (CD)-68, C-C chemokine receptor type 7 (CCR7), CD163, and arginase 1 (Arg1). The CD68(+) CD163(-) and CD68(+) CCR7(+) cells were determined to be M1 subsets, whereas the CD68(+) CD163(+) and CD68(+) Arg1(+) cell subpopulations represented M2 cells. The subsets of macrophages in the injured spinal cord at 1, 3, 5, 7, 14, and 28 days postinjury (dpi) were examined. In the sham-opened spinal cord, few M1 or M2 cells were found. After SCI, the phenotypes of both M1 and M2 cells were rapidly induced. However, M1 cells were detected and maintained at a high level for up to 28 dpi (the longest time evaluated in this study). In contrast, M2 cells were transiently detected at high levels before 7 dpi and returned to preinjury levels at 14 dpi. These results indicate that M1 cell response is rapidly induced and sustained, whereas M2 induction is transient after SCI in rat. Increasing the fraction of M2 cells and prolonging their residence time in the injured local microenvironment is a promising strategy for the repair of SCI., (© 2015 Wiley Periodicals, Inc.)

Classically activated pro-inflammatory (M1) and alternatively activated anti-inflammatory (M2) macrophages populate the local microenvironment after spinal cord injury (SCI). The former type is neurotoxic while the latter has positive effects on neuroregeneration and is less toxic. In addition, while the M1 macrophage response is rapidly induced and sustained, M2 induction is transient. A promising strategy for the repair of SCI is to increase the fraction of M2 cells and prolong their residence time. This study investigated the effect of M2 macrophages induced from bone marrow-derived macrophages on the local microenvironment and their possible role in neuroprotection after SCI. M2 macrophages produced anti-inflammatory cytokines such as interleukin (IL)-10 and transforming growth factor β and infiltrated into the injured spinal cord, stimulated M2 and helper T (Th)2 cells, and produced high levels of IL-10 and -13 at the site of injury. M2 cell transfer decreased spinal cord lesion volume and resulted in increased myelination of axons and preservation of neurons. This was accompanied by significant locomotor improvement as revealed by Basso, Beattie and Bresnahan locomotor rating scale, grid walk and footprint analyses. These results indicate that M2 adoptive transfer has beneficial effects for the injured spinal cord, in which the increased number of M2 macrophages causes a shift in the immunological response from Th1- to Th2-dominated through the production of anti-inflammatory cytokines, which in turn induces the polarization of local microglia and/or macrophages to the M2 subtype, and creates a local microenvironment that is conducive to the rescue of residual myelin and neurons and preservation of neuronal function., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Objective: The objective of this study was to investigate whether cytosolic phospholipase A2 (cPLA2 ), an important isoform of PLA2 that mediates the release of arachidonic acid, plays a role in the pathogenesis of spinal cord injury (SCI)., Methods: A combination of molecular, histological, immunohistochemical, and behavioral assessments were used to test whether blocking cPLA2 activation pharmacologically or genetically reduced cell death, protected spinal cord tissue, and improved behavioral recovery after a contusive SCI performed at the 10th thoracic level in adult mice., Results: SCI significantly increased cPLA2 expression and activation. Activated cPLA2 was localized mainly in neurons and oligodendrocytes. Notably, the SCI-induced cPLA2 activation was mediated by the extracellular signal-regulated kinase signaling pathway. In vitro, activation of cPLA2 by ceramide-1-phosphate or A23187 induced spinal neuronal death, which was substantially reversed by arachidonyl trifluoromethyl ketone, a cPLA2 inhibitor. Remarkably, blocking cPLA2 pharmacologically at 30 minutes postinjury or genetically deleting cPLA2 in mice ameliorated motor deficits, and reduced cell loss and tissue damage after SCI., Interpretation: cPLA2 may play a key role in the pathogenesis of SCI, at least in the C57BL/6 mouse, and as such could be an attractive therapeutic target for ameliorating secondary tissue damage and promoting recovery of function after SCI., (© 2014 The Authors. American Neurological Association.)

Oligodendrocyte (OL) replacement can be a promising strategy for spinal cord injury (SCI) repair. However, the poor posttransplantation survival and inhibitory properties to axonal regeneration are two major challenges that limit their use as donor cells for repair of CNS injuries. Therefore, strategies aimed at enhancing the survival of grafted oligodendrocytes as well as reducing their inhibitory properties, such as the use of more permissive oligodendrocyte progenitor cells (OPCs), also called glial restricted precursor cells (GRPs), should be highly prioritized. Schwann cell (SC) transplantation is a promising translational strategy to promote axonal regeneration after CNS injuries, partly due to their expression and secretion of multiple growth-promoting factors. Whether grafted SCs have any effect on the biological properties of grafted GRPs remains unclear. Here we report that either SCs or SC-conditioned medium (SCM) promoted the survival, proliferation, and migration of GRPs in vitro. When GRPs and SCs were cografted into the normal or injured spinal cord, robust survival, proliferation, and migration of grafted GRPs were observed. Importantly, grafted GRPs differentiated into mature oligodendrocytes and formed new myelin on axons caudal to the injury. Finally, cografts of GRPs and SCs promoted recovery of function following SCI. We conclude that cotransplantation of GRPs and SCs, the only two kinds of myelin-forming cells in the nervous system, act complementarily and synergistically to promote greater anatomical and functional recovery after SCI than when either cell type is used alone.

The previous studies suggested that some subpopulations of T lymphocytes against central nervous system (CNS) antigens, such as myelin basic protein (MBP), are neuroprotective. But there were few reports about the effect of these T cells on axon regeneration. In this study, the neonatally thymectomied (Tx) adult rats which contain few T lymphocytes were subjected to spinal cord hemisection and then passively immunized with MBP-activated T cells (MBP-T). The regeneration and dieback of transected axons of cortico-spinal tract (CST) were detected by biotin dextran amine (BDA) tracing. The behavioral assessments were performed using the Basso, Beattie, and Bresnahan locomotor rating scale. We found that passive transferring of MBP-T could attenuate axonal dieback. However, no significant axon regeneration and behavioral differences were observed among the normal, Tx and sham-Tx (sTx) rats with or without MBP-T passive immunization. These results indicate that passive transferring of MBP-T cells can attenuate axonal dieback and promote neuroprotection following spinal cord injury (SCI), but may not promote axon regeneration.

Neural stem cell (NSC) transplantation is a major focus of current research for treatment of spinal cord injury (SCI). However, it is very important to promote the survival and differentiation of NSCs into myelinating oligodendrocytes (OLs). In this study, myelin basic protein-activated T (MBP-T) cells were passively immunized to improve the SCI microenvironment. Olig2-overexpressing NSCs were infected with a lentivirus carrying the enhanced green fluorescent protein (GFP) reporter gene to generate Olig2-GFP-NSCs that were transplanted into the injured site to differentiate into OLs. Transferred MBP-T cells infiltrated the injured spinal cord, produced neurotrophic factors, and induced the differentiation of resident microglia and/or infiltrating blood monocytes into an "alternatively activated" anti-inflammatory macrophage phenotype by producing interleukin-13. As a result, the survival of transplanted NSCs increased fivefold in MBP-T cell-transferred rats compared with that of the vehicle-treated control. In addition, the differentiation of MBP-positive OLs increased 12-fold in Olig2-GFP-NSC-transplanted rats compared with that of GFP-NSC-transplanted controls. In the MBP-T cell and Olig2-GFP-NSC combined group, the number of OL-remyelinated axons significantly increased compared with those of all other groups. However, a significant decrease in spinal cord lesion volume and an increase in spared myelin and behavioral recovery were observed in Olig2-NSC- and NSC-transplanted MBP-T cell groups. Collectively, these results suggest that MBP-T cell adoptive immunotherapy combined with NSC transplantation has a synergistic effect on histological and behavioral improvement after traumatic SCI. Although Olig2 overexpression enhances OL differentiation and myelination, the effect on functional recovery may be surpassed by MBP-T cells.

Demyelination contributes to the functional deficits after spinal cord injury (SCI). Therefore, remyelination may be an important strategy to facilitate repair after SCI. Oligodendrocyte precursor cells (OPCs) are immature oligodendrocytes and can differentiate into myelin-forming cells of central nervous system under certain conditions. OPC transplantation is an attractive approach for the treatment of demyelinating diseases. In this study, we transplanted OPCs expressing green fluorescent protein (GFP-OPCs) into normal and injured rat spinal cords to evaluate the differentiation of transplanted OPCs in vivo. Unfortunately, the grafted GFP-OPCs, in spinal cord whether normal or injured, were all differentiated into astrocytes, but not oligodendrocytes. Our further study indicated that inflammatory environment might not be the key factor influencing the differentiation of OPCs. Some spinal cord components, such as bone morphogenetic proteins (BMPs), were the major factors that induced OPCs to differentiate into astrocytes. The three types of BMP receptor (BMPRIA, IB and II) could all be detected in OPCs, and the astroglial differentiation of OPCs induced by spinal cord homogenate extract (SCHE) in vitro could be blocked partly by noggin, an antagonist of BMP. These results suggested that the BMPR signal transduction pathway might be one of the key factors which determine the differentiation direction of engrafted OPCs in spinal cord., (Copyright © 2010 International Society of Differentiation. Published by Elsevier B.V. All rights reserved.)

Background: Transplantation of oligodendrocyte precursor cells (OPCs) is an attractive therapy for demyelinating diseases. Cyclosporin A (CsA) is one of the foremost immunosuppressive agents and has widespread use in tissue and cell transplantation. However, whether CsA affects survival and differentiation of engrafted OPCs in vivo is unknown. In this study, the effect of CsA on morphological, functional and immunological aspects, as well as survival and differentiation of engrafted OPCs in injured spinal cord was explored., Results: We transplanted green fluorescent protein (GFP) expressed OPCs (GFP-OPCs) into injured spinal cords of rats treated with or without CsA (10 mg/kg). Two weeks after cell transplantation, more GFP-positive cells were found in CsA-treated rats than that in vehicle-treated ones. However, the engrafted cells mostly differentiated into astrocytes, but not oligodendrocytes in both groups. In the CsA-treated group, a significant decrease in spinal cord lesion volume along with increase in spared myelin and neurons were found compared to the control group. Such histological improvement correlated well with an increase in behavioral recovery. Further study suggested that CsA treatment could inhibit infiltration of T cells and activation of resident microglia and/or macrophages derived from infiltrating monocytes in injured spinal cords, which contributes to the survival of engrafted OPCs and repair of spinal cord injury (SCI)., Conclusions: These results collectively indicate that CsA can promote the survival of engrafted OPCs in injured spinal cords, but has no effect on their differentiation. The engrafted cells mostly differentiated into astrocytes, but not oligodendrocytes. The beneficial effect of CsA on SCI and the survival of engrafted cells may be attributed to its neuroprotective effect.

Post-traumatic inflammation response has been implicated in secondary injury mechanisms after spinal cord injury (SCI). Interleukin-1 (IL-1) is a key inflammatory mediator that is increasingly expressed after SCI. The action of IL-1 is mediated through its functional receptor, type I interleukin-1 receptor (IL-1RI). However, whether this receptor is expressed after SCI remains to be elucidated. In the present study, the temporospatial expression of IL-1RI was detected in rats that received a moderate contusive SCI (a 10 g rod dropped at a height of 12.5 mm) at the ninth to tenth thoracic vertebral level using a widely used New York University impact device. Our study demonstrated that IL-1RI was slightly increased at 4 h post-injury compared to the normal or sham-operated controls, reached the peak at 8 h at mRNA level (4.44-fold, P<0.01) and 1 d at protein level (2.62-fold, P<0.01). IL-1RI remained at its elevated levels for a relatively long duration (4 h-7 days). Spatially, IL-1RI was observed throughout the entire length of a 10 mm-long cord segment containing the injury epicenter. Colocalization of IL-1RI was found in neurons, oligodendrocytes, astrocytes, and activated microglia. Our results suggest that the elevated expression of IL-1RI after SCI may contribute to posttraumatic inflammation responses of IL-1.

Previous studies by our group have demonstrated that the transplantation of exogenous platelet-derived growth factor (PDGF)-AA-overexpressing oligodendrocyte progenitor cells (OPCs) promotes tissue repair and recovery of neurological function in a rat model of spinal cord injury (SCI). However, it remains unclear whether treatment with PDGF-AA also affects endogenous oligodendrocytes (OLs) or even neurons, thus promoting further functional recovery after SCI. In the present study, we evaluated the therapeutic potential of PDGF-AA treatment by direct subcutaneous injection of PDGF-AA immediately after SCI. We demonstrated that PDGF-AA injection resulted in increased tissue sparing, myelination and functional recovery in rats following SCI. Further experimentation confirmed that PDGF-AA increased the survival of endogenous OPCs and OLs, and promoted the proliferation of OPCs and their differentiation into OLs. Moreover, PDGF-AA also protected motor neurons from death in the injured spinal cord. These results indicated that PDGF-AA administration may be an effective treatment for SCI. [ABSTRACT FROM AUTHOR]