Progression Of Spinal Cord Injury Research Articles

Dysregulated MiR-223-5p Modulates Inflammation and Oxidative Stress in Traumatic Spinal Cord Injury

ABSTRACT Aim This study aimed to evaluate the miR-223-5p expression in patients with spinal cord injury (SCI) and to determine its role in the pathogenesis of SCI. Methods The serum miR-223-5p levels were analyzed using quantitative real-time polymerase chain reaction. The diagnostic accuracy of miR-223-5p was evaluated using the receiving operating characteristic curves. LPS-induced PC12 cells were established as an in vitro inflammatory cell model. Cell apoptosis, inflammation and oxidative stress were examined. The SCI rat model was constructed to evaluate the effects of miR-223-5p on inflammatory response and motor function in rats. Results MiR-223-5p expression was upregulated in SCI patients. MiR-223-5p expression in the complete SCI group was significantly higher than that in incomplete SCI group. ROC analysis showed that miR-223-5p can distinguish SCI patients from healthy volunteers. In vitro experiments demonstrated that LPS upregulated apoptosis and inflammation in PC12 cells. Treatment with miR-223-5p inhibitor alleviated the changes in LPS-induced PC12 cells . Inhibition of miR-223-5p can alleviate the activation of inflammatory response and the effects of SCI on the motor function in rats. Conclusions MiR-223-5p is a potential diagnostic marker for SCI, and it can promote the SCI progression by regulating nerve cell survival, inflammation, and oxidative stress.

Of Potential Noncoding RNAs Related to Spinal Cord Injury Based on Competing Endogenous RNAs.

This study aims to elucidate the key regulatory molecules, specifically messenger RNAs (mRNAs), long noncoding RNAs (lncRNAs), and microRNAs (miRNAs) and their roles in the development and progression of spinal cord injury (SCI). Expression profiles (GSE45006, GSE19890, and GSE125630) for SCI were sourced from the Gene Expression Omnibus (GEO) database. By comparing rats with SCI at various time points against those without SCI, we identified differentially expressed mRNAs (DEmRNAs), lncRNAs (DElncRNAs), and miRNAs (DEmiRNAs). The GSE45006 dataset facilitated the production of DEmRNAs, which were then clustered using Mfuzz. Subsequently, we constructed a protein-protein interaction (PPI) network and anticipated interaction pairs between miRNA-mRNA and lncRNA-mRNA. These pairs were instrumental in forming a regulatory network involving lncRNA-miRNA-mRNA interactions. Additionally, we conducted functional enrichment studies on the DEmRNAs within these gene networks. A total of 2313 DEmRNAs were identified using the GSE45006 dataset, alongside 111 DEmiRNAs from GSE19890. From GSE125630, we extracted 154 DElncRNAs and 2322 DEmRNAs. Our analysis revealed 294 up-regulated DEmRNAs, grouped into the up-cluster, and 407 down-regulated DEmRNAs, forming the down-cluster. Key hub genes in the PPI network, such as Rhof, Vav1, Lyz2, Rab3a, Lyn, Cyfip1, Gns, and Nckap1l, were identified. Additionally, the study successfully constructed a competing endogenous RNA (ceRNA) network, revealing 55 unique lncRNA-miRNA-mRNA link pairs. Our research established a ceRNA network associated with SCI, identifying several critical lncRNA-miRNA-mRNA connection pairs integral to the disease's onset and progression. Notably, significant associations, including the AABR07041411.1-miR-125a-5p-Slc4a7 and the Smg1-rno-miR-331-3p-Tlr4 pairs, were observed to exert a significant influence within this biological context.

Targeting foamy macrophages by manipulating ABCA1 expression to facilitate lesion healing in the injured spinal cord

Spinal cord injury (SCI) triggers a complex cascade of events, including myelin loss, neuronal damage, neuroinflammation, and the accumulation of damaged cells and debris at the injury site. Infiltrating bone marrow derived macrophages (BMDMϕ) migrate to the epicenter of the SCI lesion, where they engulf cell debris including abundant myelin debris to become pro-inflammatory foamy macrophages (foamy Mϕ), participate neuroinflammation, and facilitate the progression of SCI. This study aimed to elucidate the cellular and molecular mechanisms underlying the functional changes in foamy Mϕ and their potential implications for SCI. Contusion at T10 level of the spinal cord was induced using a New York University (NYU) impactor (5 g rod from a height of 6.25 mm) in male mice. ABCA1, an ATP-binding cassette transporter expressed by Mϕ, plays a crucial role in lipid efflux from foamy cells. We observed that foamy Mϕ lacking ABCA1 exhibited increased lipid accumulation and a higher presence of lipid-accumulated foamy Mϕ as well as elevated pro-inflammatory response in vitro and in injured spinal cord. We also found that both genetic and pharmacological enhancement of ABCA1 expression accelerated lipid efflux from foamy Mϕ, reduced lipid accumulation and inhibited the pro-inflammatory response of foamy Mϕ, and accelerated clearance of cell debris and necrotic cells, which resulted in functional recovery. Our study highlights the importance of understanding the pathologic role of foamy Mϕ in SCI progression and the potential of ABCA1 as a therapeutic target for modulating the inflammatory response, promoting lipid metabolism, and facilitating functional recovery in SCI.

TRIM32 Inhibits NEK7 Ubiquitylation-Dependent Microglia Pyroptosis After Spinal Cord Injury.

Spinal cord injury (SCI) is a disabling disease associated with microglial activation. Tripartite motif containing 32 (TRIM32) is an E3 ubiquitin ligase that plays a role in SCI. This study aimed to explore the role of TRIM32 in SCI and its potential mechanisms. We established an SCI mouse model to assess the function of TRIM32 using quantitative real-time polymerase chain reaction (qPCR), and hematoxylin and eosin staining. Additionally, a lipopolysaccharides (LPS)-induced cell injury model was generated to explore the impact of TRIM32 on pyroptosis using qPCR, propidium iodide staining, and western blotting. The ubiquitylation of NEK7 was analyzed using western blotting, co-immunoprecipitation, and immunofluorescence staining. The results showed that TRIM32 expression was increased in SCI mice and LPS-induced BV-2 cells. Overexpression of TRIM32 ameliorated SCI in mice and suppressed pyroptosis in LPS-treated BV-2 cells. Additionally, the E3 ligase TRIM32 promoted the ubiquitylation of NEK7 at the K64 site, leading to the downregulation of NEK7 levels. Inhibiting NEK7 ubiquitylation reversed the suppression of pyroptosis by TRIM32. In conclusion, TRIM32 inhibits microglia pyroptosis by facilitating the ubiquitylation of NEK7 at the K64 site, thereby alleviating the progression of SCI. The findings suggest that TRIM32 has the potential to be a therapeutic target of SCI.

Function of GSK‑3 signaling in spinal cord injury (Review).

Spinal cord injury (SCI) is a major social problem with a heavy burden on patient physiology and psychology. Glial scar formation and irreversible neuron loss are the two key points during SCI progression. During the acute phase of spinal cord injury, glial scars form, limiting the progression of inflammation. However, in the subacute or chronic phase, glial scarring inhibits axon regeneration. Following spinal cord injury, irreversible loss of neurons leads to further aggravation of spinal cord injury. Several therapies have been developed to improve either glial scar or neuron loss; however, few therapies reach the stage of clinical trials and there are no mainstream therapies for SCI. Exploring the key mechanism of SCI is crucial for finding further treatments. Glycogen synthase kinase-3 (GSK-3) is a widely expressed kinase with important physiological and pathophysiological functions in vivo. Dysfunction of the GSK-3 signaling pathway during SCI has been widely discussed for controlling neurite growth in vitro and in vivo, improving the proliferation and neuronal differentiation of endogenous neural stem cells and functional recovery from spinal cord injury. SCI can decrease the phosphorylated (p)/total (t)-GSK-3β ratio, which leads to an increase in apoptosis, whereas treatment with GSK-3 inhibitors can promote neurogenesis. In addition, several therapies for the treatment of SCI involve signaling pathways associated with GSK-3. Furthermore, signaling pathways associated with GSK-3 also participate in the pathological process of neuropathic pain that remains following SCI. The present review summarized the roles of GSK-3 signaling in SCI to aid in the understanding of GSK-3 signaling during the pathological processes of SCI and to provide evidence for the development of comprehensive treatments.

Drug Repurposing for Spinal Cord Injury: Progress Towards Therapeutic Intervention for Primary Factors and Secondary Complications.

Spinal cord injury (SCI) encompasses a plethora of complex mechanisms like the involvement of major cell death pathways, neurodegeneration of spinal cord neurons, overexpression of glutaminergic transmission and inflammation cascade, along with different co-morbidities like neuropathic pain, urinary and sexual dysfunction, respiratory and cardiac failures, making it one of the leading causes of morbidity and mortality globally. Corticosteroids such as methylprednisolone and dexamethasone, and non-steroidal anti-inflammatory drugs such as naproxen, aspirin and ibuprofen are the first-line treatment options for SCI, inhibiting primary and secondary progression by preventing inflammation and action of reactive oxygen species. However, they are constrained by a short effective drug administration window and their pharmacological action being limited to symptomatic relief of the secondary effects related to spinal cord injury only. Although post-injury rehabilitation treatments may enable functional recovery, they take a long time to show results. Drug repurposing might be an innovative method for expanding therapy alternatives, utilising drugs that are already approved by various esteemed federal agencies throughout the world. Reutilising a drug molecule to treat SCI can eliminate the need for expensive and lengthy drug discovery processes and pave the way for new therapeutic approaches in SCI. This review summarises marketed drugs that could be repurposed based on their safety and efficacy data. We also discuss their mechanisms of action and provide a list of repurposed drugs under clinical trials for SCI therapy.

FGF21 prevents neuronal cell ferroptosis after spinal cord injury by activating the FGFR1/β-Klotho pathway

Spinal cord injury (SCI) is a kind of traumatic nervous system disease caused by neuronal death, causing symptoms like sensory, motor, and autonomic nerve dysfunction. The recovery of neurological function has always been a intractable problem that has greatly distressed individuals and society. Although the involvement of iron-dependent lipid peroxidation leading to nerve cell ferroptosis in SCI progression has been reported, the underlying mechanisms remain unaddressed. Thus, this study aimed to investigate the potential of recombinant human FGF21 (rhFGF21) in inhibiting ferroptosis of nerve cells and improving limb function after SCI, along with its underlying mechanisms. In vivo animal model showed that FGFR1, p-FGFR1, and β-Klotho protein gradually increased over time after injury, reaching a peak on the third day. Moreover, rhFGF21 treatment significantly reduced ACSL4, increased GPX4 expression, reduced iron deposition, and inhibited ferroptosis. Meanwhile, rhFGF21 decreased cell apoptosis following acute spinal cord damage. In contrast, FGFR1 inhibitor PD173074 partially reversed the rhFGF21-induced therapeutic effects. Overall, this work revealed that rhFGF21 activates the FGFR1/β-Klotho pathway to decrease ferroptosis of nerve cells, suggesting that FGF21 could be a new therapeutic target for SCI neurological rehabilitation.

A cutting-edge strategy for spinal cord injury treatment: resident cellular transdifferentiation.

Spinal cord injury causes varying degrees of motor and sensory function loss. However, there are no effective treatments for spinal cord repair following an injury. Moreover, significant preclinical advances in bioengineering and regenerative medicine have not yet been translated into effective clinical therapies. The spinal cord's poor regenerative capacity makes repairing damaged and lost neurons a critical treatment step. Reprogramming-based neuronal transdifferentiation has recently shown great potential in repair and plasticity, as it can convert mature somatic cells into functional neurons for spinal cord injury repair in vitro and in vivo, effectively halting the progression of spinal cord injury and promoting functional improvement. However, the mechanisms of the neuronal transdifferentiation and the induced neuronal subtypes are not yet well understood. This review analyzes the mechanisms of resident cellular transdifferentiation based on a review of the relevant recent literature, describes different molecular approaches to obtain different neuronal subtypes, discusses the current challenges and improvement methods, and provides new ideas for exploring therapeutic approaches for spinal cord injury.

Knockout of microglial Hv1 proton channel reduces neurotoxic A1 astrocytes and neuronal damage via the ROS/STAT3 pathway after spinal cord injury.

Spinal cord injury (SCI) causes severe functional deficits and neuronal damage, accompanied by intense glial activation. The voltage-gated proton channel Hv1, selectively expressed on microglia, is associated with SCI progression. However, the effect of Hv1 on the phenotypes and functions of reactive astrocytes after SCI remains unclear. Here, we combined Hv1 knockout (Hv1-/- ) mice and T10 spinal cord contusion to investigate the effects of microglial Hv1 on SCI pathophysiology and the phenotypes and functions of reactive astrocytes. After SCI, astrocytes proliferated and activated in the peri-injury area and exhibited an A1-dominant phenotype. Hv1 knockout reduced neurotoxic A1 astrocytes and shifted the dominant phenotype of reactive astrocytes from A1 to A2, enhancing synaptogenesis promotion, phagocytosis, and neurotrophy of astrocytes. Moreover, synaptic and axonal remodeling as well as motor recovery after SCI benefited from the improved astrocytic functions of Hv1 knockout. Furthermore, exogenous and endogenous reactive oxygen species (ROS) in astrocytes after SCI were reduced by Hv1 knockout. Our in vitro results showed that inhibition of ROS reduced the neurotoxic A1 phenotype in primary astrocytes via the STAT3 pathway. Similar to the effect of Hv1 knockout, the application of the ROS scavenger N-acetylcysteine reduced SCI-induced neurotoxic A1 astrocytes in vivo. Based on the in vivo and vitro results, we elucidated that microglial Hv1 knockout promotes synaptic and axonal remodeling in SCI mice by decreasing neurotoxic A1 astrocytes and increasing neuroprotective A2 astrocytes via the ROS/STAT3 pathway. Therefore, the Hv1 proton channel is a promising target for the treatment of SCI.

LncRNA TSIX aggravates spinal cord injury by regulating the PI3K/AKT pathway via the miR-532-3p/DDOST axis.

Long noncoding RNA (lncRNA)-X-inactive-specific transcript (TSIX) expression is upregulated in spinal cord tissues following spinal cord injury (SCI). However, the role of lncRNA-TSIX in SCI remains elusive. SCI animal model was established using C57BL/6 mice. LncRNA TSIX and miR-532-3p expression were determined usingquantitative reverse transcription polymerase chain reaction (qRT-PCR). Apoptosis, cell proliferation, and migration were evaluated by transferase dUTP nick end labeling staining, CCK-8, and Transwell assays, respectively. The interaction of miR-532-3p with lncRNA TSIX and DDOST was explored via a dual-luciferase reporter system. Hematoxylin-eosin staining and the Basso, Beattie, and Bresnahan locomotor rating (BBB) scale were performed to investigate SCI progression. The expression of the lncRNA TSIX was found to be significantly upregulated in the serum of SCI patients and spinal cord tissues of SCI mice. The overexpression of lncRNA TSIX enhanced spinal cord neural stem cell (SC-NSC) proliferation and migration in vitro while inhibiting apoptosis and inflammatory cell infiltration in vivo. Moreover, lncRNA TSIX acted as a molecular sponge for miR-532-3p, and the knockdown of miR-532-3p promoted proliferation and migration and inhibited apoptosis of SC-NSCs. Moreover, DDOST was found to be the downstream target of miR-532-3p, and DDOST overexpression showed a similar effect as miR-532-3p silencing on the proliferation, migration, and apoptosis of SC-NSCs. Furthermore, we found that lncRNA TSIX overexpression promoted the activation of the PI3K/AKT signaling pathway. LncRNA TSIX aggravates SCI by regulating the PI3K/AKT pathway via the miR-532-3p/DDOST axis, indicating potential applications for targeted therapy of SCI regeneration.

Patients with Chronic Spinal Cord Injury and a Long Period of Evolution Exhibit an Altered Cytokine Production by CD4 and CD8 T Cell Populations.

Spinal cord injury (SCI) is a disabling neurological condition coursing with serious multisystem affections and morbidities. Changes in immune cell compartments have been consistently reported in previous works, representing a critical point of study for understanding the pathophysiology and progression of SCI from acute to chronic stages. Some relevant variations in circulating T cells have been noticed in patients with chronic SCI, although the number, distribution, and function of these populations remain to be fully elucidated. Likewise, the characterization of specific T cell subpopulations and their related cytokine production can aid in understanding the immunopathological role of T cells in SCI progression. In this sense, the objective of the present study was to analyze and quantify the total number of different cytokine-producers T cells in the serum of patients with chronic SCI (n = 105) in comparison to healthy controls (n = 38) by polychromatic flow cytometry. Having this goal, we studied CD4 and CD8 lymphocytes as well as naïve, effector, and effector/central memory subpopulations. SCI patients were classified according to the duration of the lesion in chronic SCI with a short period of evolution (SCI-SP) (comprised between 1 and 5 years since initial injury), early chronic phase (SCI-ECP) (between 5 and 15 years since initial injury) and late-chronic phase (SCI-LCP) (>15 years since initial injury). Our results show that patients with chronic SCI exhibited an altered immune profile of cytokine-producer T cells, including CD4/CD8 naïve, effector, and memory subpopulations in comparison to HC. In particular, IL-10 and IL-9 production seems to be importantly altered, especially in patients with SCI-LCP, whereas changes in IL-17, TNF-α, and IFN-γ T cell populations have also been reported in this and other chronic SCI groups. In conclusion, our study demonstrates an altered profile of cytokine-producer T cells in patients with chronic SCI, with marked changes throughout the course of the disease. In more detail, we have observed significant variations in cytokine production by circulating naive, effector, and effector/central memory CD4 and CD8 T cells. Future studies should be directed to explore the possible clinical consequences of these changes or develop additional translational approaches in these groups of patients.

The involvement of EGR1 in neuron apoptosis in the in vitro model of spinal cord injury via BTG2 up-regulation

ABSTRACT Objective EGR1 has been implicated in the progression of spinal cord injury (SCI). Nevertheless, its specific mechanism in SCI remains to be investigated. Hence, this study explored the potential mechanism of EGR1 in SCI by focusing on neuron apoptosis. Methods H2O2 was utilized to treat rat neurons-dorsal spinal cord (RN-dsc) for the construction of an in vitro model of SCI. Afterwards, cell survival, apoptosis, and LDH leakage were detected to evaluate the injury degree of H2O2-treated RN-dsc. The expression of apoptosis-related proteins was also measured. Additionally, EGR1 was silenced and/or BTG2 was overexpressed in RN-dsc before H2O2 treatment to assess the impacts of EGR1 and BTG2 on H2O2-induced RN-dsc. Jasper online website was utilized to predict binding sites of EGR1 on BTG2, and dual-luciferase reporter gene and chromatin immunoprecipitation (ChIP) assays were utilized to verify the binding between EGR1 and BTG2. Results H2O2 treatment suppressed survival and promoted apoptosis in RN-dsc, accompanied by upregulated LDH, Bax, and cleaved-caspase-3 and down-regulated Bcl-2. Moreover, EGR1 and BTG2 were up-regulated in H2O2-induced RN-dsc. Mechanistically, EGR1 was bound to the promoter of BTG2 to transcriptionally activate BTG2. EGR1 knockdown diminished apoptosis and LDH, Bax, and cleaved-caspase-3 levels while elevating survival and Bcl-2 levels in H2O2-induced RN-dsc. These effects of EGR1 knockdown were abrogated by further BTG2 overexpression. Discussion Conclusively, EGR1 promotes H2O2-induced apoptosis in RN-dsc by activating BTG2 transcription.

Phosphoglycerate mutase 5 facilitates mitochondrial dysfunction and neuroinflammation in spinal tissues after spinal cord injury

Spinal cord injury (SCI) is a high incidence worldwide that causes a heavy physical and psychological burden to patients. It is urgent to further reveal the pathological mechanism and effective treatment of SCI. Mitochondrial dysfunction plays an important role in the disease progression of SCI. As a mitochondrial membrane protein, phosphoglycerate mutase 5 (PGAM5) is mainly involved in mitochondrial function and mitosis to modulate cellular physiological functions, but the roles of PGAM5 in spinal tissues remain to be unreported after SCI. The purpose of this study was to evaluate the role of PGAM5 in SCI mice and its relationship with neuroinflammation. The results showed that the mitochondrial membrane protein PGAM5 was involved in microglia activation after SCI, and PGAM5 deletion could improve mitochondrial dysfunction (including abnormal mtDNA, ATP synthases, and ATP levels, Cyt C expression, and ROS and rGSH levels) in spinal cord tissue after SCI, Arg1/iNOS mRNA level, iNOS expression, and pro-inflammatory cytokines TNF-α, IL-1β, and IL-18 levels. In vitro, H2O2 increased TNF-α, IL-1β, and IL-18 levels in BV2 cells, and PGAM5-sh and Nrf2 activators significantly reversed H2O2-induced iNOS expression and proinflammatory cytokine production. Furthermore, IP/Western blotting results revealed that PGAM5-sh treatment significantly reduced the interaction of PGAM5 with Nrf2 and enhanced the nuclear translocation of Nrf2 in BV2 cells. The data suggested that PGAM5 was involved in the cascade of oxidative stress and inflammatory response in microglia via facilitating the expression level of Nrf2 in the nucleus after SCI. It provided a reference for clarifying the pathological mechanism and therapeutic target of SCI.

Bioinformatics analysis of ferroptosis in spinal cord injury.

Ferroptosis plays a key role in aggravating the progression of spinal cord injury (SCI), but the specific mechanism remains unknown. In this study, we constructed a rat model of T10 SCI using a modified Allen method. We identified 48, 44, and 27 ferroptosis genes that were differentially expressed at 1, 3, and 7 days after SCI induction. Compared with the sham group and other SCI subgroups, the subgroup at 1 day after SCI showed increased expression of the ferroptosis marker acyl-CoA synthetase long-chain family member 4 and the oxidative stress marker malondialdehyde in the injured spinal cord while glutathione in the injured spinal cord was lower. These findings with our bioinformatics results suggested that 1 day after SCI was the important period of ferroptosis progression. Bioinformatics analysis identified the following top ten hub ferroptosis genes in the subgroup at 1 day after SCI: STAT3, JUN, TLR4, ATF3, HMOX1, MAPK1, MAPK9, PTGS2, VEGFA, and RELA. Real-time polymerase chain reaction on rat spinal cord tissue confirmed that STAT3, JUN, TLR4, ATF3, HMOX1, PTGS2, and RELA mRNA levels were up-regulated and VEGFA, MAPK1 and MAPK9 mRNA levels were down-regulated. Ten potential compounds were predicted using the DSigDB database as potential drugs or molecules targeting ferroptosis to repair SCI. We also constructed a ferroptosis-related mRNA-miRNA-lncRNA network in SCI that included 66 lncRNAs, 10 miRNAs, and 12 genes. Our results help further the understanding of the mechanism underlying ferroptosis in SCI.

Dysregulation of miR-301a-3p serves as a non-invasive biomarker and is associated with inflammatory responses in traumatic spinal cord injury.

Spinal cord injury (SCI) led by trauma is a serious damage in central nervous system. This study aimed at confirming the expression levels and clinical significance of miRNA-301a-3p (miR-301a-3p) in SCI patients, and exploring the potential mechanisms of miR-301a-3p participating in SCI progression. One hundred and thirty nine acute spinal trauma patients, included neurologically normal, incomplete and complete SCI cases, were analyzed in this study. Quantitative real-time PCR was used to measure the expression of miR-301a-3p. Receiver operating characteristic (ROC) was used to evaluate the diagnostic accuracy of serum miR-301a-3p in SCI. LPS-treated SH-SY5Y cells were used to mimic SCI inflammatory injury. The levels of IL-1β, IL-6, TNF-α were detected using enzyme-linked immunosorbent assay. Expression of serum miR-301a-3p was significantly higher in both incomplete and complete SCI patients than that in neurologically normal controls, and had a significant ability to distinguish SCI patients from controls. Additionally, complete SCI cases had markedly increased miR-301a-3p compared to incomplete cases, and the two groups of patients could be distinguished based on serum deregulated miR-301a-3p. In the SCI cell model, miR-301a-3p overexpression led to decreased cell viability. The inflammatory responses of the cell model were enhanced by miR-301a-3p, which was consistent with the findings in SCI patients that serum miR-301a-3p was positively correlated with pro-inflammatory cytokines. The expression of miR-301a-3p is increased in SCI patients, and serves as a candidate biomarker for SCI diagnosis. MiR-301a-3p may be involved in SCI progression by affecting neuronal viability and inflammation.

Decellularization alters the unfavorable regenerative adverse microenvironment of the injured spinal cord to support neurite outgrowth.

BackgroundAcellular tissue has been transplanted into the injury site as an external microenvironment to intervene with imbalance microenvironment that occurs after spinal cord injury (SCI) and stimulating axonal regeneration, although the mechanism is unclear. Given decellularization is the key means to obtain acellular tissues, we speculated changes in the internal components of tissue caused by decellularization may be the key reason why acellular tissues affect remodeling of the microenvironment.MethodsComplete spinal cord crush in a mouse model was established, and the dynamic of extracellular matrix (ECM) expression and distribution during SCI was studied with immunohistochemistry (IHC). Normal spinal cord (NSC) and 14-day injury spinal cord (ISC) were obtained to prepare the decellularized NSC (DNSC) and decellularized ISC (DISC) through a well-designed decellularization method, and the decellularization effects were evaluated by residual DNA content determination, hematoxylin and eosin staining (H&E), and IHC. Rat dorsal root ganglia (DRG) were co-cultured with NSC, ISC, DNSC, and DISC to evaluate their effect on neurite outgrowth. Furthermore, the mechanisms by which decellularized tissue promotes axonal growth were explored with proteomics analysis of the protein components and function of 14-day ISC and DISC.ResultsWe found the expression of the four main ECM components (collagen type I and IV, fibronectin, and laminin) gradually increased with the progression of SCI compared to NSC, peaking at 14 days of injury then slightly decreasing at 21 days, and the distribution of the four ECM proteins in the ISC also changed dynamically. H&E staining, residual DNA content determination, and IHC showed decellularization removed cellular components and preserved an intact ECM. The results of co-cultured DRG with NSCs, ISCs, DNSCs, and DISCs showed DNSCs and DISCs had a stronger ability in supporting neurite outgrowth than NSC and ISC. We found through proteomics that decellularization could remove proteins associated with inflammatory responses, scarring, and other pathological factors, while completely retaining the ECM proteins.ConclusionsTaken together, our findings demonstrate decellularization can optimize the imbalanced microenvironment after SCI by removing components that inhibit spinal cord regeneration, providing a theoretical basis for clinical application of acellular tissue transplantation to repair SCI.

Inflammation: A Target for Treatment in Spinal Cord Injury.

Spinal cord injury (SCI) is a significant cause of disability, and treatment alternatives that generate beneficial outcomes and have no side effects are urgently needed. SCI may be treatable if intervention is initiated promptly. Therefore, several treatment proposals are currently being evaluated. Inflammation is part of a complex physiological response to injury or harmful stimuli induced by mechanical, chemical, or immunological agents. Neuroinflammation is one of the principal secondary changes following SCI and plays a crucial role in modulating the pathological progression of acute and chronic SCI. This review describes the main inflammatory events occurring after SCI and discusses recently proposed potential treatments and therapeutic agents that regulate inflammation after insult in animal models.

Self-assembly of gelatin microcarrier-based MSC microtissues for spinal cord injury repair

Current approaches for treating spinal cord injury (SCI) are mainly based on cell transplantation. Mesenchymal stem cells (MSCs) can help slow the progression of SCI due to their trophic function. However, SCI creates a complex microenvironment that reduces cell activity and hence cellular function, ultimately resulting in poor therapeutic outcomes. To help maintain function in transplanted cells, we produced functional tissue constructs by self-assembly of MSC microtissues comprising of porous gelatin microcarriers (GM) and MSCs. These microtissues maintained cellular activity without incurring an excessive amount of apoptosis and delayed senescence in vitro. The paracrine function of MSCs also improved within microtissues, shown by the increased secretion of nerve regeneration-related factors. Microtissues were transplanted in a rat model of complete spinal cord transection, and therapeutic effects were evaluated through behavioral measurements, imaging, histology, and western blot analysis. RNA-seq of spinal cord tissues using Gene Ontology analysis further revealed that the microtissues may have induced repair in SCI through mechanisms related to neurotrophin-3 (NT-3) regulation of response mediator protein 2 (CRMP2) phosphorylation, and inhibition of inflammatory response through interleukin-17 (IL-17), Chemokine C-X-C motif Ligand 1 (CXCL1) axis. The gelatin microcarrier-based MSC microtissues we developed may be effective in providing a new treatment strategy for SCI.

LncRNA RMRP Contributes to the Development and Progression of Spinal Cord Injury by Regulating miR-766-5p/FAM83A Axis.

Spinal cord injury (SCI) is known as a central nervous system disorder. Previous studies suggested that long-non-coding RNA RMRP (LncRNA RMRP) was abnormally expressed in SCI, but the potential underlying mechanism remains to be further explored. To explore the regulatory roles of LncRNA RMRP/miR-766-5p/FAM83A axis in SCI. Spinal T9 hemisection was performed on healthy adult male Sprague Dawley (SD) rats to establish the SCI rat models. The expressions of LncRNA RMRP in spinal cord of rats in different groups were examined by RT-qPCR. Moreover, AGE1.HN and PC12 cells were treated with hypoxic condition, and expression of LncRNA RMRP was examined by RT-qPCR methods. Furthermore, hypoxic PC12 cells were transfected with LncRNA RMRP OE, and the cell viability, proliferation, and apoptosis were examined. Next, the direct targeting relationship between LncRNA RMRP and miR-766-5p, as well as miR-766-5p and FAM83A, was confirmed by dual-luciferase reporter and RNA pull-down assays. Finally, the effects of LncRNA RMRP/miR-766-5p/FAM83A axis on cell viability, proliferation, and apoptosis were examined. LncRNA RMRP was downregulated in SCI rats and over-expression of LncRNA RMRP alleviated the SCI condition. LncRNA RMRP over-expression increased the viability and proliferation, and inhibited the apoptosis of hypoxic PC12 cells in vitro. miR-766-5p was confirmed as a target of LncRNA RMRP, and FAM83A was confirmed as a target of miR-766-5p. LncRNA RMRP could regulate the proliferation and apoptosis of hypoxic PC12 cells via regulating miR-766-5p/FAM83A axis in vitro. LncRNA RMRP may contribute to the pathogenesis of SCI via regulating miR-766-5p/FAM83A axis.

Revealing Potential Spinal Cord Injury Biomarkers and Immune Cell Infiltration Characteristics in Mice.

Spinal cord injury (SCI) is a disabling condition with significant morbidity and mortality. Currently, no effective SCI treatment exists. This study aimed to identify potential biomarkers and characterize the properties of immune cell infiltration during this pathological event. To eliminate batch effects, we concurrently analyzed two mouse SCI datasets (GSE5296, GSE47681) from the GEO database. First, we identified differentially expressed genes (DEGs) using linear models for microarray data (LIMMA) and performed functional enrichment studies on those DEGs. Next, we employed bioinformatics and machine-learning methods to identify and define the characteristic genes of SCI. Finally, we validated them using immunofluorescence and qRT-PCR. Additionally, this study assessed the inflammatory status of SCI by identifying cell types using CIBERSORT. Furthermore, we investigated the link between key markers and infiltrating immune cells. In total, we identified 561 robust DEGs. We identified Rab20 and Klf6 as SCI-specific biomarkers and demonstrated their significance using qRT-PCR in the mouse model. According to the examination of immune cell infiltration, M0, M1, and M2 macrophages, along with naive CD8, dendritic cell-activated, and CD4 Follicular T cells may have a role in the progression of SCI. Therefore, Rab20 and Klf6 could be accessible targets for diagnosing and treating SCI. Moreover, as previously stated, immune cell infiltration may significantly impact the development and progression of SCI.