Spinal Cord Injury In Mice Research Articles

Methylprednisolone substituted lipid nanoparticles deliver C3 transferase mRNA for combined treatment of spinal cord injury

Spinal cord injury (SCI), characterized by the disruption of neural pathways and an increase in inflammatory cell infiltration, leads to profound and lasting neurological deficits, with a high risk of resulting in permanent disability. Currently, the therapeutic landscape for SCI is notably sparse, with limited effective treatment options available. Methylprednisolone (MP), a widely used clinical anti-inflammatory agent for SCI, requires administration in high doses that are associated with significant adverse effects. In this study, we introduce an innovative approach by substituting cholesterol with MP to engineer a novel Lipid Nanoparticle (MP-LNP). This strategy aims to enhance the localization and concentration of MP at the injury site, thereby amplifying its therapeutic efficacy while mitigating systemic side effects. Furthermore, we explore the integration of C3 transferase mRNA into MP-LNPs. C3 transferase, a potent inhibitor of the RhoA pathway, has shown promise in facilitating neurological recovery in animal models of SCI and is currently being evaluated in clinical trials. The novel formulation, MP-LNP-C3, is designed for direct administration to the injury site during decompression surgery, offering a targeted therapeutic modality for SCI. Our findings reveal several significant advantages of this approach: Firstly, the incorporation of C3 transferase mRNA into MP-LNPs does not compromise the structural integrity of the nanoparticles, ensuring efficient mRNA expression within the spinal cord. Secondly, the MP-LNP formulation effectively attenuates inflammation and reduces the adverse effects associated with high-dose MP treatment in the acute phase of SCI. Lastly, MP-LNP-C3 demonstrates notable neuroprotective properties and promotes enhanced recovery of motor function in SCI mouse models. Together, these results underscore the potential of this innovative LNP-based therapy as a promising avenue for advancing the treatment of clinical SCI.Graphical

Modulation of extrinsic and intrinsic signaling together with neuronal activation enhances forelimb motor recovery after cervical spinal cord injury.

Singular strategies for promoting axon regeneration and motor recovery after spinal cord injury (SCI) have been attempted with limited success. For instance, the deletion of RhoA and Pten (an extrinsic and intrinsic modulating factor, respectively) in corticospinal neurons promotes axon sprouting after thoracic SCI, however it is unable to restore motor function. Here, we examine the effects of combining RhoA/Pten deletion in corticospinal neurons with chemogenetic neuronal stimulation on axonal growth and motor recovery after SCI in mice. We find that this combinatorial approach promotes greater axonal growth and synaptic bouton formation in corticospinal neurons within the spinal cord compared to RhoA;Pten deletion alone. Furthermore, chemogenetic neuronal stimulation of RhoA;Pten-deleted corticospinal neurons improved forelimb performance in behavioral tasks after SCI compared to RhoA;Pten deletion alone. These results demonstrate that combination therapies pairing genetic modifications with neuronal stimulation can promote greater synaptic formation and motor recovery following SCI than either strategy alone.Significance Statement In this study, we examined whether pairing neural stimulation with genetic deletion of RhoA and Pten in corticospinal neurons would enhance axonal growth and motor recovery after spinal cord injury (SCI). We found that this combinatorial approach, relative to singular strategies, promoted greater presynaptic bouton formation in injured corticospinal neurons in the spinal cord, resulting in improved motor recovery in those mice. These results open the possibility for even greater enhancements in healing and recovery following SCI through carefully crafted, multifaceted treatments tailored to address the specific needs of individual SCI patients.

2-Mercaptoethanol enhances the yield of exosomes showing therapeutic potency in alleviating spinal cord injury mice

2-Mercaptoethanol enhances the yield of exosomes showing therapeutic potency in alleviating spinal cord injury mice

Microglial C/EBPβ-Fcgr1 regulatory axis blocking inhibits microglial pyroptosis and improves neurological recovery

CAAT/Enhancer Binding Protein β (C/EBPβ) is associated with inflammatory responses in neurodegenerative pathologies, particularly in the brain. However, the regulatory role of C/EBPβ in spinal cord injury and its impact on neurological recovery remain unknown. In this study, we observed significant upregulation of C/EBPβ in microglia after spinal cord injury in mice and was associated with neuroinflammation. Knocking down C/EBPβ in the spinal cord attenuated microglia pyroptosis, reduced the production of proinflammatory cytokines, and inhibited neuronal apoptosis. Mechanistically, C/EBPβ promoted the transcription of Fcgr1, which was involved in activating microglia pyroptosis. In both in-vivo and in-vitro experiments, knocking down Cebpb or Fcgr1, or the pyroptosis inhibitor VX765 inhibited neuronal apoptosis and improved neurological recovery in mice. These findings indicate that C/EBPβ functions as a key regulator that participates in the microglia pyroptosis-mediated neuroinflammation by activating Fcgr1 transcription.

Inhibition of CD36 ameliorates mouse spinal cord injury by accelerating microglial lipophagy.

Spinal cord injury (SCI) is a serious trauma of the central nervous system (CNS). SCI induces a unique lipid-dense environment that results in the deposition of large amounts of lipid droplets (LDs). The presence of LDs has been shown to contribute to the progression of other diseases. Lipophagy, a selective type of autophagy, is involved in intracellular LDs degradation. Fatty acid translocase CD36, a multifunctional transmembrane protein that facilitates the uptake of long-chain fatty acids, is implicated in the progression of certain metabolic diseases, and negatively regulates autophagy. However, the precise mechanisms of LDs generation and degradation in SCI, as well as whether CD36 regulates SCI via lipophagy, remain unknown. In this study, we investigated the role of LDs accumulation in microglia for SCI, as well as the regulatory mechanism of CD36 in microglia lipophagy during LDs elimination in vivo and in vitro. SCI was induced in mice by applying moderate compression on spina cord at T9-T10 level. Locomotion recovery was evaluated at days 0, 1, 3, 7 and 14 following the injury. PA-stimulated BV2 cells wasestablished as the in vitro lipid-loaded model. We observed a marked buildup of LDs in microglial cells at the site of injury post-SCI. More importantly, microglial cells with excessive LDs exhibited elevated activation and stimulated inflammatory response, which drastically triggered the pyroptosis of microglial cells. Furthermore, we found significantly increased CD36 expression, and the breakdown of lipophagy in microglia following SCI. Sulfo-N-succinimidyl oleate sodium (SSO), a CD36 inhibitor, has been shown to promote the lipophagy of microglial cells in SCI mice and PA-treated BV2 cells, which enhanced LDs degradation, ameliorated inflammatory levels and pyroptosis of microglial cells, and ultimately promoted SCI recovery. As expected, inhibition of lipophagy with Baf-A1 reversed the effects of SSO. We conclude that microglial lipophagy is essential for the removal of LDs during SCI recovery. Our research implies that CD36 could be a potential therapeutic target for the treatment and management of SCI.

Brain inflammation and cognitive decline induced by spinal cord injury can be reversed by spinal cord cell transplants.

Spinal cord injuries (SCIs) impact between 250,000 and 500,000 people worldwide annually, often resulting from road accidents or falls. These injuries frequently lead to lasting disabilities, with the severity depending on the injury's extent and location. Emerging research also links SCIs to cognitive impairments due to brain inflammation. From a treatment perspective, various approaches, including cell therapy, have been investigated. One common mechanism across cellular transplant models is the modulation of inflammation at the injury site, though it remains uncertain if these effects extend to the brain. To explore this, we induced SCI in wild-type mice and treated them with either olfactory ensheathing cells or mesenchymal stem cells. Our findings reveal that both cell types can reverse SCI-induced cognitive deficits, reduce brain inflammation, and increase hippocampal neuronal density. This study is the first, to our knowledge, to demonstrate that cells transplanted into the spinal cord can influence brain inflammation and mitigate injury-induced effects on brain functions. These results highlight the intricate relationship between the spinal cord and brain in both health and disease.

Nanoenzyme-Anchored Mitofactories Boost Mitochondrial Transplantation to Restore Locomotor Function after Paralysis Following Spinal Cord Injury.

Mitochondrial transplantation is a significant therapeutic approach for addressing mitochondrial dysfunction in patients with spinal cord injury (SCI), yet it is limited by rapid mitochondrial deactivation and low transfer efficiency. Here, high-quality mitochondria microfactories (HQ-Mitofactories) were constructed by anchoring Prussian blue nanoenzymes onto mesenchymal stem cells for effective mitochondrial transplantation to treat paralysis from SCI. Notably, the results demonstrated that HQ-Mitofactories could continuously produce vitality-boosting mitochondria with highly interconnected and elongated network structures under oxidative stress by scavenging excessive ROS. Furthermore, HQ-Mitofactories enabled efficient transfer of therapeutic mitochondria to injured neurons primarily via gap junctions, resulting in the restoration of mitochondrial homeostasis and thereby suppressing intracellular ROS burst and facilitating neuronal repair. After i.v. administration, HQ-Mitofactories migrated to the injured spinal cords of SCI mice and subsequently promoted neuronal regeneration and remyelination. Consequently, HQ-Mitofactory-treated mice successfully recovered locomotor function within 4 weeks, with 40% of the mice fully restoring walking after hindlimb paralysis. Conversely, untreated SCI exhibited completely abolished hindlimb movements. In light of real-time generation of vitality-boosting mitochondria even under oxidative stress and enabling targeted mitochondrial transfer, HQ-Mitofactories have promising therapeutic potential in the context of mitochondrial transplantation to reduce SCI-related paralysis, and more broadly impact the field of neuroregenerative medicine.

Astrocyte-conditional knockout of MOB2 inhibits the phenotypic conversion of reactive astrocytes from A1 to A2 following spinal cord injury in mice.

After spinal cord injury (SCI), reactive astrocytes in the injured area are triggered after spinal cord injury (SCI) and to polarize into A1 astrocytes with a proinflammatory phenotype or A2 astrocytes with an anti-inflammatory phenotype. Monopolar spindle binder 2 (MOB2) induces astrocyte stellation, maintains cell homeostasis, and promotes neurite outgrowth; however, its role in the phenotypic transformation of reactive astrocytes remains unclear. Here, we confirmed for the first time that MOB2 is associated with A1/A2 phenotypic switching in reactive astrocytes following SCI in mice. MOB2 modulated A1/A2 transformation in a primary astrocyte reactive cell model. Therefore, we constructed MOB2 conditional knockout mice (MOB2GFAP-CKO) and discovered that conditional knockout of MOB2 inhibited the conversion of reactive astrocytes from A1 to A2 and hindered spinal cord function recovery. Mechanistically, MOB2 increased the activation of PI3K-AKT signaling to promote A1/A2 transformation in vitro, whereas sc79 (an AKT activator) reversed the subtype transformation of reactive astrocytes and improved functional recovery in MOB2GFAP-CKO mice after SCI. Taken together, study provides the first insights into how MOB2 acts as a novel regulator to promote the conversion this of the reactive astrocyte phenotype from A1 to A2, showing great potential for the treatment of SCI.

Temporal and spatial pattern of DNA damage in neurons following spinal cord Injury in mice

BackgroundDeficient DNA repair and excessive DNA damage contribute to neurodegenerative disease. However, the role of DNA damage and repair in spinal cord injury (SCI) is unclear. SCI, a debilitating disruption of the structural and biological network of the spinal cord, is characterized by oxidative stress. Nevertheless, the pathophysiological mechanisms leading to neuronal loss following SCI remain incompletely defined. Methods: Using a contusion model, a severe SCI was induced at the L1 spinal level in C57Bl/6J mice. The temporal and spatial presence of DNA damage was then determined via immunolabeling for the DNA damage marker, γH2AX, from 1 h post-injury (hpi) to 28 days post-injury (dpi). Results: Our analysis revealed that increased DNA damage foci were present from 1 hpi to 3 dpi in SCI mice relative to controls (sham surgery and naive), with the damage signal spreading over time longitudinally from the affected area to more rostral and caudal regions. Co-labeling of γH2AX with NeuN revealed neuronal specificity of DNA damage, with increased early cell death (pan-nuclear γH2AX) peaking at 1 dpi and apoptosis (cleaved Caspase-3) arising later at 3 dpi. Conclusion: Our study indicates a possible role of DNA damage in neuronal loss following SCI and highlights the need for early interventions targeting DNA repair to preserve neuronal tissue.

Neuronal Dual-Specificity Phosphatase 26 Inhibition via Reactive-Oxygen-Species Responsive Mesoporous-Silica-Loaded Hydrogel for Spinal Cord Injury Repair.

Spinal cord injury (SCI) remains a formidable challenge in biomedical research, as the silencing of intrinsic regenerative signals in most spinal neurons results in an inability to reestablish neural circuits. In this study, we found that neurons with low axonal regeneration after SCI showed decreased extracellular signal-regulated kinase (ERK) phosphorylation levels. However, the expression of dual specificity phosphatase 26 (DUSP26)─which negatively regulates ERK phosphorylation─was reduced considerably in neurons undergoing spontaneous axonal regeneration. Therefore, we developed a system named F10@MS@UV-HG that integrated a DUSP26-specific inhibitor into reactive oxygen species-responsive nanoparticles and embedded them in photosensitive hydrogels. This system effectively downregulated DUSP26 expression in primary neurons and enhanced ERK phosphorylation, ultimately promoting axonal outgrowth. When transplanted into an SCI mouse model, the system achieved sustained drug release, specifically targeting the DUSP26/ERK/ELK1 pathway in the spinal neurons and facilitating short-term axonal regeneration. Additionally, long-term repair effects─including improved myelination and enhanced motor function─were evident in the SCI mice transplanted with F10@MS@UV-HG. The results suggested that activating ERK signaling by modulating DUSP26 expression in neurons after SCI could effectively promote axonal regeneration and functional recovery. Thus, the developed F10@MS@UV-HG system exhibits enormous potential as a therapeutic approach for patients with SCI.

An optogenetic mouse model of hindlimb spasticity after spinal cord injury.

Spasticity is a common comorbidity of spinal cord injury (SCI), disrupting motor function and resulting in significant discomfort. While elements of post-SCI spasticity can be assessed using pre-clinical SCI models, the robust measurement of spasticity severity can be difficult due to its periodic and spontaneous appearance. Electrical stimulation of sensory afferents can elicit spasticity-associated motor responses, such as spasms; however, placing surface electrodes on the hindlimbs of awake animals can induce stress or encumbrance that could influence the expression of behaviour. Therefore, we have generated a mouse model of SCI-related spasticity that utilizes optogenetics to activate a subset of cutaneous VGLUT2+ sensory afferents to produce reliable incidences of spasticity-associated responses in the hindlimb. To examine the efficacy of this optogenetic SCI spasticity model, a T9-T10 complete transection injury was performed in Islet1-Cre+/-;VGLUT2-Flp+/-;CreON-FlpON-CatCh+/- mice, followed by the implantation of EMG electrodes into the left and right gastrocnemius and tibialis anterior muscles. EMG recordings were performed during episodic optogenetic stimulation (1-2 sessions per week until 5 weeks post-injury (wpi); n = 10 females, 5 males). A subset of these mice (n = 3 females, 2 males) was also tested at 10 wpi. During each recording session, an optic fiber coupled to a 470 nm wavelength LED was used to deliver 9 × 100 ms light pulses to the palmar surface of each hind paw. The results of these recordings demonstrated significant increases in the amplitude of EMG responses to the light stimulus from 2 wpi to 10 wpi, suggesting increased excitability of cutaneous sensorimotor pathways. Interestingly, this effect was significantly greater in the female cohort than in the males. Incidences of prolonged involuntary muscle contraction in response to the stimulus (fictive spasms) were also detected through EMG and visual observation during the testing period, supporting the presence of spasticity. As such, the optogenetic mouse model developed for this study appears to elicit spasticity-associated behaviours in SCI mice reliably and may be valuable for studying SCI-related limb spasticity mechanisms and therapeutic.

LncRNA SNHG6 Knockdown Promotes Microglial M2 Polarization and Alleviates Spinal Cord Injury via Regulating the miR-182-5p/NEUROD4 Axis.

Spinal cord injury (SCI) is one of the devastating neurological disorders that leads to a loss of motor and sensory functions. Long non-coding RNA small nucleolar RNA host gene 6 (lncRNA SNHG6) plays a crucial role in inflammatory regulation across various diseases. This study investigates the role of SNHG6 in SCI development and its underlying regulatory mechanisms. Two experimental models were established: an in vitro model using LPS-challenged (100ng/mL) mouse microglia BV2 cells and an in vivo model employing controlled spinal cord impact in mice. SNHG6, miR-182-5p, and NEUROD4 expression levels were quantified through RT-qPCR and Western blot. Functional and histological assessments were performed using the Basso mouse scale (BMS) and Nissl staining, respectively. Putative binding sites between SNHG6 and miR-182-5p, as well as between miR-182-5p and NEUROD4, were predicted using the ENCORI/starBase platform. These molecular interactions were validated through dual-luciferase reporter assays and RNA pull-down experiments, with further confirmation by qRT-PCR and Western blot analyses. Both LPS-stimulated BV2 cells and spinal cord tissues from SCI mice exhibited elevated SNHG6 expression. Downregulation of SNHG6 enhanced LPS-induced polarization of BV2 cells from M1-type to M2-type, significantly modulated the expression of pro-inflammatory factors (TNF-α, IL-1β, and IL-6) and anti-inflammatory factors (TGF-β, IL-10, and IL-13), and reduced injury severity in SCI mice. Our mechanistic studies revealed that SNHG6 functions as a molecular sponge for miR-182-5p to regulate NEUROD4 expression. This study demonstrates that SNHG6 knockdown promotes microglial M2-type polarization and alleviates inflammatory responses through modulation of the miR-182-5p/NEUROD4 axis, suggesting SNHG6 as a potential therapeutic target for SCI treatment.

Acute Extrinsic Activation of the RANKL Pathway Decreases Wound Healing and Functional Recovery After Spinal Cord Injury in Mice.

Manipulating wound healing-associated signaling after SCI presents a promising avenue for increasing the recovery of function after injury. This study explores the potential of targeting molecular regulators of wound healing, initially identified in nonneural tissues, to enhance outcomes after SCI. Astrocytes, pivotal in central nervous system wound healing, play a crucial role in tissue remodeling and recovery. However, the optimal manipulation of astrogliosis for beneficial outcomes remains elusive. Previous research demonstrated a transcriptional response in astrocytes resembling epithelial-to-mesenchymal transitions (EMTs) after CNS injury. Here, we investigate the extrinsic manipulation of wound healing through the Receptor Activator of Nuclear-factor Kappa-Β (RANK) pathway, known for its involvement in nonneural tissue remodeling and linked to EMT pathway. Using a severe thoracic spinal cord contusion mouse model, we demonstrate that acute activation of the RANK pathway with RANK ligand (RANKL) adversely affects tissue remodeling, resulting in larger lesion volumes and delayed recovery of posture and locomotion. These findings suggest that early perturbations in the tight molecular regulation of tissue remodeling negatively impact the wound-healing process after SCI. The study provides a proof-of-concept demonstration that exogenous nonneural remodeling ligands can modify astrocyte responses and functional recovery after SCI, raising questions about the optimal time frame for beneficial remodeling interventions during injury progression. These insights open new avenues for therapeutic strategies aimed at improving functional outcomes following SCI.

Spinal cord injury enhances lung inflammation and exacerbates immune response following exposure to LPS.

The severity of spinal cord injury (SCI) is closely tied to pulmonary function, especially in cases of higher SCI levels. Despite this connection, the underlying pathological mechanisms in the lungs post-SCI are not well understood. Previous research has established a connection between disrupted sympathetic regulation and splenocyte apoptosis in high thoracic SCI, leading to pulmonary dysfunction. The aim of this study was to investigate whether mice with low-level SCI exhibit increased susceptibility to acute lung injury by eliciting systemic inflammatory responses that operate independently of the sympathetic nervous system. Here, we employed T9 contusion SCI and exposed mice to aerosolized lipopolysaccharide (LPS) to simulate lung inflammation associated with acute respiratory distress syndrome (ARDS). Twenty-four hours post-LPS exposure, lung tissues and bronchoalveolar lavage (BAL) fluid were analyzed. LPS markedly induced proinflammatory gene expression (SAA3, IRG1, NLRP3, IL-1beta, MCP-1) and cytokine release (IL-1beta, IL-6, MCP-1) in SCI mice compared to controls, indicating an exaggerated inflammatory response. Infiltration of Ly6G/C positive neutrophils and macrophages was significantly higher in SCI mice lungs post-LPS exposure. Interestingly, spleen size and weight did not differ between control and SCI mice, suggesting that T9 SCI alone does not cause spleen atrophy. Notably, bone-marrow-derived macrophages (BMDMs) from SCI mice exhibited hyper-responsiveness to LPS. This study demonstrated an increase in lung inflammation and immune responses subsequent to low-level T9 SCI, underscoring the widespread influence of systemic inflammation post-SCI, especially pronounced in specific organs like the lungs.

Baicalin ameliorates neuroinflammation by targeting TLR4/MD2 complex on microglia via PI3K/AKT/NF-κB signaling pathway.

This study aims to elucidate the target and mechanism of baicalin, a clinically utilized drug, in the treatment of neuroinflammatory diseases. Neuroinflammation, characterized by the activation of glial cells and the release of various pro-inflammatory cytokines, plays a critical role in the pathogenesis of various diseases, including spinal cord injury (SCI). The remission of such diseases is significantly dependent on the improvement of inflammatory microenvironment. Toll-like receptor 4/myeloid differentiation protein 2 (TLR4/MD2) complex plays an important role in pathogen recognition and innate immune activation. baicalin, a natural flavonoid, is renowned for its potent anti-inflammatory property. In this study, we discovered that baicalin significantly reduced the activation of glial cells and the levels of pro-inflammatory cytokines at the lesion site of SCI mice, thereby mitigating demyelination and neuronal damage. By directly occupying the active pocket of TLR4/MD2 complex on microglia, baicalin inhibited PI3K/AKT/NF-κB pathway, thereby exerting its anti-inflammatory effect. These findings were corroborated in mice induced by lipopolysaccharide, a TLR4 agonist. Furthermore, baicalin indirectly altered phenotype of astrocytes by reducing secretion of TNF-α, IL-1α, and C1q levels from microglia. Our work demonstrated that baicalin effectively alleviated neuroinflammation by directly targeting microglia and indirectly modulating astrocytes phenotype. As a natural flavonoid, baicalin holds significant potential as a therapeutic candidate for diseases characterized by neuroinflammation.

Single-Cell Multi-omics Assessment of Spinal Cord Injury Blocking via Cerium-doped Upconversion Antioxidant Nanoenzymes.

Spinal cord injury (SCI) impairs the central nervous system and induces the myelin-sheath-deterioration because of reactive oxygen species (ROS), further hindering the recovery of function. Herein, the simultaneously emergency treatment and dynamic luminescence severity assessment (SETLSA) strategy is designed for SCI based on cerium (Ce)-doped upconversion antioxidant nanoenzymes (Ce@UCNP-BCH). Ce@UCNP-BCH can not only efficiently eliminate the SCI localized ROS, but dynamically monitor the oxidative state in the SCI repair process using a ratiometric luminescence signal. Moreover, the classic basso mouse scale score and immunofluorescence analysis together exhibit that Ce@UCNP-BCH effectively facilitates the regeneration of spinal cord including myelin sheath, and promotes the functional recovery of SCI mice. Particularly, the study combines snATAC-eq and snRNA-seq to reveal the heterogeneity of spinal cord tissue following Ce@UCNP-BCH treatment. The findings reveal a significant increase in myelinating oligodendrocytes, as well as higher expression of myelination-related genes, and the study also reveals the gene regulatory dynamics of remyelination after treatment. Besides, the ETLSA strategy synergistically boosts ROS consumption through the superoxide dismutase (SOD)-related pathways after SOD-siRNA treatment. In conclusion, this SETLSA strategy with simultaneously blocking and dynamic monitoring oxidative stress has enriched the toolkit for promoting SCI repair.

Cycloastragenol promotes dorsal column axon regeneration in mice.

Cycloastragenol (CAG) has a wide range of pharmacological effects, including anti-inflammatory, antiaging, antioxidative, and antitumorigenic properties. In addition, our previous study showed that CAG administration can promote axonal regeneration in peripheral neurons. However, whether CAG can activate axon regeneration central nervous system (CNS) remains unknown. Here, we established a novel mouse model for visualizing spinal cord dorsal column axon regeneration involving the injection of AAV2/9-Cre into the lumbar 4/5 dorsal root ganglion (DRG) of Rosa-tdTomato reporter mice. We then treated mice by intraperitoneal administration of CAG. Our results showed that intraperitoneal CAG injections significantly promoted the growth of vitro-cultured DRG axons as well as the growth of dorsal column axons over the injury site in spinal cord injury (SCI) mice. Our results further indicate that CAG administration can promote the recovery of sensory and urinary function in SCI mice. Together, our findings highlight the therapeutic potential of CAG in spinal cord injury repair.

Lentiviral Injection of Inter-α-Trypsin Inhibitor Heavy Chain 4 Promotes Female Spinal Cord Injury Mice Recuperation by Diminishing Peripheral and Central Inflammation.

Inter-alpha-trypsin inhibitor heavy chain H4 (ITIH4) acts as a mediator of inflammation and extracellular matrix stabilization. The current study intended to delve into the impact of ITIH4 on locomotor performance, nerve injury, neuroinflammation, systemic inflammation, and the downstream pathway in spinal cord injury (SCI) mice. Overexpression lentivirus of ITIH4 (LV-ITIH4) and negative control lentivirus (LV-NC) were intravenously injected into adult C57BL/6 mice on 7days before SCI surgery. All mice were euthanized on day 28 after SCI surgery, and their blood samples and spinal cord tissues were collected. Decreased relative gene expression and protein levels of ITIH4 were observed in SCI mice. LV-ITIH4 improved the locomotor performance compared to LV-NC in SCI mice. In spinal cord of SCI mice, LV-ITIH4 reduced apoptosis and increased survival of neurons compared to LV-NC. By comparison with LV-NC, LV-ITIH4 also reduced relative gene expressions of interleukin (IL)-6 and tumor necrosis factor-α in spinal cord of SCI mice. Moreover, LV-ITIH4 reduced microglia M1 polarization compared with LV-NC in spinal cord of SCI mice. In the serum, LV-ITIH4 decreased the protein levels of IL-6 and IL-1β compared to LV-NC in SCI mice. LV-ITIH4 also inhibited the nuclear factor kappa-B (NF-κB) pathway compared to LV-NC in spinal cord of SCI mice. ITIH4 enhances locomotor performance in SCI mice, and it inhibits nerve injury, neuroinflammation, systemic inflammation, and the NF-κB pathway in SCI mice.

Systemic administrations of protamine heal subacute spinal cord injury in mice.

Spinal cord injury (SCI) results in damage to neural circuits that cause long-term locomotor and sensory disability. The objective of the present study is to evaluate whether a clinical drug, protamine, can be employed as a therapeutic agent for SCI. First, we examined the rescue effect of protamine on dystrophic endballs (DEs) cultured on a chondroitin sulfate (CS) gradient coating. Consequently, axons with DE, which are unable to grow through the CS barrier, resumed growth after protamine treatment and were able to pass through the barrier. In addition, we tested whether protamine resolves the DE phenotype, accumulation of autophagosomes. The results demonstrated that protamine has significantly reduced the density of LC3 in DEs. Subsequently, mice were administered 1mg/kg protamine via the tail vein one week following a contusion injury to the thoracic spinal cord. The hindlimb movements of the mice were evaluated in order to assess the therapeutic effect of protamine. Eleven venous administrations of protamine improved the symptoms. The current study has demonstrated that protamine cancels the CS inhibitory effect on axonal regrowth. Administrations of protamine were observed to alleviate hindlimb motor dysfunction in SCI mice. Our results suggest an effective therapeutic agent for SCI and a possibility for drug repositioning. It would be of interest to see if protamine also exerts a therapeutic effect in brain injury.

Effect of Sakuranetin on Microglia-Mediated Neuroinflammation After Spinal Cord Injury.

Objective To investigate the effects of sakuranetin (SK) on motor functions in the mouse model of spinal cord injury (SCI) and decipher the mechanism. Methods Fifty-four C57BL/6J mice were randomized into sham,SCI,and SK groups.The mice in the sham group underwent only laminectomy at T9,while those in the SCI and SK groups were subjected to spinal cord contusion injury at T9.Behavioral tests were conducted at different time points after surgery to evaluate the motor functions of mice in each group.The pathological changes in the tissue were observed to assess the extent of SCI in each group.The role and mechanism of SK in SCI were predicted by gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) enrichment analyses.Reverse transcription real-time fluorescence quantitative PCR,ELISA,and immunofluorescence were employed to evaluate the inflammation and activation of microglia in SCI mice.BV2 cells in vitro were classified into control (Con),lipopolysaccharide (LPS),and LPS+SK groups.The effects of SK intervention on the release of inflammatory cytokines and the activation of BV2 cells were evaluated.Furthermore,the phosphatidylinositol-3-kinase(PI3K)/protein kinase B (AKT) signaling pathway activator insulin-like growth factor-1 (IGF-1) was used to treat the SK-induced BV2 cells in vitro (SK+IGF-1 group),and SK was used to treat the IGF-1-induced BV2 cells in vitro (IGF-1+SK group).Western blotting was conducted for molecular mechanism validation. Results Behavioral tests and histological staining results showed that compared with the SCI group,the SK group exhibited improved motor abilities and reduced area of damage in the spinal cord tissue (all P<0.001).The GO enrichment analysis predicted that SK may be involved in the inflammation following SCI.The KEGG enrichment analysis predicted that SK regulated the PI3K/Akt pathway to exert the neuroprotective effect.The results from in vitro and in vivo experiments showed that SK lowered the levels of tumor necrosis factor-α,interleukin-6,and interleukin-1β and inhibited the activation of microglia (all P<0.05).The results of Western blotting showed that SK down-regulated the phosphorylation levels of PI3K and Akt (all P<0.001) and inhibited the IGF-1-induced elevation of PI3K and Akt phosphorylation levels (all P<0.001).Conversely,IGF-1 had the opposite effects (P=0.001,P<0.001).The results of reverse transcription real-time fluorescence quantitative PCR,ELISA,and immunofluorescence showed that the SK+IGF-1 group had higher levels of inflammatory cytokines and more activated microglia than the SK group(all P<0.05). Conclusion SK may suppress the activation of the PI3K/Akt pathway to inhibit the inflammation mediated by SCI-induced activation of microglia,ameliorate the pathological damage of the spinal cord tissue,and promote the recovery of motor functions in SCI mice.