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Spinal cord injury (SCI) is a devastating neurotraumatic condition characterized by severe motor dysfunction and paralysis. Accumulating evidence suggests that DNA damage is involved in SCI pathology. However, the underlying mechanisms remain elusive. Although checkpoint kinase 1 (Chk1)-regulated DNA damage is involved in critical cellular processes, its role in SCI regulation remains unclear. This study aimed to explore the role and potential mechanism of Chk1 in SCI-induced motor dysfunction. Adult female C57BL/6J mice subjected to T9-T10 spinal cord contusions were used as models of SCI. Western blotting, immunoprecipitation, histomorphology, and Chk1 knockdown or overexpression achieved by adeno-associated virus were performed to explore the underlying mechanisms. Levels of p-Chk1 and γ-H2AX (a cellular DNA damage marker) were upregulated, while ferroptosis-related protein levels, including glutathione peroxidase 4 (GPX4) and x-CT were downregulated, in the spinal cord and hippocampal tissues of SCI mice. Functional experiments revealed increased Basso Mouse Scale (BMS) scores, indicating that Chk1 downregulation promoted motor function recovery after SCI, whereas Chk1 overexpression aggravated SCI-induced motor dysfunction. In addition, Chk1 downregulation reversed the SCI-increased levels of GPX4 and x-CT expression in the spinal cord and hippocampus, while immunoprecipitation assays revealed strengthened interactions between p-Chk1 and GPX4 in the spinal cord after SCI. Finally, Chk1 downregulation promoted while Chk1 overexpression inhibited NeuN cellular immunoactivity in the spinal cord after SCI, respectively. Collectively, these preliminary results imply that Chk1 is a novel regulator of SCI-induced motor dysfunction, and that interventions targeting Chk1 may represent promising therapeutic targets for neurotraumatic diseases such as SCI., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Spinal cord injury (SCI) induces the disruption of the blood-spinal cord barrier (BSCB) and the failure of axonal growth. SCI activates a complex series of responses, including cell apoptosis and endoplasmic reticulum (ER) stress. Pericytes play a critical role in maintaining BSCB integrity and facilitating tissue growth and repair. However, the roles of pericytes in SCI and the potential mechanisms underlying the improvements in functional recovery in SCI remain unclear. Recent evidence indicates that irisflorentin exerts neuroprotective effects against Parkinson's disease; however, whether it has potential protective roles in SCI or not is still unknown. In this study, we found that the administration of irisflorentin significantly inhibited pericyte apoptosis, protected BSCB integrity, promoted axonal growth, and ultimately improved locomotion recovery in a rat model of SCI. In vitro, we found that the positive effects of irisflorentin on axonal growth were likely to be mediated by regulating the crosstalk between pericytes and neurons. Furthermore, irisflorentin effectively ameliorated ER stress caused by incubation with thapsigargin (TG) in pericytes. Meanwhile, the protective effect of irisflorentin on BSCB disruption is strongly related to the reduction of pericyte apoptosis via inhibition of ER stress. Collectively, our findings demonstrate that irisflorentin is beneficial for functional recovery after SCI and that pericytes are a valid target of interest for future SCI therapies., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2024 Elsevier Inc. All rights reserved.)

After spinal cord injury, astrocytes undergo a reactive process and form an astroglial scar, which impedes the regeneration of axons. The role of Runx2 in promoting the transformation of astrocytes in the central nervous system is well-established. However, it remains unclear whether Runx2 also plays a role in the development of astroglial scar, and the precise underlying mechanism has yet to be identified. Recently, our study using cell culture and animal models has demonstrated that Runx2 actually suppresses astrocyte activation and the formation of astroglial scar following injury. The initial results demonstrated an increase in the expression of Runx2 in astrocytes following in vivo injury. Subsequently, the overexpression of Runx2 resulted in the inhibition of astrocyte activation, reduction in the total area of astroglial scar, and restoration of neural function after 14 days of injury. However, these effects were reversed by CADD522. These findings indicate that Runx2 could potentially serve as a therapeutic intervention for spinal cord injury (SCI). Furthermore, our findings suggest that the Nuclear-matrix-targeting signal (NMTS) of Runx2 is associated with its effect. In summary, the study's results propose that targeting Runx2 may be a promising treatment approach for reactive astrocytes and astroglial scar in the recovery of SCI., Competing Interests: Declarations. Ethics Approval: The animal study was reviewed and approved by The Animal Care and Use Committee of Wenzhou Medical University. Consent for Publication: Not applicable. Consent to Participate: Not applicable. Competing Interests: The authors declare no competing interests., (© 2024. The Author(s).)

After spinal cord injury (SCI), significant alterations in the tissue microenvironment lead to mitochondrial dysfunction, inducing apoptosis and inhibiting the remodeling of neural circuits, thereby impeding recovery. Although previous studies have demonstrated a marked decrease in pH at the injury site, creating an acidic microenvironment, the impact of improving this acidic microenvironment on SCI recovery has not been investigated. This study prepared a lysine@hollow mesoporous silica nanoparticle/gelatin methacrylate (GelMA) (L@H/G) composite hydrogel. The L@H/G composite hydrogel was demonstrated to release lysine and efficiently improve the acidic microenvironment slowly. Significantly, the composite hydrogel reduced cell apoptosis, promoted nerve regeneration, inhibited glial scar formation, and ultimately enhanced motor function recovery in mice with SCI. Mechanistically, the L@H/G hydrogel improved the mitochondrial tricarboxylic acid (TCA) cycle and fatty acid metabolism, restoring energy supply and facilitating mitochondrial function recovery. To the best of our knowledge, this is the first report confirming that improving the acidic microenvironment could promote SCI repair, providing a potential therapeutic strategy for SCI.

Endoplasmic reticulum (ER) stress-induced apoptosis and autophagy flux blockade significantly contribute to neuronal pathology of spinal cord injury (SCI). Yet, the molecular interplay between these two distinctive pathways in mediating the pathology of SCI remains largely unexplored. Currently, we aimed at exploring the crucial role of Stub1 in maintaining ER homeostasis and regulating autophagic flux after SCI. Our results demonstrate that Stub1 reduces ER stress induced neuronal apoptosis, promotes axonal regeneration, inhibits glial scar formation and fosters functional recovery by restoring autophagic flux following SCI. Stub1 enhances autophagic flux following SCI by alleviating the permeabilization of lysosomal membrane through activating TFEB. Importantly, we showed that Stub1 promotes the activation of TFEB by targeting HDAC2 for ubiquitination and degradation. Furthermore, the neuroprotective effect of Stub1 on SCI was abrogated by chloroquine administration, underscoring the essential role of Stub1-mediated enhancement of autophagic flux in its protective effects against SCI. Collectively, our data highlights the vital role of Stub1 in regulating ER stress and autophagy flux after SCI, and propose its potential as a promising target for neuroprotective interventions in SCI., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Background: Increasing evidence suggests that acute traumatic spinal cord injury (SCI)-induced defects in autophagy and autophagy-lysosomal pathway (ALP) may contribute to endothelial barrier disruption following injury. Recently, Kruppel-like factor 2 (KLF2) was reported as a key molecular switch on regulating autophagy. Whether KLF2 coordinates endothelial endothelial ALP in SCI is not known. Methods: Genetic manipulations of KLF2 were performed in bEnd.3 cells and SCI model. Western blot, qRT-PCR, immunofluorescence staining and Lyso-Tracker Red staining, Evans blue dye extravasation, behavioral assessment via Basso mouse scale (BMS), electrophysiology and footprint analysis were performed. Results: In SCI, autophagy flux disruption in endothelial cells contributes to TJ proteins degradation, leading to blood-spinal cord barrier (BSCB) impairment. Furthermore, the KLF2 level was decreased in SCI, overexpression of which alleviated TJ proteins loss and BSCB damage, which improve motor function recovery in SCI mice, while knockdown of KLF2 displayed the opposite effects. At the molecular level, KLF2 overexpression alleviated the TJ proteins degradation and the endothelial permeability by tuning the ALP dysfunction caused by SCI and oxygen glucose deprivation (OGD). Conclusions: Endothelial KLF2 as one of the key contributors to SCI-mediated ALP dysfunction and BSCB disruption. KLF2 could be a promising pharmacological target for the management and treatment of SCI., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

The repair of spinal cord injury (SCI) is still a tough clinical challenge and needs innovative therapies. Mitochondrial function is significantly compromised after SCI and has emerged as an important factor causing neuronal apoptosis and hindering functional recovery. In this study, umbilical cord mesenchymal stem cells (UCMSC), which are promising seed cells for nerve regeneration, and basic fibroblast growth factor (bFGF) that have been demonstrated to have a variety of effects on neural regeneration were jointly immobilized in extracellular matrix (ECM) and heparin-poloxamer (HP) to create a polymer bioactive system that brings more hope and possibility for the treatment of SCI. Our results in vitro and in vivo showed that the UCMSC-bFGF-ECM-HP thermosensitive hydrogel has good therapeutic effects, mainly in reducing apoptosis and improving the mitochondrial function. It showed promising utility for the functional recovery of impaired mitochondrial function by promoting mitochondrial fusion, reducing pathological mitochondrial fragmentation, increasing mitochondrial energy supply, and improving the metabolism of MDA, LDH, and ROS. In addition, we uncovered a distinct molecular mechanism underlying the protective effects associated with activating p21-activated kinase 1 (PAK1) and mitochondrial sirtuin 4 (SIRT4) by the UCMSC-bFGF-ECM-HP hydrogel. The expansion of new insights into the molecular relationships between PAK1 and SIRT4, which links the mitochondrial function in SCI, can lay the foundation for future applications and help to provide promising interventions of stem-cell-based biological scaffold therapies and potential therapeutic targets for the clinical formulation of SCI treatment strategies.

The disability rate of spinal cord injury (SCI) is extremely high, and stem cell inhibition is one of the most effective schemes in treating the spinal cord, but the survival rate is extremely low after stem cell transplantation, so it cannot be widely used in clinic. Studies have revealed that loading stem cells with biological scaffolds can effectively improve the survival rate and effect after stem cell transplantation. Therefore, this research was devised to analyze the repair effect of thiolated chitosan nanocarriers scaffold carrying de-epithelized human amniotic epithelial cells (HAECs) on SCI. And we used thiolated chitosan as nanocarriers, aiming to provide a reliable theoretical basis for future clinical practice. Through experiments, we concluded that the Tarlov and BBB scores of rats with SCI were raised under the intervention of thiolated chitosan carrying HAECs, while the inflammatory factors in serum, oxidative stress reaction in spinal cord tissue, apoptosis rate of nerve cells, and autophagy protein expression were all suppressed. Thus, the thiolated chitosan carrying HAECs may be applied to treat SCI by suppressing autophagy protein expression, oxidative stress response, and release of inflammatory factors in spinal cord tissue, which may be a new clinical therapy for SCI in the future. Even though we cannot understand exactly the therapeutic mechanism of thiolated chitosan carrying HAECs for SCI, the real clinical application of thiolated chitosan carrying HAECs needs to be confirmed by human experiments., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2022 Lijuan Zhu et al.)

Axon growth and neuronal apoptosis are considered to be crucial therapeutic targets against spinal cord injury (SCI). Growing evidences have reported stimulation of glucagon-like peptide-1 (GLP-1)/GLP-1 receptor (GLP-1R) signalling axis provides neuroprotection in experimental models of neurodegeneration disease. Endogenous GLP-1 is rapidly degraded by dipeptidyl peptidase-IV (DPP4), resulting in blocking of GLP-1/GLP1R signalling process. Sitagliptin, a highly selective inhibitor of DPP4, has approved to have beneficial effects on diseases in which neurons damaged. However, the roles and the underlying mechanisms of sitagliptin in SCI repairing remain unclear. In this study, we used a rat model of SCI and PC12 cells/primary cortical neurons to explore the mechanism of sitagliptin underlying SCI recovery. We discovered the expression of GLP-1R decreased in the SCI model. Administration of sitagliptin significantly increased GLP-1R protein level, alleviated neuronal apoptosis, enhanced axon regeneration and improved functional recovery following SCI. Nevertheless, treatment with exendin9-39, a GLP-1R inhibitor, remarkably reversed the protective effect of sitagliptin. Additionally, we detected the AMPK/PGC-1α signalling pathway was activated by sitagliptin stimulating GLP-1R. Taken together, sitagliptin may be a potential agent for axon regrowth and locomotor functional repair via GLP-1R-induced AMPK/ PGC-1α signalling pathway after SCI., (© 2020 The Authors. Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)

Background and Aim: Increasing evidence suggests that spinal cord injury (SCI)-induced defects in autophagic flux may contribute to an impaired ability for neurological repair following injury. Transcription factor E3 (TFE3) plays a crucial role in oxidative metabolism, lysosomal homeostasis, and autophagy induction. Here, we investigated the role of TFE3 in modulating autophagy following SCI and explored its impact on neurological recovery. Methods: Histological analysis via HE, Nissl and Mason staining, survival rate analysis, and behavioral testing via BMS and footprint analysis were used to determine functional recovery after SCI. Quantitative real-time polymerase chain reaction, Western blotting, immunofluorescence, TUNEL staining, enzyme-linked immunosorbent assays, and immunoprecipitation were applied to examine levels of autophagy flux, ER-stress-induced apoptosis, oxidative stress, and AMPK related signaling pathways. In vitro studies using PC12 cells were performed to discern the relationship between ROS accumulation and autophagy flux blockade. Results: Our results showed that in SCI, defects in autophagy flux contributes to ER stress, leading to neuronal death. Furthermore, SCI enhances the production of reactive oxygen species (ROS) that induce lysosomal dysfunction to impair autophagy flux. We also showed that TFE3 levels are inversely correlated with ROS levels, and increased TFE3 levels can lead to improved outcomes. Finally, we showed that activation of TFE3 after SCI is partly regulated by AMPK-mTOR and AMPK-SKP2-CARM1 signaling pathways. Conclusions: TFE3 is an important regulator in ROS-mediated autophagy dysfunction following SCI, and TFE3 may serve as a promising target for developing treatments for SCI., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

After spinal cord injury (SCI), the inhibitory molecules derived from scars at the lesion sites and the limited regenerative capacity of neuronal axons pose difficulties for the recovery after SCI. Remodeling of cytoskeleton structures including microtubule assembly and tubulin post-translational modification are widely accepted to play a crucial role in initiation of growth cone and regrowth of injured axon. Although increasing studies have focused on the association between tubulin acetylation and autophagy due to the role of tubulin acetylation in organelles and substances transport, there are no studies exploring the effect of tubulin acetylation on autophagy after spinal cord injury (SCI). Here, we found that histone deacetylase 6 (HDAC6) was significantly up-regulated after SCI, while inhibition of HDAC6 by Tubastatin A induced functional recovery after SCI. In view of enzyme-dependent and -independent mechanisms of HDAC6 to adjust diverse cellular processes, such as autophagy, the ubiquitin proteasome system and post-translational modification of tubulin, we mainly focused on the significance of HDAC6 in axonal regeneration and autophagy after SCI. Western blotting, Co-immunoprecipitation and immunofluorescence staining were conducted to showed that Tubastatin A treatment in nocodazole-treated cells and mice suffering from SCI prompted acetylation and stabilization of microtubules and thus restored transport function, which may contribute to restored autophagic flux and increased axonal length. Whereas inhibition of degradation of autolysosomes by bafilomycin A1 (Baf-A1) reversed functional recovery caused by Tubastatin A, revealing the association between tubulin acetylation and autophagy, which supports HDAC6 inhibition as a potential target for SCI treatment., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Spinal cord injury (SCI) is a devastating disease that may lead to lifelong disability. Thus, seeking for valid drugs that are beneficial to promoting axonal regrowth and elongation after SCI has gained wide attention. Metformin, a glucose-lowering agent, has been demonstrated to play roles in various central nervous system (CNS) disorders. However, the potential protective effect of metformin on nerve regeneration after SCI is still unclear. In this study, we found that the administration of metformin improved functional recovery after SCI through reducing neuronal cell apoptosis and repairing neurites by stabilizing microtubules via PI3K/Akt signaling pathway. Inhibiting the PI3K/Akt pathway with LY294002 partly reversed the therapeutic effects of metformin on SCI in vitro and vivo. Furthermore, metformin treatment weakened the excessive activation of oxidative stress and improved the mitochondrial function by activating the nuclear factor erythroid-related factor 2 (Nrf2) transcription and binding to the antioxidant response element (ARE). Moreover, treatment with Nrf2 inhibitor ML385 partially abolished its antioxidant effect. We also found that the Nrf2 transcription was partially reduced by LY294002 in vitro. Taken together, these results revealed that the role of metformin in nerve regeneration after SCI was probably related to stabilization of microtubules and inhibition of the excessive activation of Akt-mediated Nrf2/ARE pathway-regulated oxidative stress and mitochondrial dysfunction. Overall, our present study suggests that metformin administration may provide a potential therapy for SCI., Competing Interests: The authors confirm that there are no conflicts of interest., (Copyright © 2020 Haoli Wang et al.)

Axon retraction greatly limits functional recovery after spinal cord injury (SCI) and neuron polarization, which affects processes including axon formation and development, is a promising target for promoting axon regeneration. Increasing microtubule stability has been demonstrated to improve intrinsic axon regeneration processes and is critically related to endoplasmic reticulum (ER)-mitochondria interactions. We used real-time polymerase chain reaction, Western blotting, and immunofluorescence to screen a variety of natural compounds, and found that Loureirin B (LrB) effectively promoted neuron polarization and axon regeneration in vitr o and in vivo . LrB significantly inhibited ER stress and thereby promoted mitochondrial functions by regulating mitochondrial fusion. Further, LrB reactivated the Akt/GSK-3β pathway, which plays critical roles in cell survival and microtubule stabilization. Taken together, our results suggest that the effects of LrB on neuron regeneration involve the inhibition of ER stress-induced mitochondrial dysfunction and activation of the Akt/GSK-3β pathway, which further promotes microtubule stabilization. LrB may therefore be a promising candidate for facilitating recovery following SCI.

Proper selection and effective delivery of combination drugs targeting multiple pathophysiological pathways key to spinal cord injury (SCI) hold promise to address the thus far scarce clinical therapeutics for improving recovery after SCI. In this study, we aim to develop a clinically feasible way for targeted delivery of multiple drugs with different physiochemical properties to the SCI site, detail the underlying mechanism of neural recovery, and detect any synergistic effect related to combination therapy. Methods: Liposomes (LIP) modified with a scar-targeted tetrapeptide (cysteine-alanine-glutamine-lysine, CAQK) were first constructed to simultaneously encapsulate docetaxel (DTX) and brain-derived neurotrophic factor (BDNF) and then were further added into a thermosensitive heparin-modified poloxamer hydrogel (HP) with affinity-bound acidic fibroblast growth factor (aFGF-HP) for local administration into the SCI site (CAQK-LIP-GFs/DTX@HP) in a rat model. In vivo fluorescence imaging was used to examine the specificity of CAQK-LIP-GFs/DTX binding to the injured site. Multiple comprehensive evaluations including biotin dextran amine anterograde tracing and magnetic resonance imaging were used to detect any synergistic effects and the underlying mechanisms of CAQK-LIP-GFs/DTX@HP both in vivo (rat SCI model) and in vitro (primary neuron). Results: The multiple drugs were effectively delivered to the injured site. The combined application of GFs and DTX supported neuro-regeneration by improving neuronal survival and plasticity, rendering a more permissive extracellular matrix environment with improved regeneration potential. In addition, our combination therapy promoted axonal regeneration via moderation of microtubule function and mitochondrial transport along the regenerating axon. Conclusion: This novel multifunctional therapeutic strategy with a scar-homing delivery system may offer promising translational prospects for the clinical treatment of SCI., Competing Interests: Competing Interests: The authors have declared that no competing interest exists.

Spinal cord injury (SCI) is a severe nervous system disease that may lead to lifelong disability. Studies have shown that autophagy plays a key role in various diseases; however, the mechanisms regulating cross-talk between autophagy, inflammation, and endoplasmic reticulum (ER) stress during SCI recovery remain unclear. This study was designed to investigate the mechanism by which chloroquine (CQ) inhibits autophagy-associated inflammation and ER stress in rats during their recovery from acute SCI. We evaluated the locomotor function, level of autophagy, and levels of inflammatory cytokines and ER-stress-associated proteins and examined the degradation of the key regulator of inflammation inhibitor of kappa B alpha (I-κBα) through autophagy by analyzing the colocalization of I-κBα, p62, and microtubule-associated protein 1 light chain 3-II. In addition, overexpression of the p62 and activating transcription factor 4 (ATF4) silencing plasmids was used to verify the important roles for autophagic degradation and ER stress. In this study, locomotor function is improved, and autophagy and inflammation are significantly inhibited by, CQ treatment in the model rats. In addition, CQ significantly inhibits the degradation of ubiquitinated I-κBα and blocks the nuclear translocation of nuclear factor kappa B p65 and expression of inflammatory factors. Overexpression of p62 increases I-κBα degradation and improves inflammatory responses. Moreover, CQ treatment also inhibits the activation of ER stress in the rat SCI model, and the ATF4 signaling pathway is required for ER-stress-induced activation of autophagy. These findings reveal a novel mechanism underlying the beneficial effects of CQ on the recovery of SCI, particularly the mechanisms regulating cross-talk between autophagy, inflammation, and ER stress.

Fibroblast growth factor 13 (FGF13), a nonsecretory protein of the FGF family, plays a crucial role in developing cortical neurons by stabilizing the microtubule. In previous studies, we showed that regulation of microtubule dynamics was instrumental for both growth cone initiation and for promoting regrowth of injured axon. However, the expression and effect of FGF13 in spinal cord or after spinal cord injury (SCI) remains undefined. Here, we demonstrated a role of FGF13 in regulating microtubule dynamics and in enhancing axon regeneration after SCI. Administration of FGF13 not only promoted neuronal polarization, axon formation, and growth cone initiation in vitro, but it also facilitated functional recovery following SCI. In addition, we found that upregulation of FGF13 in primary cortical neurons was accompanied by enhanced mitochondrial function, which is essential for axon regeneration. Our study has defined a novel mechanism underlying the beneficial effect of FGF13 on axon regeneration, pointing out that FGF13 may serve as a potential candidate for treating SCI or other central nervous system (CNS) injury.

After spinal cord injury (SCI), the destruction of blood-spinal cord barrier (BSCB) is shown to accelerate gathering of noxious blood-derived components in the nervous system, leading to secondary neurodegenerative damages. SCI activates endoplasmic reticulum stress (ER stress), which is considered to evoke secondary damages of neurons and glia. Recent evidence indicates that Dl-3-n-butylphthalide (NBP) has the neuroprotective effect in ischaemic brain injury, but whether it has protective effects on SCI or not is largely unclear. Here, we show that NBP prevented BSCB disruption after SCI via inhibition of ER stress. Following a moderate contusion injury of the T9 level of spinal cord, NBP was administered by oral gavage and further treated once a day. NBP significantly attenuated BSCB permeability and breakdown of adherens junction (AJ) and tight junction (TJ) proteins, then improved locomotion recovery following SCI. The protective role of NBP on BSCB disruption is associated with the restrain of ER stress caused by SCI. Furthermore, NBP considerably constrained the expression of ER stress-associated proteins and degradation of TJ and AJ in human brain microvascular endothelial cells (HBMECs) treated with TG. In conclusion, our results indicate that ER stress is associated with the disruption of BSCB integrity after injury, NBP attenuates BSCB disruption via inhibiting ER stress and improve functional recovery following SCI., Competing Interests: Competing Interests: The authors have declared that no competing interest exists.

In this study, we examined the neuroprotective effects and anti-inflammatory properties of Dl-3-n-butylphthalide (NBP) in Sprague-Dawley (SD) rats following traumatic spinal cord injury (SCI) as well as microglia activation and inflammatory response both in vivo and in vitro. Our results showed that NBP improved the locomotor recovery of SD rats after SCI an significantly diminished the lesion cavity area of the spinal cord, apoptotic activity in neurons, and the number of TUNEL-positive cells at 7 days post-injury. NBP inhibited activation of microglia, diminished the release of inflammatory mediators, and reduced the upregulation of microglial TLR4/NF-κB expression at 1 day post-injury. In a co-culture system with BV-2 cells and PC12 cells, NBP significantly reduced the cytotoxicity of BV-2 cells following lipopolysaccharide (LPS) stimulation. In addition, NBP reduced the activation of BV-2 cells, diminished the release of inflammatory mediators, and inhibited microglial TLR4/NF-κB expression in BV-2 cells. Our findings demonstrate that NBP may have neuroprotective and anti-inflammatory properties in the treatment of SCI by inhibiting the activation of microglia via TLR4/NF-κB signalling., (© 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

Therapeutics used to treat central nervous system (CNS) injury were designed to repair neurites and inhibit cell apoptosis. Previous studies have shown that neuron-derived FGF10 exerts potential neuroprotective effects after cerebral ischemia injury. However, little is known about the role of endogenous FGF10 in the recovery process after spinal cord injury (SCI). In this study, we found that FGF10 is mainly produced by neuron and microglia/macrophages, and its expression is increased after SCI. Exogenous treatment of FGF10 improved functional recovery after injury by reducing apoptosis, as well as repairing neurites via FGFR2/PI3K/Akt pathway. On another hand, inhibiting the PI3K/Akt pathway with LY294002 partially reversed the therapeutic effects of FGF10. In addition, small interfering RNA knockdown of FGFR2 suppressed PI3K/Akt pathway activation by FGF10 and abolished its anti-apoptotic and neurite repair effects in vitro. Furthermore, FGF10 treatment inhibited the activation and proliferation of microglia/macrophages through regulation of TLR4/NF-κB pathway, and attenuated the release of pro-inflammatory cytokines after SCI. Thus, the increased expression of FGF10 after acute SCI is an endogenous self-protective response, suggesting that FGF10 could be a potential treatment for CNS injury.

Acidic fibroblast growth factor (aFGF) exerts a protective effect on spinal cord injury (SCI) but is limited by the lack of physicochemical stability and the ability to cross the blood spinal cord barrier (BSCB). As promising biomaterials, hydrogels contain substantial amounts of water and a three-dimensional porous structure and are commonly used to load and deliver growth factors. Heparin can not only enhance growth factor loading onto hydrogels but also can stabilize the structure and control the release behavior. Herein, a novel aFGF-loaded thermosensitive heparin-poloxamer (aFGF-HP) hydrogel was developed and applied to provide protection and regeneration after SCI. To assess the effects of the aFGF-HP hydrogel, BSCB restoration, neuron and axonal rehabilitation, glial scar inhibition, inflammatory response suppression, and motor recovery were studied both in vivo and in vitro. The aFGF-HP hydrogels exhibited sustained release of aFGF and protected the bioactivity of aFGF in vitro. Compared to groups intravenously administered either drug-free HP hydrogel or aFGF alone, the aFGF-HP hydrogel group revealed prominent and attenuated disruption of the BSCB, reduced neuronal apoptosis, reactive astrogliosis, and increased neuron and axonal rehabilitation both in vivo and in vitro. This work provides an effective approach to enhance recovery after SCI and provide a successful strategy for SCI protection.

Spinal cord injury induces the disruption of blood-spinal cord barrier and triggers a complex array of tissue responses, including endoplasmic reticulum (ER) stress and autophagy. However, the roles of ER stress and autophagy in blood-spinal cord barrier disruption have not been discussed in acute spinal cord trauma. In the present study, we respectively detected the roles of ER stress and autophagy in blood-spinal cord barrier disruption after spinal cord injury. Besides, we also detected the cross-talking between autophagy and ER stress both in vivo and in vitro. ER stress inhibitor, 4-phenylbutyric acid, and autophagy inhibitor, chloroquine, were respectively or combinedly administrated in the model of acute spinal cord injury rats. At day 1 after spinal cord injury, blood-spinal cord barrier was disrupted and activation of ER stress and autophagy were involved in the rat model of trauma. Inhibition of ER stress by treating with 4-phenylbutyric acid decreased blood-spinal cord barrier permeability, prevented the loss of tight junction (TJ) proteins and reduced autophagy activation after spinal cord injury. On the contrary, inhibition of autophagy by treating with chloroquine exacerbated blood-spinal cord barrier permeability, promoted the loss of TJ proteins and enhanced ER stress after spinal cord injury. When 4-phenylbutyric acid and chloroquine were combinedly administrated in spinal cord injury rats, chloroquine abolished the blood-spinal cord barrier protective effect of 4-phenylbutyric acid by exacerbating ER stress after spinal cord injury, indicating that the cross-talking between autophagy and ER stress may play a central role on blood-spinal cord barrier integrity in acute spinal cord injury. The present study illustrates that ER stress induced by spinal cord injury plays a detrimental role on blood-spinal cord barrier integrity, on the contrary, autophagy induced by spinal cord injury plays a furthersome role in blood-spinal cord barrier integrity in acute spinal cord injury.

After spinal cord injury (SCI), disruption of blood-spinal cord barrier (BSCB) elicits blood cell infiltration such as neutrophils and macrophages, contributing to permanent neurological disability. Previous studies show that epidermal growth factor (EGF) produces potent neuroprotective effects in SCI models. However, little is known that whether EGF contributes to the integrity of BSCB. The present study is performed to explore the mechanism of BSCB permeability changes which are induced by EGF treatment after SCI in rats. In this study, we demonstrate that EGF administration inhibits the disruption of BSCB permeability and improves the locomotor activity in SCI model rats. Inhibition of the PI3K/Akt pathways by a specific inhibitor, LY294002, suppresses EGF-induced Rac1 activation as well as tight junction (TJ) and adherens junction (AJ) expression. Furthermore, the protective effect of EGF on BSCB is related to the activation of Rac1 both in vivo and in vitro. Blockade of Rac1 activation with Rac1 siRNA downregulates EGF-induced TJ and AJ proteins expression in endothelial cells. Taken together, our results indicate that EGF treatment preserves BSCB integrity and improves functional recovery after SCI via PI3K-Akt-Rac1 signalling pathway., (© 2016 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

Spinal cord injury (SCI) induces the disruption of the blood-spinal cord barrier (BSCB), which leads to infiltration of blood cells, inflammatory responses and neuronal cell death, with subsequent development of spinal cord secondary damage. Recent reports pointed to an important role of retinoic acid (RA), the active metabolite of the vitamin A, in the induction of the blood-brain barrier (BBB) during human and mouse development, however, it is unknown whether RA plays a role in maintaining BSCB integrity under the pathological conditions such as SCI. In this study, we investigated the BSCB protective role of RA both in vivo and in vitro and demonstrated that autophagy are involved in the BSCB protective effect of RA. Our data show that RA attenuated BSCB permeability and also attenuated the loss of tight junction molecules such as P120, β-catenin, Occludin and Claudin5 after injury in vivo as well as in brain microvascular endothelial cells. In addition, RA administration improved functional recovery of the rat model of trauma. We also found that RA could significantly increase the expression of LC3-II and decrease the expression of p62 both in vivo and in vitro. Furthermore, combining RA with the autophagy inhibitor chloroquine (CQ) partially abolished its protective effect on the BSCB and exacerbated the loss of tight junctions. Together, our studies indicate that RA improved functional recovery in part by the prevention of BSCB disruption via the activation of autophagic flux after SCI.

Spinal cord injury (SCI) induces the disruption of the blood-spinal cord barrier (BSCB) which leads to infiltration of blood cells, an inflammatory response, and neuronal cell death, resulting spinal cord secondary damage. Retinoic acid (RA) has a neuroprotective effect in both ischemic brain injury and SCI, however the relationship between BSCB disruption and RA in SCI is still unclear. In this study, we demonstrated that autophagy and ER stress are involved in the protective effect of RA on the BSCB. RA attenuated BSCB permeability and decreased the loss of tight junction (TJ) molecules such as P120, β-catenin, Occludin and Claudin5 after injury in vivo as well as in Brain Microvascular Endothelial Cells (BMECs). Moreover, RA administration improved functional recovery in the rat model of SCI. RA inhibited the expression of CHOP and caspase-12 by induction of autophagic flux. However, RA had no significant effect on protein expression of GRP78 and PDI. Furthermore, combining RA with the autophagy inhibitor chloroquine (CQ) partially abolished its protective effect on the BSCB via exacerbated ER stress and subsequent loss of tight junctions. Taken together, the neuroprotective role of RA in recovery from SCI is related to prevention of of BSCB disruption via the activation of autophagic flux and the inhibition of ER stress-induced cell apoptosis. These findings lay the groundwork for future translational studies of RA for CNS diseases, especially those related to BSCB disruption.

Background: Endoplasmic reticulum (ER) stress-induced apoptosis plays a major role in various diseases, including spinal cord injury (SCI). Nerve growth factor (NGF) show neuroprotective effect and improve the recovery of SCI, but the relations of ER stress-induced apoptosis and the NGF therapeutic effect in SCI still unclear., Methods: Young adult female Sprague-Dawley rats's vertebral column was exposed and a laminectomy was done at T9 vertebrae and moderate contusion injuries were performed using a vascular clip. NGF stock solution was diluted with 0.9% NaCl and administered intravenously at a dose of 20 μg/kg/day after SCI and then once per day until they were executed. Subsequently, the rats were executed at 1d, 3 d, 7d and 14d. The locomotor activities of SCI model rats were tested by the 21-point Basso-Beattie-Bresnahan (BBB) locomotion scale, inclined plane test and footprint analysis. In addition, Western blot analysis was performed to identify the expression of ER-stress related proteins including CHOP, GRP78 and caspase-12 both in vivo and in vitro. The level of cell apoptosis was determined by TUNEL in vivo and Flow cytometry in vitro. Relative downstream signals Akt/GSK-3β and ERK1/2were also analyzed with or without inhibitors in vitro., Results: Our results demonstrated that ER stress-induced apoptosis was involved in the injury of SCI model rats. NGF administration improved the motor function recovery and increased the neurons survival in the spinal cord lesions of the model rats. NGF decreases neuron apoptosis which measured by TUNEL and inhibits the activation of caspase-3 cascade. The ER stress-induced apoptosis response proteins CHOP, GRP78 and caspase-12 are inhibited by NGF treatment. Meanwhile, NGF administration also increased expression of growth-associated protein 43 (GAP43). The administration of NGF activated downstream signals Akt/GSK-3β and ERK1/2 in ER stress cell model in vitro., Conclusion: The neuroprotective role of NGF in the recovery of SCI is related to the inhibition of ER stress-induced cell death via the activation of downstream signals, also suggested a new trend of NGF translational drug development in the central neural system injuries which involved in the regulation of chronic ER stress.

Promoting blood–spinal cord barrier (BSCB) repair at the early stage plays a crucial role in treatment of spinal cord injury (SCI). Excessive activation of autophagy can prevent recovery of BSCB after SCI. Basic fibroblast growth factor (bFGF) has been shown to promote BSCB repair and locomotor function recovery in SCI. However, the therapeutic effect of bFGF via direct administration on SCI is limited because of its rapid degradation and dilution at injury site. Based on these considerations, controlled release of bFGF in the lesion area is becoming an attractive strategy for SCI repair. At present, we have designed a sustained-release system of bFGF (called ALG-bFGF) using sodium alginate hydrogel, which is able to load large amounts of bFGF and suitable for in situ administration of bFGF in vivo. Here, traumatic SCI mice models and oxygen glucose deprivation (OGD)–stimulated human brain microvascular endothelial cells were performed to explore the effects and the underlying mechanisms of ALG-bFGF in promoting SCI repair. After a single in situ injection of ALG-bFGF hydrogel into the injured spinal cord, sustained release of bFGF from ALG hydrogel distinctly prevented BSCB destruction and improved motor functional recovery in mice after SCI, which showed better therapeutic effect than those in mice treated with bFGF solution or ALG. Evidences have demonstrated that autophagy is involved in maintaining BSCB integrity and functional restoration in animals after SCI. In this study, SCI/OGD exposure–induced significant upregulations of autophagy activation-related proteins (Beclin1, ATG5, LC3II/I) were distinctly decreased by ALG-bFGF hydrogel near the baseline and not less than it both in vivo and in vitro , and this inhibitory effect contributed to prevent BSCB destruction. Finally, PI3K inhibitor LY294002 and KLF4 inhibitor NSC-664704 were applied to further explore the underlying mechanism by which ALG-bFGF attenuated autophagy activation to alleviate BSCB destruction after SCI. The results further indicated that ALG-bFGF hydrogel maintaining BSCB integrity by inhibiting autophagy activation was regulated by PI3K/Akt/FOXO1/KLF4 pathway. In summary, our current study revealed a novel mechanism by which ALG-bFGF hydrogel improves BSCB and motor function recovery after SCI, providing an effective therapeutic strategy for SCI repair. [ABSTRACT FROM AUTHOR]

An amendment to this paper has been published and can be accessed via the original article. [ABSTRACT FROM AUTHOR]

This article has been retracted. Please see the Retraction Notice for more detail: https://doi.org/10.1007/s11064-020-03149-1. [ABSTRACT FROM AUTHOR]

• A three-dimensional scaffold hydrogel (SLIP-bFGF/DTX@SF) were synthesized. • The hydrogel delivery bFGF and DTX for spinal cord injury treatment. • The hydrogel prolonged drug half-life at the injured spinal cord area, bridged cysts by remodeling ECM. • The hydrogel promoted microtubule stabilization and enhanced endogeneous axonal regeneration. Spinal cord injury (SCI) remains a challenging clinical problem worldwide. Combination therapy is a promising therapeutic strategy for SCI. Recent investigations have reported that docetaxel (DTX) facilitates intrinsic axonal regeneration, and basic fibroblast growth factor (bFGF) regulates neuronal survival, plasticity, and extracellular matrix (ECM) after SCI. We constructed a novel biocompatible liposome (LIP) with a silk fibroin (SF) hydrogel core (SLIP) to simultaneously deliver bFGF and DTX, which have different physiochemical characteristics. Subsequently, SLIP-bFGF/DTX was added to the SF hydrogel to form a three-dimensional scaffold gel (SLIP-bFGF/DTX@SF). We evaluated the pharmacological properties of SLIP@SF and investigated histological and functional outcomes. A SLIP-bFGF/DTX@SF treatment effectively prolonged the drug half-life in the injured spinal cord area, bridged cysts by remodeling the extracellular matrix (ECM), and improved the motor function of rats with SCI. Additionally, this hydrogel demonstrated a strong affinity for the polysaccharides enriched in the lesion area providing a unique therapeutic approach for the treatment of SCI. Furthermore, the mechanisms of inhibition of microtubule dynamics and reshaping of the ECM by the drugs were demonstrated to reflected the pathological features of SCI. This novel multidrug delivery system achieves a significant functional improvement a simultaneously addresses multiple key aspects of SCI pathology indicating favorable clinical translational prospects. [ABSTRACT FROM AUTHOR]