Axon Extension Research Articles

Pip5k1γ regulates axon formation by limiting Rap1 activity.

During their differentiation, neurons establish a highly polarized morphology by forming axons and dendrites. Cortical and hippocampal neurons initially extend several short neurites that all have the potential to become an axon. One of these neurites is then selected as the axon by a combination of positive and negative feedback signals that promote axon formation and prevent the remaining neurites from developing into axons. Here, we show that Pip5k1γ is required for the formation of a single axon as a negative feedback signal that regulates C3G and Rap1 through the generation of phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2). Impairing the function of Pip5k1γ results in a hyper-activation of the Fyn/C3G/Rap1 pathway, which induces the formation of supernumerary axons. Application of a hyper-osmotic shock to modulate membrane tension has a similar effect, increasing Rap1 activity and inducing the formation of supernumerary axons. In both cases, the induction of supernumerary axons can be reverted by expressing constitutively active Pip5k. Our results show that PI(4,5)P2-dependent membrane properties limit the activity of C3G and Rap1 to ensure the extension of a single axon.

Distinct transcriptomic profile of satellite cells contributes to preservation of neuromuscular junctions in extraocular muscles of ALS mice.

Amyotrophic lateral sclerosis (ALS) is a fatal neuromuscular disorder characterized by progressive weakness of almost all skeletal muscles, whereas extraocular muscles (EOMs) are comparatively spared. While hindlimb and diaphragm muscles of end-stage SOD1G93A (G93A) mice (a familial ALS mouse model) exhibit severe denervation and depletion of Pax7+satellite cells (SCs), we found that the pool of SCs and the integrity of neuromuscular junctions (NMJs) are maintained in EOMs. In cell sorting profiles, SCs derived from hindlimb and diaphragm muscles of G93A mice exhibit denervation-related activation, whereas SCs from EOMs of G93A mice display spontaneous (non-denervation-related) activation, similar to SCs from wild-type mice. Specifically, cultured EOM SCs contain more abundant transcripts of axon guidance molecules, including Cxcl12, along with more sustainable renewability than the diaphragm and hindlimb counterparts under differentiation pressure. In neuromuscular co-culture assays, AAV-delivery of Cxcl12 to G93A-hindlimb SC-derived myotubes enhances motor neuron axon extension and innervation, recapitulating the innervation capacity of EOM SC-derived myotubes. G93A mice fed with sodium butyrate (NaBu) supplementation exhibited less NMJ loss in hindlimb and diaphragm muscles. Additionally, SCs derived from G93A hindlimb and diaphragm muscles displayed elevated expression of Cxcl12 and improved renewability following NaBu treatment in vitro. Thus, the NaBu-induced transcriptomic changes resembling the patterns of EOM SCs may contribute to the beneficial effects observed in G93A mice. More broadly, the distinct transcriptomic profile of EOM SCs may offer novel therapeutic targets to slow progressive neuromuscular functional decay in ALS and provide possible 'response biomarkers' in pre-clinical and clinical studies.

Distinct transcriptomic profile of satellite cells contributes to preservation of neuromuscular junctions in extraocular muscles of ALS mice

Amyotrophic lateral sclerosis (ALS) is a fatal neuromuscular disorder characterized by progressive weakness of almost all skeletal muscles, whereas extraocular muscles (EOMs) are comparatively spared. While hindlimb and diaphragm muscles of end-stage SOD1G93A (G93A) mice (a familial ALS mouse model) exhibit severe denervation and depletion of Pax7+satellite cells (SCs), we found that the pool of SCs and the integrity of neuromuscular junctions (NMJs) are maintained in EOMs. In cell sorting profiles, SCs derived from hindlimb and diaphragm muscles of G93A mice exhibit denervation-related activation, whereas SCs from EOMs of G93A mice display spontaneous (non-denervation-related) activation, similar to SCs from wild-type mice. Specifically, cultured EOM SCs contain more abundant transcripts of axon guidance molecules, including Cxcl12, along with more sustainable renewability than the diaphragm and hindlimb counterparts under differentiation pressure. In neuromuscular co-culture assays, AAV-delivery of Cxcl12 to G93A-hindlimb SC-derived myotubes enhances motor neuron axon extension and innervation, recapitulating the innervation capacity of EOM SC-derived myotubes. G93A mice fed with sodium butyrate (NaBu) supplementation exhibited less NMJ loss in hindlimb and diaphragm muscles. Additionally, SCs derived from G93A hindlimb and diaphragm muscles displayed elevated expression of Cxcl12 and improved renewability following NaBu treatment in vitro. Thus, the NaBu-induced transcriptomic changes resembling the patterns of EOM SCs may contribute to the beneficial effects observed in G93A mice. More broadly, the distinct transcriptomic profile of EOM SCs may offer novel therapeutic targets to slow progressive neuromuscular functional decay in ALS and provide possible ‘response biomarkers’ in pre-clinical and clinical studies.

Fabrication of Artificial Nerve Conduits Used in a Long Nerve Gap: Current Reviews and Future Studies.

There are many commercially available artificial nerve conduits, used mostly to repair short gaps in sensory nerves. The stages of nerve regeneration in a nerve conduit are fibrin matrix formation between the nerve stumps joined to the conduit, capillary extension and Schwann cell migration from both nerve stumps, and, finally, axon extension from the proximal nerve stump. Artificial nerves connecting transected nerve stumps with a long interstump gap should be biodegradable, soft and pliable; have the ability to maintain an intrachamber fibrin matrix structure that allows capillary invasion of the tubular lumen, inhibition of scar tissue invasion and leakage of intratubular neurochemical factors from the chamber; and be able to accommodate cells that produce neurochemical factors that promote nerve regeneration. Here, we describe current progress in the development of artificial nerve conduits and the future studies needed to create nerve conduits, the nerve regeneration of which is compatible with that of an autologous nerve graft transplanted over a long nerve gap.

Constructing Linear-Oriented Pre-Vascularized Human Spinal Cord Tissues for Spinal Cord Injury Repair.

Repairing spinal cord injury (SCI) is a global medical challenge lacking effective clinical treatment. Developing human-engineered spinal cord tissues that can replenish lost cells and restore a regenerative microenvironment offers promising potential for SCI therapy. However, creating vascularized human spinal cord-like tissues (VSCT) that mimic the diverse cell types and longitudinal parallel structural features of spinal cord tissues remains a significant hurdle. In the present study, VSCTs are engineered using embryonic human spinal cord-derived neural and endothelial cells on linear-ordered collagen scaffolds (LOCS). Studies have shown that astrocytes and endothelial cells align along the scaffolds in VSCT, supporting axon extension from various human neurons myelinated by oligodendrocytes. After transplantation into SCI rats, VSCT survives at the injury sites and promotes endogenous neural regeneration and vascularization, ultimately reducing scarring and enhancing behavioral functional recovery. It suggests that pre-vascularization of engineered spinal cord tissues is beneficial for SCI treatment and highlights the important role of exogenous endothelial cells in tissue engineering.

Repulsive Sema3E-Plexin-D1 signaling coordinates both axonal extension and steering via activating an autoregulatory factor, Mtss1.

Axon guidance molecules are critical for neuronal pathfinding because they regulate directionality and growth pace during nervous system development. However, the molecular mechanisms coordinating proper axonal extension and turning are poorly understood. Here, metastasis suppressor 1 (Mtss1), a membrane protrusion protein, ensured axonal extension while sensitizing axons to the Semaphorin 3E (Sema3E)-Plexin-D1 repulsive cue. Sema3E-Plexin-D1 signaling enhanced Mtss1 expression in projecting striatonigral neurons. Mtss1 localized to the neurite axonal side and regulated neurite outgrowth in cultured neurons. Mtss1 also aided Plexin-D1 trafficking to the growth cone, where it signaled a repulsive cue to Sema3E. Mtss1 ablation reduced neurite extension and growth cone collapse in cultured neurons. Mtss1-knockout mice exhibited fewer striatonigral projections and irregular axonal routes, and these defects were recapitulated in Plxnd1- or Sema3e-knockout mice. These findings demonstrate that repulsive axon guidance activates an exquisite autoregulatory program coordinating both axonal extension and steering during neuronal pathfinding.

Generation of contractile forces by three-dimensional bundled axonal tracts in micro-tissue engineered neural networks.

Axonal extension and retraction are ongoing processes that occur throughout all developmental stages of an organism. The ability of axons to produce mechanical forces internally and respond to externally generated forces is crucial for nervous system development, maintenance, and plasticity. Such axonal mechanobiological phenomena have typically been evaluated in vitro at a single-cell level, but these mechanisms have not been studied when axons are present in a bundled three-dimensional (3D) form like in native tissue. In an attempt to emulate native cortico-cortical interactions under in vitro conditions, we present our approach to utilize previously described micro-tissue engineered neural networks (micro-TENNs). Here, micro-TENNs were comprised of discrete populations of rat cortical neurons that were spanned by 3D bundled axonal tracts and physically integrated with each other. We found that these bundled axonal tracts inherently exhibited an ability to generate contractile forces as the microtissue matured. We therefore utilized this micro-TENN testbed to characterize the intrinsic contractile forces generated by the integrated axonal tracts in the absence of any external force. We found that contractile forces generated by bundled axons were dependent on microtubule stability. Moreover, these intra-axonal contractile forces could simultaneously generate tensile forces to induce so-called axonal "stretch-growth" in different axonal tracts within the same microtissue. The culmination of axonal contraction generally occurred with the fusion of both the neuronal somatic regions along the axonal tracts, therefore perhaps showing the innate tendency of cortical neurons to minimize their wiring distance, a phenomenon also perceived during brain morphogenesis. In future applications, this testbed may be used to investigate mechanisms of neuroanatomical development and those underlying certain neurodevelopmental disorders.

Clickable Granular Hydrogel Scaffolds for Delivery of Neural Progenitor Cells to Sites of Spinal Cord Injury.

Spinal cord injury (SCI) is a serious condition with limited treatment options. Neural progenitor cell (NPC) transplantation is a promising treatment option, and the identification of novel biomaterial scaffolds that support NPC engraftment and therapeutic activity is a top research priority. The objective of this study is to evaluate in situ assembled poly (ethylene glycol) (PEG)-based granular hydrogels for NPC delivery in a murine model of SCI. Microgel precursors are synthesized by using thiol-norbornene click chemistry to react four-armed PEG-amide-norbornene with enzymatically degradable and cell adhesive peptides. Unreacted norbornene groups are utilized for in situ assembly into scaffolds using a PEG-di-tetrazine linker. The granular hydrogel scaffolds exhibit good biocompatibility and do not adversely affect the inflammatory response after SCI. Moreover, when used to deliver NPCs, the granular hydrogel scaffolds supported NPC engraftment, do not adversely affect the immune response to the NPC grafts, and successfully support graft differentiation toward neuronal or astrocytic lineages as well as axonal extension into the host tissue. Collectively, these data establish PEG-based granular hydrogel scaffolds as a suitable biomaterial platform for NPC delivery and justify further testing, particularly in the context of more severe SCI.

Role of SIRT1 in Potentially Toxic Trace Elements (Lead, Fluoride, Aluminum and Cadmium) Associated Neurodevelopmental Toxicity.

The formation of the central nervous system is a meticulously planned and intricate process. Any modification to this process has the potential to disrupt the structure and operation of the brain, which could result in deficiencies in neurological growth. When neurotoxic substances are present during the early stages of development, they can be exceptionally dangerous. Prenatally, the immature brain is extremely vulnerable and is therefore at high risk in pregnant women associated with occupational exposures. Lead, fluoride, aluminum, and cadmium are examples of possibly toxic trace elements that have been identified as an environmental concern in the aetiology of a number of neurological and neurodegenerative illnesses. SIRT1, a member of the sirtuin family has received most attention for its potential neuroprotective properties. SIRT1 is an intriguing therapeutic target since it demonstrates important functions to increase neurogenesis and cellular lifespan by modulating multiple pathways. It promotes axonal extension, neurite growth, and dendritic branching during the development of neurons. Additionally, it contributes to neurogenesis, synaptic plasticity, memory development, and neuroprotection. This review summarizes the possible role of SIRT1 signalling pathway in potentially toxic trace elements -induced neurodevelopmental toxicity, highlighting some molecular pathways such as mitochondrial biogenesis, CREB/BDNF and PGC-1α/NRF1/TFAM.

Cell adhesion and actin dynamics factors promote axonal extension and synapse formation in transplanted Drosophila photoreceptor cells.

Vision is formed by the transmission of light stimuli to the brain through axons extending from photoreceptor cells. Damage to these axons leads to loss of vision. Despite research on neural circuit regeneration through transplantation, achieving precise axon projection remains challenging. To achieve optic nerve regeneration by transplantation, we employed the Drosophila visual system. We previously established a transplantation method for Drosophila utilizing photoreceptor precursor cells extracted from the eye disc. However, little axonal elongation of transplanted cells into the brain, the lamina, was observed. We verified axonal elongation to the lamina by modifying the selection process for transplanted cells. Moreover, we focused on N-cadherin (Ncad), a cell adhesion factor, and Twinstar (Tsr), which has been shown to promote actin reorganization and induce axon elongation in damaged nerves. Overexpression of Ncad and tsr promoted axon elongation to the lamina, along with presynaptic structure formation in the elongating axons. Furthermore, overexpression of Neurexin-1 (Nrx-1), encoding a protein identified as a synaptic organizer, was found to not only promote presynapse formation but also enhance axon elongation. By introducing Ncad, tsr, and Nrx-1, we not only successfully achieved axonal projection of transplanted cells to the brain beyond the retina, but also confirmed the projection of transplanted cells into a deeper ganglion, the medulla. The present study offers valuable insights to realize regeneration through transplantation in a more complex nervous system.

The secretome of macrophages has a differential impact on spinal cord injury recovery according to the polarization protocol.

The inflammatory response after spinal cord injury (SCI) is an important contributor to secondary damage. Infiltrating macrophages can acquire a spectrum of activation states, however, the microenvironment at the SCI site favors macrophage polarization into a pro-inflammatory phenotype, which is one of the reasons why macrophage transplantation has failed. In this study, we investigated the therapeutic potential of the macrophage secretome for SCI recovery. We investigated the effect of the secretome in vitro using peripheral and CNS-derived neurons and human neural stem cells. Moreover, we perform a pre-clinical trial using a SCI compression mice model and analyzed the recovery of motor, sensory and autonomic functions. Instead of transplanting the cells, we injected the paracrine factors and extracellular vesicles that they secrete, avoiding the loss of the phenotype of the transplanted cells due to local environmental cues. We demonstrated that different macrophage phenotypes have a distinct effect on neuronal growth and survival, namely, the alternative activation with IL-10 and TGF-β1 (M(IL-10+TGF-β1)) promotes significant axonal regeneration. We also observed that systemic injection of soluble factors and extracellular vesicles derived from M(IL-10+TGF-β1) macrophages promotes significant functional recovery after compressive SCI and leads to higher survival of spinal cord neurons. Additionally, the M(IL-10+TGF-β1) secretome supported the recovery of bladder function and decreased microglial activation, astrogliosis and fibrotic scar in the spinal cord. Proteomic analysis of the M(IL-10+TGF-β1)-derived secretome identified clusters of proteins involved in axon extension, dendritic spine maintenance, cell polarity establishment, and regulation of astrocytic activation. Overall, our results demonstrated that macrophages-derived soluble factors and extracellular vesicles might be a promising therapy for SCI with possible clinical applications.

A materiobiology-inspired sericin nerve guidance conduit extensively activates regeneration-associated genes of Schwann cells for long-gap peripheral nerve repair

Peripheral nerve injury (PNI) is one of the most common causes of adult disability. Following PNI, Schwann cells (SCs) dedifferentiate, proliferate, and upregulate a variety of regeneration-associated genes (RAGs), including neurotrophic factors, ultimately promoting nerve regeneration. Even though certain reported biomaterials could upregulate one or several neurotrophic factors of SCs, none of them was able to activate these RAGs extensively. Herein, this work represents a novel manganese organic framework material (Mn-MOF) capable of effectively upregulating RAGs of SCs. In vitro cell coculture assay revealed that these Mn-MOF treated SCs enhanced the axon extension of PC12 cells, further confirming the axon regeneration-promoting activity of this framework. STAT3 and C-JUN signaling pathways were involved in Mn-MOF-induced RAGs activation. Using sericin, a well-proved biomaterial with neuroprotective properties, a Mn-MOF modified sericin conduit (MOF-SS) was prepared via π-π interactions between the cyclobenzene of framework materials and the aromatic acid of natural proteins. The microstructures and physiochemical properties of MOF-SS were comprehensively characterized, and its RAGs-activating effect on SCs was well confirmed. When applied in vivo to bridge 13-mm sciatic nerve defect, this conduit significantly promoted nerve microstructural regeneration, enhanced electrical conduction performance of regenerated nerve, improved functional recovery, and more importantly, alleviated muscle atrophy. In summary, this work represents a novel Mn-MOF modified sericin conduit capable of extensively activating RAGs of SCs, significantly promoting long-gap PNI repair.

Analysis of Neuronal Morphology by Two-Photon Microscopy.

During the development of mammalian brains, pyramidal neurons in the cerebral cortex form highly organized six layers with different functions. These neurons undergo developmental processes such as axon extension, dendrite outgrowth, and synapse formation. A proper integration of the neuronal connectivity through dynamic changes of dendritic branches and spines is required for learning and memory. Disruption of these crucial developmental processes is associated with many neurodevelopmental and neurodegenerative disorders. To investigate the complex dendritic architecture, several useful staining tools and genetic methods to label neurons have been well established. Monitoring the dynamics of dendritic spine in a single neuron is still a challenging task. Here, we provide a methodology that combines in vivo two-photon brain imaging and in utero electroporation, which sparsely labels cortical neurons with fluorescent proteins. This protocol may help elucidate the dynamics of microstructure and neural complexity in living rodents under normal and disease conditions.

Exosomes derived from CD271+CD56+ bone marrow mesenchymal stem cell subpopoulation identified by single-cell RNA sequencing promote axon regeneration after spinal cord injury.

Rationale: Spinal cord injury (SCI) results in neural tissue damage. However, the limited regenerative capacity of adult mammals' axons upon SCI leads to persistent neurological dysfunction. Thus, exploring the pathways that can enhance axon regeneration in injured spinal cord is of great significance. Methods: Through the utilization of single-cell RNA sequencing in this research, a distinct subpopulation of bone marrow mesenchymal stem cells (BMSCs) that exhibits the capacity to facilitate axon regeneration has been discovered. Subsequently, the CD271+CD56+ BMSCs subpopulation was isolated using flow cytometry, and the exosomes derived from this subpopulation (CD271+CD56+ BMSC-Exos) were extracted and incorporated into a hydrogel to create a sustained release system. The aim was to investigate the therapeutic effects of CD271+CD56+ BMSC-Exos and elucidate the underlying mechanisms involved in promoting axon regeneration and neural function recovery. Results: The findings indicate that CD271+CD56+ BMSC-Exos share similar physical and chemical properties with conventional exosomes. Importantly, in an SCI model, in situ implantation of CD271+CD56+ BMSC-Exos hydrogel resulted in increased expression of NF and synaptophysin, markers associated with axon regeneration and synapse formation, respectively. This intervention also contributed to improved neural function recovery. In vitro experiments demonstrated that CD271+CD56+ BMSC-Exos treatment significantly enhanced axon extension distance and increased the number of branches in dorsal root ganglion axons. Moreover, further investigation into the molecular mechanisms underlying CD271+CD56+ BMSC-Exos-mediated axon regeneration revealed the crucial involvement of the miR-431-3p/RGMA axis. Conclusion: In summary, the implantation of CD271+CD56+ BMSC-Exos hydrogel presents a promising and effective therapeutic approach for SCI.

Mimicked Spinal Cord Fibers Trigger Axonal Regeneration and Remyelination after Injury.

Spinal cord injury (SCI) causes tissue structure damage and composition changes of the neural parenchyma, resulting in severe consequences for spinal cord function. Mimicking the components and microstructure of spinal cord tissues holds promise for restoring the regenerative microenvironment after SCI. Here, we have utilized electrospinning technology to develop aligned decellularized spinal cord fibers (A-DSCF) without requiring synthetic polymers or organic solvents. A-DSCF preserves multiple types of spinal cord extracellular matrix proteins and forms a parallel-oriented structure. Compared to aligned collagen fibers (A-CF), A-DSCF exhibits stronger mechanical properties, improved enzymatic stability, and superior functionality in the adhesion, proliferation, axonal extension, and myelination of differentiated neural progenitor cells (NPCs). Notably, axon extension or myelination has been primarily linked to Agrin (AGRN), Laminin (LN), or Collagen type IV (COL IV) proteins in A-DSCF. When transplanted into rats with complete SCI, A-DSCF loaded with NPCs improves the survival, maturation, axon regeneration, and motor function of the SCI rats. These findings highlight the potential of structurally and compositionally biomimetic scaffolds to promote axonal extension and remyelination after SCI.

A scaffold containing zinc oxide for Schwann cell-mediated axon growth

Objective. Schwann cells (SCs) transplanted in damaged nervous tissue promote axon growth, which may support the recovery of function lost after injury. However, SC transplant-mediated axon growth is often limited and lacks direction. Approach. We have developed a zinc oxide (ZnO) containing fibrous scaffold consisting of aligned fibers of polycaprolactone (PCL) with embedded ZnO nanoparticles as a biodegradable, bifunctional scaffold for promoting and guiding axon growth. This scaffold has bifunctional properties wherein zinc is released providing bioactivity and ZnO has well-known piezoelectric properties where piezoelectric materials generate electrical activity in response to minute deformations. In this study, SC growth, SC-mediated axon extension, and the presence of myelin basic protein (MBP), as an indicator of myelination, were evaluated on the scaffolds containing varying concentrations of ZnO in vitro. SCs and dorsal root ganglion (DRG) neurons were cultured, either alone or in co-culture, on the scaffolds. Main results. Findings demonstrated that scaffolds with 1 wt.% ZnO promoted the greatest SC growth and SC-mediated axon extension. The presence of brain-derived neurotrophic factor (BDNF) was also determined. BDNF increased in co-cultures for all scaffolds as compared to SCs or DRGs cultured alone on all scaffolds. For co-cultures, cells on scaffolds with low levels of ZnO (0.5 wt.% ZnO) had the highest amount of BDNF as compared to cells on higher ZnO-containing scaffolds (1 and 2 wt.%). MBP immunostaining was only detected in co-cultures on PCL control scaffolds (without ZnO). Significance. The results of this study demonstrate the potential of the ZnO-containing scaffolds for SC-mediated axon growth and its potential for use in nervous tissue repair.

An electroconductive hydrogel with injectable and self-healing properties accelerates peripheral nerve regeneration and motor functional recovery

Peripheral nerve regeneration and functional recovery are slow due to disruption of bioelectrical signal transmission. As a new therapeutic strategy, conductive hydrogels have been shown to promote the regeneration of electroactive tissue, especially nerve tissue. However, conventional conductive hydrogels made of physically doped conductive particles suffer from fragility, agglomeration, poor durability, and discontinuous transmission of electrical signals, severely limiting the migration of Schwann cells (SCs) and the remyelination of axons. Herein, a novel water-soluble conductive material was synthesized by grafting polyaniline (PANI) onto carboxymethyl chitosan (CMCS). Dual cross-linked hydrogels with injectable, self-healing, and conductive properties were created by adding CMCS-PANI (CP) to the dynamic gel network generated by the Schiff base reaction between aldehyde-based hyaluronic acid (ALHA) and CMCS. The prepared self-healing ALHA/CMCS/CP (ACCP) hydrogel had good biocompatibility, elastic modulus, and electrical conductivity that matched the sciatic nerve. The in vitro experiments indicated that ACCP3 (3 wt% CP) promoted the proliferation and migration of the SCs. Injection of ACCP3 into the crush injury site of the sciatic nerve reduced tissue resistivity, increased nerve conduction velocity, decreased the sciatic nerve functional index, enhanced the expression of neuronal axon-specific proteins, induced axon extension and remyelination, and prevented muscle denervation atrophy. Overall, the injectable self-healing conductive ACCP3 hydrogel, as a safe and effective neural tissue substitute, opens up a new way for the treatment of peripheral nerve injury and has potential clinical application.

Value of Edema-like Marrow Signal Intensity in Diagnosis of Joint Pain: Radiologists' Perspective.

Musculoskeletal pain is a significant contributor to disability. The mechanism and target of the treatment should be optimized by imaging, but currently no accepted gold standard exists to image pain. In addition to end-organ pathology, other mediators also contribute to nociception, such as angiogenesis, axonal extension, immunologic modulation, and central sensitization. Recent research indicates that local inflammation is a significant contributor to pain in the extremities; therefore, we focus here on edema-like marrow signal intensity (ELMSI). We examine both the relevance of ELMSI for pain and novel imaging techniques.

Shinbaro2 enhances axonal extension beyond the glial scar for functional recovery in rats with contusive spinal cord injury

Spinal cord injury (SCI) is a devastating event that often results in the inflammatory condition of glial scar tissue formation, impeding neural regeneration and recovery. Reducing the inflammatory response and inhibiting glial formation are promising strategies for improving SCI outcomes. Here, we introduce a new role for Shinbaro2 (Sh2), known for its anti-inflammatory and pain-reducing effects, in ameliorating glial scars formed in the damaged spinal cord and promoting axon growth after SCI. Sh2 was applied at various concentrations to cultivate primary spinal cord neurons. Concentrations of 1 and 2 mg/mL effectively enhanced cell viability and axonal outgrowth in spinal cord neurons subjected to hydrogen peroxide or laceration injury. Sh2 helped reduce neuroinflammation by increasing anti-inflammatory M2 macrophages (arginase 1) and decreasing inflammatory cells, ultimately reducing lesion size. In scar formation, Sh2 inhibited the expression of β-catenin and nestin in reactive astrocytes in the injured spinal cord. Moreover, Sh2 suppressed the expression of chondroitin sulfate proteoglycans and SOX9, which are involved in scar formation. Furthermore, Sh2 promoted the sprouting of serotonergic axons and the growth of neurofibrillary tangles, enhancing motor function recovery in SCI. These findings highlight the potential of Sh2 as an SCI therapeutic intervention, offering hope for neural and functional restoration in individuals with this debilitating condition.

Stat3 Has a Different Role in Axon Growth During Development Than It Does in Axon Regeneration After Injury.

Signal transducer and activator of transcription 3 (STAT3) is essential for neural development and regeneration as a key transcription factor and mitochondrial activator. However, the mechanism of Stat3 in axon development and regeneration has not been fully understood. In this study, using zebrafish posterior lateral line (PLL) axons, we demonstrate that Stat3 plays distinct roles in PLL axon embryonic growth and regeneration. Our experiments indicate that stat3 is required for PLL axon extension. In stat3 mutant zebrafish, the PLL axon ends were stalled at the level of the cloaca, and expression of stat3 rescues the PLL axon growth in a cell-autonomous manner. Jak/Stat signaling inhibition did not affect PLL axon growth indicating Jak/Stat was dispensable for PLL axon growth. In addition, we found that Stat3 was co-localized with mitochondria in PLL axons and important for the mitochondrial membrane potential and ATPase activity. The PLL axon growth defect of stat3 mutants was mimicked and rescued by rotenone and DCHC treatment, respectively, which suggests that Stat3 regulates PLL axon growth through mitochondrial Stat3. By contrast, mutation of stat3 or Jak/Stat signaling inhibition retarded PLL axon regeneration. Meanwhile, we also found Schwann cell migration was also inhibited in stat3 mutants. Taken together, Stat3 is required for embryonic PLL axon growth by regulating the ATP synthesis efficiency of mitochondria, whereas Stat3 stimulates PLL axon regeneration by regulating Schwann cell migration via Jak/Stat signaling. Our findings show a new mechanism of Stat3 in axon growth and regeneration.