RNA sequencing of exosomes secreted by fibroblast and Schwann cells elucidates mechanisms underlying peripheral nerve regeneration

Exosomes exhibit complex biological functions and mediate a variety of biological processes,such as promoting axonal regeneration and functional recove ry after injury.Long non-coding RNAs(IncRNAs)have been reported to play a crucial role in axonal regeneration.Howeve r,the role of the IncRNA-microRNAmessenger RNA(mRNA)-competitive endogenous RNA(ceRNA)network in exosome-mediated axonal regeneration remains unclear.In this study,we performed RNA transcriptome sequencing analysis to assess mRNA expression patterns in exosomes produced by cultured fibroblasts(FC-EXOs)and Schwann cells(SCEXOs).Diffe rential gene expression analysis,Gene Ontology analysis,Kyoto Encyclopedia of Genes and Genomes analysis,and protein-protein intera ction network analysis were used to explo re the functions and related pathways of RNAs isolated from FC-EXOs and SC-EXOs.We found that the ribosome-related central gene Rps5 was enriched in FC-EXOs and SC-EXOs,which suggests that it may promote axonal regeneration.In addition,using the miRWalk and Starbase prediction databases,we constructed a regulatory network of ceRNAs targeting Rps5,including 27 microRNAs and five IncRNAs.The ceRNA regulatory network,which included Ftx and Miat,revealed that exsosome-derived Rps5 inhibits scar formation and promotes axonal regeneration and functional recovery after nerve injury.Our findings suggest that exosomes derived from fibro blast and Schwann cells could be used to treat injuries of peripheral nervous system.

Gp78 regulates PMP22 and causes ER stress and autophagy in EV71-VP1-overexpressing mouse Schwann cells

Background:During Enterovirus type 71(EV71)infection,the structural viral protein 1(VP1)activates endoplasmic reticulum(ER)stress associated with peripheral myelin protein 22(PMP22)accumulation and induces autophagy.However,the specific mechanism behind this process remains elusive.Methods:In this research,we used the VP1-overexpressing mouse Schwann cells(SCs)models co-transfected with a PMP22 silencing or Autocrine motility factor receptor(AMFR/gp78)overexpressing vector to explore the regulation of gp78 on PMP22 and its relationship with autophagy and apoptosis.Results:The activity of gp78 could be influenced by EV71-VP1,leading to a decrease in the ubiquitination and degradation of PMP22,resulting in PMP22 accumulation in ER.In VP1-overexpressing mouse SCs,all three ER stress sensors,including pancreatic endoplasmic reticulum kinase(PERK),activating transcription factor 6(ATF6)and inositol-requiring enzyme 1(IRE1)and the related downstream signals(C/EBP-homologous protein(CHOP)and Caspase 12)were activated,as well as the ER-resident chaperone Glucose-regulated protein 78(GRP78).In addition,VP1 upregulated the autophagy marker Microtubule-associated protein 1 light chain 3 beta(LC3B),while PMP22 silencing or gp78 overexpression reversed the phenomenon.Meanwhile,PMP22 silencing or gp78 overexpression increased proliferation of EV71-VP1-transfected mouse SCs.Conclusion:Gp78 could regulate PMP22 accumulation through ubiquitination degradation and cause ER stress and autophagy in EV71-VP1-overexpressing mouse SCs.Therefore,the gp78/PMP22/ER stress axis might emerge as a promising therapeutic target for myelin and neuronal damage induced by EV71 infection.

Mechanism of inflammatory response and therapeutic effects of stem cells in ischemic stroke:current evidence and future perspectives

Ischemic stroke is a leading cause of death and disability worldwide,with an increasing trend and tendency for onset at a younger age.China,in particular,bears a high burden of stroke cases.In recent years,the inflammatory response after stroke has become a research hotspot:understanding the role of inflammatory response in tissue damage and repair following ischemic stroke is an important direction for its treatment.This review summarizes several major cells involved in the inflammatory response following ischemic stroke,including microglia,neutrophils,monocytes,lymphocytes,and astrocytes.Additionally,we have also highlighted the recent progress in various treatments for ischemic stroke,particularly in the field of stem cell therapy.Overall,understanding the complex interactions between inflammation and ischemic stroke can provide valuable insights for developing treatment strategies and improving patient outcomes.Stem cell therapy may potentially become an important component of ischemic stroke treatment.

Chondroitinase ABC combined with Schwann cell transplantation enhances restoration of neural connection and functional recovery following acute and chronic spinal cord injury

Schwann cell transplantation is considered one of the most promising cell-based therapy to repair injured spinal cord due to its unique growth-promoting and myelin-forming properties.A the Food and Drug Administration-approved Phase I clinical trial has been conducted to evaluate the safety of transplanted human autologous Schwann cells to treat patients with spinal cord injury.A major challenge for Schwann cell transplantation is that grafted Schwann cells are confined within the lesion cavity,and they do not migrate into the host environment due to the inhibitory barrier formed by injury-induced glial scar,thus limiting axonal reentry into the host spinal cord.Here we introduce a combinatorial strategy by suppressing the inhibitory extracellular environment with injection of lentivirus-mediated transfection of chondroitinase ABC gene at the rostral and caudal borders of the lesion site and simultaneously leveraging the repair capacity of transplanted Schwann cells in adult rats following a mid-thoracic contusive spinal cord injury.We report that when the glial scar was degraded by chondroitinase ABC at the rostral and caudal lesion borders,Schwann cells migrated for considerable distances in both rostral and caudal directions.Such Schwann cell migration led to enhanced axonal regrowth,including the serotonergic and dopaminergic axons originating from supraspinal regions,and promoted recovery of locomotor and urinary bladder functions.Importantly,the Schwann cell survival and axonal regrowth persisted up to 6 months after the injury,even when treatment was delayed for 3 months to mimic chronic spinal cord injury.These findings collectively show promising evidence for a combinatorial strategy with chondroitinase ABC and Schwann cells in promoting remodeling and recovery of function following spinal cord injury.

miRNA-21-5p is an important contributor to the promotion of injured peripheral nerve regeneration using hypoxia-pretreated bone marrow-derived neural crest cells

Our previous study found that rat bone marrow–derived neural crest cells(acting as Schwann cell progenitors)have the potential to promote long-distance nerve repair.Cell-based therapy can enhance peripheral nerve repair and regeneration through paracrine bioactive factors and intercellular communication.Nevertheless,the complex contributions of various types of soluble cytokines and extracellular vesicle cargos to the secretome remain unclear.To investigate the role of the secretome and extracellular vesicles in repairing damaged peripheral nerves,we collected conditioned culture medium from hypoxia-pretreated neural crest cells,and found that it significantly promoted the repair of sensory neurons damaged by oxygen-glucose deprivation.The mRNA expression of trophic factors was highly expressed in hypoxia-pretreated neural crest cells.We performed RNA sequencing and bioinformatics analysis and found that miR-21-5p was enriched in hypoxia-pretreated extracellular vesicles of neural crest cells.Subsequently,to further clarify the role of hypoxia-pretreated neural crest cell extracellular vesicles rich in miR-21-5p in axonal growth and regeneration of sensory neurons,we used a microfluidic axonal dissociation model of sensory neurons in vitro,and found that hypoxia-pretreated neural crest cell extracellular vesicles promoted axonal growth and regeneration of sensory neurons,which was greatly dependent on loaded miR-21-5p.Finally,we constructed a miR-21-5p-loaded neural conduit to repair the sciatic nerve defect in rats and found that the motor and sensory functions of injured rat hind limb,as well as muscle tissue morphology of the hind limbs,were obviously restored.These findings suggest that hypoxia-pretreated neural crest extracellular vesicles are natural nanoparticles rich in miRNA-21-5p.miRNA-21-5p is one of the main contributors to promoting nerve regeneration by the neural crest cell secretome.This helps to explain the mechanism of action of the secretome and extracellular vesicles of neural crest cells in repairing damaged peripheral nerves,and also promotes the application of miR-21-5p in tissue engineering regeneration medicine.

Treating amyotrophic lateral sclerosis with allogeneic Schwann cell-derived exosomal vesicles: a case report

Schwann cells are essential for the maintenance and function of motor neurons,axonal networks,and the neuromuscular junction.In amyotrophic lateral sclerosis,where motor neuron function is progressively lost,Schwann cell function may also be impaired.Recently,important signaling and potential trophic activities of Schwann cell-derived exosomal vesicles have been reported.This case report describes the treatment of a patient with advanced amyotrophic lateral sclerosis using serial intravenous infusions of allogeneic Schwann cell-derived exosomal vesicles,marking,to our knowledge,the first instance of such treatment.An 81-year-old male patient presented with a 1.5-year history of rapidly progressive amyotrophic lateral sclerosis.After initial diagnosis,the patient underwent a combination of generic riluzole,sodium phenylbutyrate for the treatment of amyotrophic lateral sclerosis,and taurursodiol.The patient volunteered to participate in an FDA-approved single-patient expanded access treatment and received weekly intravenous infusions of allogeneic Schwann cell-derived exosomal vesicles to potentially restore impaired Schwann cell and motor neuron function.We confirmed that cultured Schwann cells obtained from the amyotrophic lateral sclerosis patient via sural nerve biopsy appeared impaired(senescent)and that exposure of the patient’s Schwann cells to allogeneic Schwann cell-derived exosomal vesicles,cultured expanded from a cadaver donor improved their growth capacity in vitro.After a period of observation lasting 10 weeks,during which amyotrophic lateral sclerosis Functional Rating Scale-Revised and pulmonary function were regularly monitored,the patient received weekly consecutive infusions of 1.54×1012(×2),and then consecutive infusions of 7.5×1012(×6)allogeneic Schwann cell-derived exosomal vesicles diluted in 40 mL of Dulbecco’s phosphate-buffered saline.None of the infusions were associated with adverse events such as infusion reactions(allergic or otherwise)or changes in vital signs.Clinical lab serum neurofilament and cytokine levels measured prior to each infusion varied somewhat without a clear trend.A more sensitive in-house assay suggested possible inflammasome activation during the disease course.A trend for clinical stabilization was observed during the infusion period.Our study provides a novel approach to address impaired Schwann cells and possibly motor neuron function in patients with amyotrophic lateral sclerosis using allogeneic Schwann cell-derived exosomal vesicles.Initial findings suggest that this approach is safe.

Meningeal lymphatic vessel crosstalk with central nervous system immune cells in aging and neurodegenerative diseases

Meningeal lymphatic vessels form a relationship between the nervous system and periphery, which is relevant in both health and disease. Meningeal lymphatic vessels not only play a key role in the drainage of brain metabolites but also contribute to antigen delivery and immune cell activation. The advent of novel genomic technologies has enabled rapid progress in the characterization of myeloid and lymphoid cells and their interactions with meningeal lymphatic vessels within the central nervous system. In this review, we provide an overview of the multifaceted roles of meningeal lymphatic vessels within the context of the central nervous system immune network, highlighting recent discoveries on the immunological niche provided by meningeal lymphatic vessels. Furthermore, we delve into the mechanisms of crosstalk between meningeal lymphatic vessels and immune cells in the central nervous system under both homeostatic conditions and neurodegenerative diseases, discussing how these interactions shape the pathological outcomes. Regulation of meningeal lymphatic vessel function and structure can influence lymphatic drainage, cerebrospinal fluid-borne immune modulators, and immune cell populations in aging and neurodegenerative disorders, thereby playing a key role in shaping meningeal and brain parenchyma immunity.

Small extracellular vesicles derived from human induced pluripotent stem cell-differentiated neural progenitor cells mitigate retinal ganglion cell degeneration in a mouse model of optic nerve injury

Several studies have found that transplantation of neural progenitor cells(NPCs)promotes the survival of injured neurons.However,a poor integration rate and high risk of tumorigenicity after cell transplantation limits their clinical application.Small extracellular vesicles(sEVs)contain bioactive molecules for neuronal protection and regeneration.Previous studies have shown that stem/progenitor cell-derived sEVs can promote neuronal survival and recovery of neurological function in neurodegenerative eye diseases and other eye diseases.In this study,we intravitreally transplanted sEVs derived from human induced pluripotent stem cells(hiPSCs)and hiPSCs-differentiated NPCs(hiPSC-NPC)in a mouse model of optic nerve crush.Our results show that these intravitreally injected sEVs were ingested by retinal cells,especially those localized in the ganglion cell layer.Treatment with hiPSC-NPC-derived sEVs mitigated optic nerve crush-induced retinal ganglion cell degeneration,and regulated the retinal microenvironment by inhibiting excessive activation of microglia.Component analysis further revealed that hiPSC-NPC derived sEVs transported neuroprotective and anti-inflammatory miRNA cargos to target cells,which had protective effects on RGCs after optic nerve injury.These findings suggest that sEVs derived from hiPSC-NPC are a promising cell-free therapeutic strategy for optic neuropathy.

Therapeutic potential of stem cells in subarachnoid hemorrhage

Aneurysm rupture can result in subarachnoid hemorrhage,a condition with potentially severe consequences,such as disability and death.In the acute stage,early brain injury manifests as intracranial pressure elevation,global cerebral ischemia,acute hydrocephalus,and direct blood–brain contact due to aneurysm rupture.This may subsequently cause delayed cerebral infarction,often with cerebral vasospasm,significantly affecting patient outcomes.Chronic complications such as brain volume loss and chronic hydrocephalus can further impact outcomes.Investigating the mechanisms of subarachnoid hemorrhage-induced brain injury is paramount for identifying effective treatments.Stem cell therapy,with its multipotent differentiation capacity and anti-inflammatory effects,has emerged as a promising approach for treating previously deemed incurable conditions.This review focuses on the potential application of stem cells in subarachnoid hemorrhage pathology and explores their role in neurogenesis and as a therapeutic intervention in preclinical and clinical subarachnoid hemorrhage studies.

Photobiomodulation:a novel approach to promote trans-differentiation of adipose-derived stem cells into neuronal-like cells

Photobiomodulation,originally used red and near-infrared lasers,can alter cellular metabolism.It has been demonstrated that the visible spectrum at 451-540 nm does not necessarily increase cell proliferation,near-infrared light promotes adipose stem cell proliferation and affects adipose stem cell migration,which is necessary for the cells homing to the site of injury.In this in vitro study,we explored the potential of adipose-derived stem cells to differentiate into neurons for future translational regenerative treatments in neurodegenerative disorders and brain injuries.We investigated the effects of various biological and chemical inducers on trans-differentiation and evaluated the impact of photobiomodulation using 825 nm near-infrared and 525 nm green laser light at 5 J/cm2.As adipose-derived stem cells can be used in autologous grafting and photobiomodulation has been shown to have biostimulatory effects.Our findings reveal that adipose-derived stem cells can indeed trans-differentiate into neuronal cells when exposed to inducers,with pre-induced cells exhibiting higher rates of proliferation and trans-differentiation compared with the control group.Interestingly,green laser light stimulation led to notable morphological changes indicative of enhanced trans-differentiation,while near-infrared photobiomodulation notably increased the expression of neuronal markers.Through biochemical analysis and enzyme-linked immunosorbent assays,we observed marked improvements in viability,proliferation,membrane permeability,and mitochondrial membrane potential,as well as increased protein levels of neuron-specific enolase and ciliary neurotrophic factor.Overall,our results demonstrate the efficacy of photobiomodulation in enhancing the trans-differentiation ability of adipose-derived stem cells,offering promising prospects for their use in regenerative medicine for neurodegenerative disorders and brain injuries.

Small extracellular vesicles derived from cerebral endothelial cells with elevated microRNA 27a promote ischemic stroke recovery

Axonal remodeling is a critical aspect of ischemic brain repair processes and contributes to spontaneous functional recovery.Our previous in vitro study demonstrated that exosomes/small extracellular vesicles(sEVs)isolated from cerebral endothelial cells(CEC-sEVs)of ischemic brain promote axonal growth of embryonic cortical neurons and that microRNA 27a(miR-27a)is an elevated miRNA in ischemic CEC-sEVs.In the present study,we investigated whether normal CEC-sEVs engineered to enrich their levels of miR-27a(27a-sEVs)further enhance axonal growth and improve neurological outcomes after ischemic stroke when compared with treatment with non-engineered CEC-sEVs.27a-sEVs were isolated from the conditioned medium of healthy mouse CECs transfected with a lentiviral miR-27a expression vector.Small EVs isolated from CECs transfected with a scramble vector(Scra-sEVs)were used as a control.Adult male mice were subjected to permanent middle cerebral artery occlusion and then were randomly treated with 27a-sEVs or Scra-sEVs.An array of behavior assays was used to measure neurological function.Compared with treatment of ischemic stroke with Scra-sEVs,treatment with 27a-sEVs significantly augmented axons and spines in the peri-infarct zone and in the corticospinal tract of the spinal grey matter of the denervated side,and significantly improved neurological outcomes.In vitro studies demonstrated that CEC-sEVs carrying reduced miR-27a abolished 27a-sEV-augmented axonal growth.Ultrastructural analysis revealed that 27a-sEVs systemically administered preferentially localized to the pre-synaptic active zone,while quantitative reverse transcription-polymerase chain reaction and Western Blot analysis showed elevated miR-27a,and reduced axonal inhibitory proteins Semaphorin 6A and Ras Homolog Family Member A in the peri-infarct zone.Blockage of the Clathrin-dependent endocytosis pathway substantially reduced neuronal internalization of 27a-sEVs.Our data provide evidence that 27a-sEVs have a therapeutic effect on stroke recovery by promoting axonal remodeling and improving neurological outcomes.Our findings also suggest that suppression of axonal inhibitory proteins such as Semaphorin 6A may contribute to the beneficial effect of 27a-sEVs on axonal remodeling.

Advances in therapies using mesenchymal stem cells and their exosomes for treatment of peripheral nerve injury:state of the art and future perspectives

“Peripheral nerve injury”refers to damage or trauma affecting nerves outside the brain and spinal cord.Peripheral nerve injury results in movements or sensation impairments,and represents a serious public health problem.Although severed peripheral nerves have been effectively joined and various therapies have been offered,recovery of sensory or motor functions remains limited,and efficacious therapies for complete repair of a nerve injury remain elusive.The emerging field of mesenchymal stem cells and their exosome-based therapies hold promise for enhancing nerve regeneration and function.Mesenchymal stem cells,as large living cells responsive to the environment,secrete various factors and exosomes.The latter are nano-sized extracellular vesicles containing bioactive molecules such as proteins,microRNA,and messenger RNA derived from parent mesenchymal stem cells.Exosomes have pivotal roles in cell-to-cell communication and nervous tissue function,offering solutions to changes associated with cell-based therapies.Despite ongoing investigations,mesenchymal stem cells and mesenchymal stem cell-derived exosome-based therapies are in the exploratory stage.A comprehensive review of the latest preclinical experiments and clinical trials is essential for deep understanding of therapeutic strategies and for facilitating clinical translation.This review initially explores current investigations of mesenchymal stem cells and mesenchymal stem cell-derived exosomes in peripheral nerve injury,exploring the underlying mechanisms.Subsequently,it provides an overview of the current status of mesenchymal stem cell and exosomebased therapies in clinical trials,followed by a comparative analysis of therapies utilizing mesenchymal stem cells and exosomes.Finally,the review addresses the limitations and challenges associated with use of mesenchymal stem cell-derived exosomes,offering potential solutions and guiding future directions.

Induced neural stem cells regulate microglial activation through Akt-mediated upregulation of CXCR4 and Crry in a mouse model of closed head injury

Microglial activation that occurs rapidly after closed head injury may play important and complex roles in neuroinflammation-associated neuronal damage and repair.We previously reported that induced neural stem cells can modulate the behavior of activated microglia via CXCL12/CXCR4 signaling,influencing their activation such that they can promote neurological recovery.However,the mechanism of CXCR4 upregulation in induced neural stem cells remains unclear.In this study,we found that nuclear factor-κB activation induced by closed head injury mouse serum in microglia promoted CXCL12 and tumor necrosis factor-αexpression but suppressed insulin-like growth factor-1 expression.However,recombinant complement receptor 2-conjugated Crry(CR2-Crry)reduced the effects of closed head injury mouse serum-induced nuclear factor-κB activation in microglia and the levels of activated microglia,CXCL12,and tumor necrosis factor-α.Additionally,we observed that,in response to stimulation(including stimulation by CXCL12 secreted by activated microglia),CXCR4 and Crry levels can be upregulated in induced neural stem cells via the interplay among CXCL12/CXCR4,Crry,and Akt signaling to modulate microglial activation.In agreement with these in vitro experimental results,we found that Akt activation enhanced the immunoregulatory effects of induced neural stem cell grafts on microglial activation,leading to the promotion of neurological recovery via insulin-like growth factor-1 secretion and the neuroprotective effects of induced neural stem cell grafts through CXCR4 and Crry upregulation in the injured cortices of closed head injury mice.Notably,these beneficial effects of Akt activation in induced neural stem cells were positively correlated with the therapeutic effects of induced neural stem cells on neuronal injury,cerebral edema,and neurological disorders post–closed head injury.In conclusion,our findings reveal that Akt activation may enhance the immunoregulatory effects of induced neural stem cells on microglial activation via upregulation of CXCR4 and Crry,thereby promoting induced neural stem cell–mediated improvement of neuronal injury,cerebral edema,and neurological disorders following closed head injury.

Perilipin-2 mediates ferroptosis in oligodendrocyte progenitor cells and myelin injury after ischemic stroke

Differentiation of oligodendrocyte progenitor cells into mature myelin-forming oligodendrocytes contributes to remyelination.Failure of remyelination due to oligodendrocyte progenitor cell death can result in severe nerve damage.Ferroptosis is an iron-dependent form of regulated cell death caused by membrane rupture induced by lipid peroxidation,and plays an important role in the pathological process of ischemic stroke.However,there are few studies on oligodendrocyte progenitor cell ferroptosis.We analyzed transcriptome sequencing data from GEO databases and identified a role of ferroptosis in oligodendrocyte progenitor cell death and myelin injury after cerebral ischemia.Bioinformatics analysis suggested that perilipin-2(PLIN2)was involved in oligodendrocyte progenitor cell ferroptosis.PLIN2 is a lipid storage protein and a marker of hypoxia-sensitive lipid droplet accumulation.For further investigation,we established a mouse model of cerebral ischemia/reperfusion.We found significant myelin damage after cerebral ischemia,as well as oligodendrocyte progenitor cell death and increased lipid peroxidation levels around the infarct area.The ferroptosis inhibitor,ferrostatin-1,rescued oligodendrocyte progenitor cell death and subsequent myelin injury.We also found increased PLIN2 levels in the peri-infarct area that co-localized with oligodendrocyte progenitor cells.Plin2 knockdown rescued demyelination and improved neurological deficits.Our findings suggest that targeting PLIN2 to regulate oligodendrocyte progenitor cell ferroptosis may be a potential therapeutic strategy for rescuing myelin damage after cerebral ischemia.

Exosomes originating from neural stem cells undergoing necroptosis participate in cellular communication by inducing TSC2 upregulation of recipient cells following spinal cord injury

We previously demonstrated that inhibiting neural stem cells necroptosis enhances functional recovery after spinal cord injury.While exosomes are recognized as playing a pivotal role in neural stem cells exocrine function,their precise function in spinal cord injury remains unclear.To investigate the role of exosomes generated following neural stem cells necroptosis after spinal cord injury,we conducted singlecell RNA sequencing and validated that neural stem cells originate from ependymal cells and undergo necroptosis in response to spinal cord injury.Subsequently,we established an in vitro necroptosis model using neural stem cells isolated from embryonic mice aged 16-17 days and extracted exosomes.The results showed that necroptosis did not significantly impact the fundamental characteristics or number of exosomes.Transcriptome sequencing of exosomes in necroptosis group identified 108 differentially expressed messenger RNAs,104 long non-coding RNAs,720 circular RNAs,and 14 microRNAs compared with the control group.Construction of a competing endogenous RNA network identified the following hub genes:tuberous sclerosis 2(Tsc2),solute carrier family 16 member 3(Slc16a3),and forkhead box protein P1(Foxp1).Notably,a significant elevation in TSC2 expression was observed in spinal cord tissues following spinal cord injury.TSC2-positive cells were localized around SRY-box transcription factor 2-positive cells within the injury zone.Furthermore,in vitro analysis revealed increased TSC2 expression in exosomal receptor cells compared with other cells.Further assessment of cellular communication following spinal cord injury showed that Tsc2 was involved in ependymal cellular communication at 1 and 3 days post-injury through the epidermal growth factor and midkine signaling pathways.In addition,Slc16a3 participated in cellular communication in ependymal cells at 7 days post-injury via the vascular endothelial growth factor and macrophage migration inhibitory factor signaling pathways.Collectively,these findings confirm that exosomes derived from neural stem cells undergoing necroptosis play an important role in cellular communication after spinal cord injury and induce TSC2 upregulation in recipient cells.

Müller cells are activated in response to retinal outer nuclear layer degeneration in rats subjected to simulated weightlessness conditions

A microgravity environment has been shown to cause ocular damage and affect visual acuity,but the underlying mechanisms remain unclear.Therefore,we established an animal model of weightlessness via tail suspension to examine the pathological changes and molecular mechanisms of retinal damage under microgravity.After 4 weeks of tail suspension,there were no notable alterations in retinal function and morphology,while after 8 weeks of tail suspension,significant reductions in retinal function were observed,and the outer nuclear layer was thinner,with abundant apoptotic cells.To investigate the mechanism underlying the degenerative changes that occurred in the outer nuclear layer of the retina,proteomics was used to analyze differentially expressed proteins in rat retinas after 8 weeks of tail suspension.The results showed that the expression levels of fibroblast growth factor 2(also known as basic fibroblast growth factor)and glial fibrillary acidic protein,which are closely related to Müller cell activation,were significantly upregulated.In addition,Müller cell regeneration and Müller cell gliosis were observed after 4 and 8 weeks,respectively,of simulated weightlessness.These findings indicate that Müller cells play an important regulatory role in retinal outer nuclear layer degeneration during weightlessness.

Small molecule inhibitor DDQ-treated hippocampal neuronal cells show improved neurite outgrowth and synaptic branching

The process of neurite outgrowth and branching is a crucial aspect of neuronal development and regeneration.Axons and dendrites,sometimes referred to as neurites,are extensions of a neuron's cellular body that are used to start networks.Here we explored the effects of diethyl(3,4-dihydroxyphenethylamino)(quinolin-4-yl)methylphosphonate(DDQ)on neurite developmental features in HT22 neuronal cells.In this work,we examined the protective effects of DDQ on neuronal processes and synaptic outgrowth in differentiated HT22cells expressing mutant Tau(mTau)cDNA.To investigate DDQ chara cteristics,cell viability,biochemical,molecular,western blotting,and immunocytochemistry were used.Neurite outgrowth is evaluated through the segmentation and measurement of neural processes.These neural processes can be seen and measured with a fluorescence microscope by manually tracing and measuring the length of the neurite growth.These neuronal processes can be observed and quantified with a fluorescent microscope by manually tracing and measuring the length of the neuronal HT22.DDQ-treated mTau-HT22 cells(HT22 cells transfected with cDNA mutant Tau)were seen to display increased levels of synaptophysin,MAP-2,andβ-tubulin.Additionally,we confirmed and noted reduced levels of both total and p-Tau,as well as elevated levels of microtubule-associated protein 2,β-tubulin,synaptophysin,vesicular acetylcholine transporter,and the mitochondrial biogenesis protein-pe roxisome prolife rator-activated receptor-gamma coactivator-1α.In mTa u-expressed HT22 neurons,we observed DDQ enhanced the neurite characteristics and improved neurite development through increased synaptic outgrowth.Our findings conclude that mTa u-HT22(Alzheimer's disease)cells treated with DDQ have functional neurite developmental chara cteristics.The key finding is that,in mTa u-HT22 cells,DDQ preserves neuronal structure and may even enhance nerve development function with mTa u inhibition.

Hypoxia-preconditioned bone marrow-derived mesenchymal stem cells protect neurons from cardiac arrest-induced pyroptosis

Cardiac arrest can lead to severe neurological impairment as a result of inflammation,mitochondrial dysfunction,and post-cardiopulmonary resuscitation neurological damage.Hypoxic preconditioning has been shown to improve migration and survival of bone marrow–derived mesenchymal stem cells and reduce pyroptosis after cardiac arrest,but the specific mechanisms by which hypoxia-preconditioned bone marrow–derived mesenchymal stem cells protect against brain injury after cardiac arrest are unknown.To this end,we established an in vitro co-culture model of bone marrow–derived mesenchymal stem cells and oxygen–glucose deprived primary neurons and found that hypoxic preconditioning enhanced the protective effect of bone marrow stromal stem cells against neuronal pyroptosis,possibly through inhibition of the MAPK and nuclear factor κB pathways.Subsequently,we transplanted hypoxia-preconditioned bone marrow–derived mesenchymal stem cells into the lateral ventricle after the return of spontaneous circulation in an 8-minute cardiac arrest rat model induced by asphyxia.The results showed that hypoxia-preconditioned bone marrow–derived mesenchymal stem cells significantly reduced cardiac arrest–induced neuronal pyroptosis,oxidative stress,and mitochondrial damage,whereas knockdown of the liver isoform of phosphofructokinase in bone marrow–derived mesenchymal stem cells inhibited these effects.To conclude,hypoxia-preconditioned bone marrow–derived mesenchymal stem cells offer a promising therapeutic approach for neuronal injury following cardiac arrest,and their beneficial effects are potentially associated with increased expression of the liver isoform of phosphofructokinase following hypoxic preconditioning.

Gamma-glutamyl transferase 5 overexpression in cerebrovascular endothelial cells improves brain pathology,cognition,and behavior in APP/PS1 mice

In patients with Alzheimer’s disease,gamma-glutamyl transferase 5(GGT5)expression has been observed to be downregulated in cerebrovascular endothelial cells.However,the functional role of GGT5 in the development of Alzheimer’s disease remains unclear.This study aimed to explore the effect of GGT5 on cognitive function and brain pathology in an APP/PS1 mouse model of Alzheimer’s disease,as well as the underlying mechanism.We observed a significant reduction in GGT5 expression in two in vitro models of Alzheimer’s disease(Aβ_(1-42)-treated hCMEC/D3 and bEnd.3 cells),as well as in the APP/PS1 mouse model.Additionally,injection of APP/PS1 mice with an adeno-associated virus encoding GGT5 enhanced hippocampal synaptic plasticity and mitigated cognitive deficits.Interestingly,increasing GGT5 expression in cerebrovascular endothelial cells reduced levels of both soluble and insoluble amyloid-βin the brains of APP/PS1 mice.This effect may be attributable to inhibition of the expression ofβ-site APP cleaving enzyme 1,which is mediated by nuclear factor-kappa B.Our findings demonstrate that GGT5 expression in cerebrovascular endothelial cells is inversely associated with Alzheimer’s disease pathogenesis,and that GGT5 upregulation mitigates cognitive deficits in APP/PS1 mice.These findings suggest that GGT5 expression in cerebrovascular endothelial cells is a potential therapeutic target and biomarker for Alzheimer’s disease.

Effect of Rat Schwann Cell Secretion on Proliferation and Differentiation of Human Neural Stem Cells

Objective To investigate the effect of rat Schwann cell secretion on the proliferation and differentiation of human embryonic neural stem cells (NSCs). Methods The samples were divided into three groups. In Group One, NSCs were cultured in DMED/F12 in which Schwann cells had grown for one day. In Group Two, NSCs and Schwann cells were co-cultured. In Group Three, NSCs were cultured in DMEM/F12. The morphology of NSCs was checked and β-tubulin, GalC, hoechst 33342 and GFAP labellings were detected. Results In Group One, all neural spheres were attached to the bottom and differentiated. The majority of them were p-tubulin positive while a few of cells were GFAP or GalC positive. In Group Two, neural spheres remained undifferentiatied and their proliferation was inhibited in places where Schwann cells were robust. In places where there were few Schwann cells, NSCs performed in a similar manner as in Group One. In Group Three, the cell growth state deteriorated day after day. On the 7th day, most NSCs died. Conclusion The secretion of rat Schwann cells has a growth supportive and differentiation-inducing effect on human NSCs.