Emerging strategies for nerve repair and regeneration in ischemic stroke:neural stem cell therapy

Ischemic stroke is a major cause of mortality and disability worldwide,with limited treatment options available in clinical practice.The emergence of stem cell therapy has provided new hope to the field of stroke treatment via the restoration of brain neuron function.Exogenous neural stem cells are beneficial not only in cell replacement but also through the bystander effect.Neural stem cells regulate multiple physiological responses,including nerve repair,endogenous regeneration,immune function,and blood-brain barrier permeability,through the secretion of bioactive substances,including extracellular vesicles/exosomes.However,due to the complex microenvironment of ischemic cerebrovascular events and the low survival rate of neural stem cells following transplantation,limitations in the treatment effect remain unresolved.In this paper,we provide a detailed summary of the potential mechanisms of neural stem cell therapy for the treatment of ischemic stroke,review current neural stem cell therapeutic strategies and clinical trial results,and summarize the latest advancements in neural stem cell engineering to improve the survival rate of neural stem cells.We hope that this review could help provide insight into the therapeutic potential of neural stem cells and guide future scientific endeavors on neural stem cells.

Novel advancements in threedimensional neural tissue engineering and regenerative medicine

Neurological diseases and injuries present some of the great- est challenges in modern medicine, often causing irrevers- ible and lifelong burdens in the people whom they afflict. Conditions of stroke, traumatic brain injury, spinal cord injury, and neurodegenerative diseases have devastating con- sequences on millions of people each year, and yet there are currently no therapies or interventions that can repair the structure of neural circuits and restore neural tissue function in the brain and spinal cord. Despite the challenges of over- coming these limitations, there are many new approaches under development that hold much promise. Neural tissue engineering aims to restore and influence the function of damaged or diseased neural tissue generally through the use of stem cells and biomaterials. Many types of biomaterials may be implemented in various designs to influence the survival, differentiation, and function of developing stem cells, as well as to guide neurite extension and morphological architecture of cell cultures. Such designs may aim to reca- pitulate the cellular interactions, extracellular matrix char- acteristics, biochemical factors, and sequences of events that occur in neurodevelopment, in addition to supporting cell survival, differentiation, and integration into innate neural tissue.

Engineering personalized neural tissue using functionalized transcription factors

Diseases and disorders of the central nervous system often require significant interventions to restore lost function due to their com- plexity. Examples of such disorders include Parkinson＇s disease, Alzheimer＇s disease, multiple sclerosis, traumatic brain injury, and spinal cord in）ury. These diseases and disorders result trom healthy cells being destroyed, which in turn causes dysfunction in the cen- tral nervous system, The death of these cells can trigger a cascade of events that affect the rest of the body, causing symptoms that become progressively worse over time. Developing strategies for repairing the damage to the central nervous system remains chal- lenging, in part due to its inability to regenerate.

Optogenetics and its application in neural degeneration and regeneration

Neural degeneration and regeneration are important topics in neurological diseases. There are limited options for therapeutic interventions in neurological diseases that provide simultaneous spatial and temporal control of neurons. This drawback increases side effects due to non-specific targeting. Optogenetics is a technology that allows precise spatial and temporal control of cells. Therefore, this technique has high potential as a therapeutic strategy for neurological diseases. Even though the application of optogenetics in understanding brain functional organization and complex behaviour states have been elaborated, reviews of its therapeutic potential especially in neurodegeneration and regeneration are still limited. This short review presents representative work in optogenetics in disease models such as spinal cord injury, multiple sclerosis, epilepsy, Alzheimer＇s disease and Parkinson＇s disease. It is aimed to provide a broader perspective on optogenetic therapeutic potential in neurodegeneration and neural regeneration.

LLaMA-LoRA Neural Prompt Engineering:A Deep Tuning Framework for Automatically Generating Chinese Text Logical Reasoning Thinking Chains

The exption of Chinese natural language processing(NLP)has stimulated research in the broader NLP domain.However,existing large language models have limitations in comprehending and reasoning in Chinese.This paper addresses these limitations by enhancing Chinese language models comprehension and reasoning capabilities while minimizing resource requirements.We propose LLaMA-LoRA,a neural prompt engineering framework that builds upon the LLaMA-13B model and incorporates the Low-Rank Adaptation(LoRA)of Large Language Models technique for refinement.Chain-of-Thought(CoT)are crucial for generating intermediate reasoning chains in language models,but their effectiveness can be limited by isolated language patterns.Erroneous reasoning resulting from conventional prompts negatively impacts model performance.Automatic prompts are introduced to encourage reasoning chain generation and accurate answer inference.Training the model with an extensive corpus of Chinese CoT data enhances its comprehension and reasoning abilities.The LLaMA-LoRA model demonstrates exceptional performance across numerous Chinese language tasks,surpassing benchmark performance achieved by related language models such as GPT-3.5,Chat-GLM,and OpenAssistant,delivering accurate,comprehensive,and professional answers.The availability of our open-source model code facilitates further research in the field of Chinese text logical reasoning thinking chains.

Fault Diagnosis for Manifold Absolute Pressure Sensor(MAP) of Diesel Engine Based on Elman Neural Network Observer

Intake system of diesel engine is a strong nonlinear system, and it is difficult to establish accurate model of intake system; and bias fault and precision degradation fault of MAP of diesel engine can＇t be diagnosed easily using model-based methods. Thus, a fault diagnosis method based on Elman neural network observer is proposed. By comparing simulation results of intake pressure based on BP network and Elman neural network, lower sampling error magnitude is gained using Elman neural network, and the error is less volatile. Forecast accuracy is between 0.015？0.017 5 and sample error is controlled within 0？0.07. Considering the output stability and complexity of solving comprehensively, Elman neural network with a single hidden layer and with 44 nodes is presented as intake system observer. By comparing the relations of confidence intervals of the residual value between the measured and predicted values, error variance and failures in various fault types. Then four typical MAP faults of diesel engine can be diagnosed： complete failure fault, bias fault, precision degradation fault and drift fault. The simulation results show： intake pressure is observable and selection of diagnostic strategy parameter reasonably can increase the accuracy of diagnosis;the proposed fault diagnosis method only depends on data and structural parameters of observer, not depends on the nonlinear model of air intake system. A fault diagnosis method is proposed not depending system model to observe intake pressure, and bias fault and precision degradation fault of MAP of diesel engine can be diagnosed based on residuals.

Biotinylated dextran amine as a neural tracer in the rat corticospinal tract

BACKGROUND： The corticospinal tract is the core structure of cerebral control of extremity movement and plasticity, which are prerequisites for movement rehabilitation after brain injury. The measurement and assessment of plasticity changes within the corticospinal tract has become one of the key goals in this field. OBJECTIVE： To explore the effects of biotinylated dextran amine （BDA） as a neural tracer in the rat corticospinal tract and the possibilities of assessing plasticity within the corticospinal tract. DESIGN： An observational experiment. SETTING： Department of Acupuncture of Chinese Medical College, Chongqing Medical University, Department of Neurology, the Second Affiliated Hospital, Chongqing Medical University. MATERIALS： Eighteen male adult Sprague Dawley （SD） rats of clean grade, weighing 200-250 g, were provided by the experimental animal center of Chongqing Medical University. The animal procedures in this study were in accordance with the animal ethics standards. BDA was provided by Vector Laboratories Company （USA, catalogue Sp- 1140; serial number R0721 ）. METHODS. This experiment was performed in the Laboratory of Chongqing Medical University between September and December 2006. Adult SD rats were used in the experiment and 15% BDA was injected slowly with a mini-syringe through two round （3 mm diameter） holes into the left sensory and motor cortex. The center of one hole was located 3 mm anterior from the anterior fontanel and 1.5 mm left of the midline; the second hole was located 1.5 mm posterior from the anterior fontanel and 4 mm left of the midline. Three injections were made at each hole at three different levels： 1.4, 1.2, and 1 mm ventral from the surface of the flat skull. After 14 days, the brains and spinal cords were removed and frozen. Sections were cut on a cryostat and BDA transportation absorbed by axons was observed under a fluorescence microscope. MAIN OUTCOME MEASURES： Axonal absorption and transportation of BDA was observed under fluorescence microscope. RESULTS： Eighteen SD rats were enrolled in this experiment; 12 rats were included in the final analysis and six were eliminated, resulting in a dropout rate of 33% （6/18）. BDA injected into the left cortex was absorbed in the axons, and fluorescence was observed throughout the pyramidal neurons and axons of the left cerebral cortex. At 14 days after rejection, BDA was detected in the midbrain and cervical enlargement along the CST, and axonal structures and Ranvier nodes were clearly observed with 200x magnification. CONCLUSION： BDA injected into the cerebral cortex effectively traces the corticospinal tract and is biologically stable over long distance transportation. In addition, the method of BDA tracing is fairly simple to perform.

Restoring nervous system structure and function using tissue engineered living scaffolds

Neural tissue engineering is premised on the integration of engineered living tissue with the host nervous system to directly restore lost function or to augment regenerative capacity following ner- vous system injury or neurodegenerative disease. Disconnection of axon pathways - the long-distance fibers connecting specialized regions of the central nervous system or relaying peripheral signals - is a common feature of many neurological disorders and injury. However, functional axonal regenera- tion rarely occurs due to extreme distances to targets, absence of directed guidance, and the presence of inhibitory factors in the central nervous system, resulting in devastating effects on cognitive and sensorimotor function. To address this need, we are pursuing multiple strategies using tissue engi- neered ＂living scaffolds＂, which are preformed three-dimensional constructs consisting of living neural cells in a defined, often anisotropic architecture. Living scaffolds are designed to restore function by serving as a living labeled pathway for targeted axonal regeneration - mimicking key developmental mechanisms- or by restoring lost neural circuitry via direct replacement of neurons and axonal tracts. We are currently utilizing preformed living scaffolds consisting of neuronal dusters spanned by long axonal tracts as regenerative bridges to facilitate long-distance axonal regeneration and for targeted neurosurgical reconstruction of local circuits in the brain. Although there are formidable challenges in predinical and clinical advancement, these living tissue engineered constructs represent a promising strategy to facilitate nervous system repair and functional recovery.

Chitosan tube bridging autologous nerve segments for the repair of long-segmental sciatic nerve defects

BACKGROUND：Previous tissue-engineered nerve studies have focused on artificial nerve and nerve cell cultures.The effects of regeneration chambers with autologous nerve bridging for the repair of nerve defects remain unclear.OBJECTIVE：To explore the feasibility and advantages of chitosan tube bridging autologous nerve segments for repairing 12-mm sciatic nerve defects in rats.DESIGN,TIME AND SETTING：A randomized,controlled,animal study using nerve tissue engineering was performed at the Animal Laboratory and Laboratory of Histology and Embryology,Liaoning Medical University from June 2008 to March 2009.MATERIALS：Chitosan powder was purchased from Jinan Haidebei Marine Bioengineering,China.METHODS：A sciatic nerve segment of approximately 8 mm was excised from the posterior margin of the piriformis muscle of Sprague Dawley rats.The two nerve ends shrank to form a 12-mm defect,and the nerve defect was repaired using a chitosan tube bridging autologous nerve segment （bridge group）,a chitosan tube-encapsulated autologous nerve segment （encapsulation group）,and a chitosan tube alone （chitosan tube alone group）,respectively.MAIN OUTCOME MEASURES：Histological and ultrastructural changes of the injured sciatic nerve;number of regenerated myelinated nerve fibers; nerve conduction velocity; leg muscle atrophy; and sciatic nerve functional index.RESULTS：At 4 months after implantation,the chitosan tube was absorbed.The tube was thin,but maintained the original shape,and vascular proliferation was observed around the tube.In the bridge group,regenerative myelinated nerve fibers were thick and orderly,with a thick myelin sheath and intact axonal structure.The number of myelinated nerve fibers and nerve conduction velocity were significantly greater compared with the other groups （P〈 0.01）.Moreover,nerve and muscle function was significantly improved following chitosan tube bridging autologous nerve segment treatment compared with the other groups （P〈 0.05 or P 〈 0.01）.CONCLUSION：Chitosan tube bridging autologous nerve segments exhibited better repair effects on nerve defects compared with chitosan tubeencapsulated autologous nerve segments and a chitosan tube alone.This method provided a simple and effective treatment for long-segmental nerve defects.

The use of hydrogel-delivered extracellular vesicles in recovery of motor function in stroke:a testable experimental hypothesis for clinical translation including behavioral and neuroimaging assessment approaches

Neural tissue engineering,nanotechnology and neuroregeneration are diverse biomedical disciplines that have been working together in recent decades to solve the complex problems linked to central nervous system(CNS)repair.It is known that the CNS demonstrates a very limited regenerative capacity because of a microenvironment that impedes effective regenerative processes,making development of CNS therapeutics challenging.Given the high prevalence of CNS conditions such as stroke that damage the brain and place a severe burden on afflicted individuals and on society,it is of utmost significance to explore the optimum methodologies for finding treatments that could be applied to humans for restoration of function to pre-injury levels.Extracellular vesicles(EVs),also known as exosomes,when derived from mesenchymal stem cells,are one of the most promising approaches that have been attempted thus far,as EVs deliver factors that stimulate recovery by acting at the nanoscale level on intercellular communication while avoiding the risks linked to stem cell transplantation.At the same time,advances in tissue engineering and regenerative medicine have offered the potential of using hydrogels as bio-scaffolds in order to provide the stroma required for neural repair to occur,as well as the release of biomolecules facilitating or inducing the reparative processes.This review introduces a novel experimental hypothesis regarding the benefits that could be offered if EVs were to be combined with biocompatible injectable hydrogels.The rationale behind this hypothesis is presented,analyzing how a hydrogel might prolong the retention of EVs and maximize the localized benefit to the brain.This sustained delivery of EVs would be coupled with essential guidance cues and structural support from the hydrogel until neural tissue remodeling and regeneration occur.Finally,the importance of including nonhuman primate models in the clinical translation pipeline,as well as the added benefit of multi-modal neuroimaging analysis to establish non-invasive,in vivo,quantifiable imagingbased biomarkers for CNS repair are discussed,aiming for more effective and safe clinical translation of such regenerative therapies to humans.

Self-assembled IKVAV Peptide Nanofibers Promote Adherence of PC12 Cells

Lack of biocompatibility and bioactivity is a big problem for the synthetic materials that have been generated for neural tissue engineering. To get around the problem and generate better scaffold for neural tissue repair, we intended to generate nano-fibers by self-assembly of polypeptide IKVAV. Bioactive IKVAV Peptide-Amphiphile （IKVAV-PA） was first synthesized and purified, the property of which was analyzed and determined by high-performance liquid chromatography （HPLC） and mass spectrometry （MS）. Then, by addition of hydrogen chloride （HC1）, self-assembly of IKVAV-PA was induced in vitro and nano-fibers formed as shown by transmission electron microscopy （TEM）. The effect of IKVAV nanofibers on adherence of PCI2 cells was assayed in cell culture and the results showed that the rates of adherence of PC12 increased significantly when the density of IKVAV was within a certain range （0.58 μg/cm^2 to 15.6 μg/cm^2）. However, its effect on the rates of adherence did not significantly alter with time, whether after 1 hour or 3 hours of culture. In general, we showed that IKVAV-PA can successfully self-assemble to form nanofiber, and promote rapid and stable adherence of PC12 cells, and the effect of the self-assembled IKVAV to promote PCI2 cells adherence is dosage-dependent within a certain range of densities.

Neuron-fibrous scaffold interfaces in the peripheral nervous system: a perspective on the structural requirements

The nerves of the peripheral nervous system are not able to effectively regenerate in cases of severe neural injury.This can result in debilitating consequences,including morbidity and lifelong impairments affecting the quality of the patient’s life.Recent findings in neural tissue engineering have opened promising avenues to apply fibrous tissue-engineered scaffolds to promote tissue regeneration and functional recovery.These scaffolds,known as neural scaffolds,are able to improve neural regeneration by playing two major roles,namely,by being a carrier for transplanted peripheral nervous system cells or biological cues and by providing structural support to direct growing nerve fibers towards the target area.However,successful implementation of scaffold-based therapeutic approaches calls for an appropriate design of the neural scaffold structure that is capable of up-and down-regulation of neuron-scaffold interactions in the extracellular matrix environment.This review discusses the main challenges that need to be addressed to develop and apply fibrous tissue-engineered scaffolds in clinical practice.It describes some promising solutions that,so far,have shown to promote neural cell adhesion and growth and a potential to repair peripheral nervous system injuries.

Optimization of clay material mixture ratio and filling process in gypsum mine goaf

Because there is neither waste rock nor mill tailings in the gypsum mine, and the buildings on the goaf of gypsum mine are needed to be protected, the research proposed the scheme of the clay filling technology. Gypsum, cement, lime and water glass were used as adhesive, and the strength of different material ratios were investigated in this study. The influence factors of clay strength were obtained in the order of cement, gypsum, water glass and lime. The results show that the cement content is the determinant influence factor, and gypsum has positive effects, while the water glass can enhance both clay strength and the fluidity of the filing slurry. Furthermore, combining chaotic optimization method with neural network, the optimal ratio of composite cementing agent was obtained. The results show that the optimal ratio of water glass, cement, lime and clay (in quality) is 1.17:6.74:4.17:87.92 in the process of bottom self-flow filling, while the optimal ratio is 1.78:9.58:4.71:83.93 for roof-contacted filling. A novel filling process to fill in gypsum mine goaf with clay is established. The engineering practice shows that the filling cost is low, thus, notable economic benefit is achieved.

A partition-type tubular scaffold loaded with PDGF-releasing microspheres for spinal cord repair facilitates the directional migration and growth of cells

The best tissue-engineered spinal cord grafts not only match the structural characteristics of the spinal cord but also allow the seed cells to grow and function in situ.Platelet-derived growth factor（PDGF） has been shown to promote the migration of bone marrow stromal cells;however,cytokines need to be released at a steady rate to maintain a stable concentration in vivo.Therefore,new methods are needed to maintain an optimal concentration of cytokines over an extended period of time to effectively promote seed cell localization,proliferation and differentiation.In the present study,a partition-type tubular scaffold matching the anatomical features of the thoracic 8–10 spinal cord of the rat was fabricated using chitosan and then subsequently loaded with chitosan-encapsulated PDGF-BB microspheres（PDGF-MSs）.The PDGF-MS-containing scaffold was then examined in vitro for sustained-release capacity,biocompatibility,and its effect on neural progenitor cells differentiated in vitro from multilineage-differentiating stress-enduring cells（MUSE-NPCs）.We found that pre-freezing for 2 hours at-20°C significantly increased the yield of partition-type tubular scaffolds,and 30 μL of 25% glutaraldehyde ensured optimal crosslinking of PDGF-MSs.The resulting PDGF-MSs cumulatively released 52% of the PDGF-BB at 4 weeks in vitro without burst release.The PDGF-MS-containing tubular scaffold showed suitable biocompatibility towards MUSE-NPCs and could promote the directional migration and growth of these cells.These findings indicate that the combination of a partition-type tubular scaffold,PDGF-MSs and MUSENPCs may be a promising model for the fabrication of tissue-engineered spinal cord grafts.

Scaffolds with anisotropic structure for neural tissue engineering

Nervous system injuries remain a great challenge due to limited natural tissue regeneration capabilities.Neural tissue engineering has been regarded as a promising approach for repairing nerve defects,which utilizes external biomaterial scaffolds to allow cells to migrate to the injury site and repair the tissue.Particularly,scaffolds with anisotropic structures biomimicking the native extracellular matrix(ECM)can effectively guide neural orientation and reconnection.Here,the advancements of scaffolds with anisotropic structures in the field of neural tissue engineering are presented.The fabrication strategies of scaffolds with anisotropic structures and their effects in vitro and in vivo are highlighted.We also discuss the challenges and provide a perspective of this field.

Cartilage oligomeric matrix protein enhances the vascularization of acellular nerves

Vascularization of acellular nerves has been shown to contribute to nerve bridging.In this study,we used a 10-mm sciatic nerve defect model in rats to determine whether cartilage oligomeric matrix protein enhances the vascularization of injured acellular nerves.The rat nerve defects were treated with acellular nerve grafting（control group） alone or acellular nerve grafting combined with intraperitoneal injection of cartilage oligomeric matrix protein（experimental group）.As shown through two-dimensional imaging,the vessels began to invade into the acellular nerve graft from both anastomotic ends at day 7 post-operation,and gradually covered the entire graft at day 21.The vascular density,vascular area,and the velocity of revascularization in the experimental group were all higher than those in the control group.These results indicate that cartilage oligomeric matrix protein enhances the vascularization of acellular nerves.

Increased connectivity of hiPSC-derived neural networks in multiphase granular hydrogel scaffolds

To reflect human development,it is critical to create a substrate that can support long-term cell survival,differentiation,and maturation.Hydrogels are promising materials for 3D cultures.However,a bulk structure consisting of dense polymer networks often leads to suboptimal microenvironments that impedes nutrient exchange and cell-to-cell interaction.Herein,granular hydrogel-based scaffolds were used to support 3D human induced pluripotent stem cell(hiPSC)-derived neural networks.A custom designed 3D printed toolset was developed to extrude hyaluronic acid hydrogel through a porous nylon fabric to generate hydrogel granules.Cells and hydrogel granules were combined using a weaker secondary gelation step,forming self-supporting cell laden scaffolds.At three and seven days,granular scaffolds supported higher cell viability compared to bulk hydrogels,whereas granular scaffolds supported more neurite bearing cells and longer neurite extensions(65.52±11.59μm)after seven days compared to bulk hydrogels(22.90±4.70μm).Long-term(three-month)cultures of clinically relevant hiPSC-derived neural cells in granular hydrogels supported well established neuronal and astrocytic colonies and a high level of neurite extension both inside and beyond the scaffold.This approach is significant as it provides a simple,rapid and efficient way to achieve a tissue-relevant granular structure within hydrogel cultures.

State-of-the-art MEMS and microsystem tools for brain research

Mapping brain activity has received growing worldwide interest because it is expected to improve disease treatment and allow for the development of important neuromorphic computational methods.MEMS and microsystems are expected to continue to offer new and exciting solutions to meet the need for high-density,high-fidelity neural interfaces.Herein,the state-of-the-art in recording and stimulation tools for brain research is reviewed,and some of the most significant technology trends shaping the field of neurotechnology are discussed.

Three-dimensional direct laser writing of biomimetic neuron interfaces in the era of artificial intelligence:principles,materials,and applications

The creation of biomimetic neuron interfaces(BNIs)has become imperative for different research fields from neural science to artificial intelligence.BNIs are two-dimensional or three-dimensional(3D)artificial interfaces mimicking the geometrical and functional characteristics of biological neural networks to rebuild,understand,and improve neuronal functions.The study of BNI holds the key for curing neuron disorder diseases and creating innovative artificial neural networks(ANNs).To achieve these goals,3D direct laser writing(DLW)has proven to be a powerful method for BNI with complex geometries.However,the need for scaled-up,high speed fabrication of BNI demands the integration of DLW techniques with ANNs.ANNs,computing algorithms inspired by biological neurons,have shown their unprecedented ability to improve efficiency in data processing.The integration of ANNs and DLW techniques promises an innovative pathway for efficient fabrication of large-scale BNI and can also inspire the design and optimization of novel BNI for ANNs.This perspective reviews advances in DLW of BNI and discusses the role of ANNs in the design and fabrication of BNI.