Regeneration Of Damaged Nerves Research Articles

Preparation of bilayer tissue-engineered polyurethane/poly-L-lactic acid nerve conduits and their in vitro characterization for use in peripheral nerve regeneration

BackgroundDue to loss of peripheral nerve structure and/or function resulting from trauma, accidents, and other causes, peripheral nerve injuries continue to be a major clinical problem. These injuries can cause partial or total loss of sensory, motor, and autonomic capabilities as well as neuropathic pain. PNI affects between 13 and 23 out of every 100,000 people annually in developed countries. Regeneration of damaged nerves and restoration of function after peripheral nerve injury remain significant therapeutic challenges. Although autologous nerve graft transplantation is a viable therapy option in several clinical conditions, donor site morbidity and a lack of donor tissue often hinder full functional recovery. Biomimetic conduits used in tissue engineering to encourage and direct peripheral nerve regeneration by providing a suitable microenvironment for nerve ingrowth are only one example of the cutting-edge methods made possible by this field. Many innate extracellular matrix (ECM) structures of different tissues can be successfully mimicked by nanofibrous scaffolds. Nanofibrous scaffolds can closely mimic the surface structure and morphology of native ECMs of many tissues.MethodsIn this study, we have produced bilayer nanofibrous nerve conduit based on poly-lactic acid/polyurethane/multiwall carbon nanotube (PLA/PU/MWCNT), for application as composite scaffolds for static nerve tissue engineering. The contact angle was indicated to show the hydrophilicity properties of electrospun nanofibers. The SEM images were analyzed to determine the fiber’s diameters, scaffold morphology, and endometrial stem cell adhesion. Moreover, MTT assay and DAPI staining were used to show the viability and proliferation of endometrial stem cells.ResultsThe constructed bilayer PLA/PU/MWCNT scaffolds demonstrated the capacity to support cell attachment, and the vitality of samples was assessed using SEM, MTT assay, and DAPI staining technique.ConclusionsAccording to an in vitro study, electrospun bilayer PLA/PU/MWCNT scaffolds can encourage the adhesion and proliferation of human endometrial stem cells (hEnSCs) and create the ideal environment for increasing cell survival.

En route to next-generation nerve repair: static passive magnetostimulation modulates neurite outgrowth

Objective. Regeneration of damaged nerves is required for recovery following nervous system injury. While neural cell behavior may be modified by neuromodulation techniques, the impact of static direct current (DC) magnetic stimulation remains unclear. Approach. This study quantifies the effects of DC magnetostimulation on primary neuronal outgrowth in vitro. The extension of neurites of dorsal root ganglia (DRG) subjected to two different low-strength (mT) static magnetic flux configurations was investigated. Main results. After 3 d of 1 h in-plane (IP) magnetic field stimulation, a 62.5% increase in neurite outgrowth area was seen relative to unstimulated controls. The combined action of in-plane + out-of-plane (IP + OOP) magnetic field application produced a directional outgrowth bias parallel to the IP field direction. At the same time, the diverse magnetic field conditions produced no changes in two soluble neurotrophic factors, nerve growth factor and brain-derived neurotrophic factor, released from resident glia. Significance. These results demonstrate the potential for DC magnetostimulation to enhance neuronal regrowth and improve clinical outcomes.

Improvement of sciatic nerve regeneration by multichannel nanofibrous membrane-embedded electro-conductive conduits functionalized with laminin

Multichannel structures in the design of nerve conduits offer potential advantages for regeneration of damaged nerves. However, lack of biochemical cues and electrical stimulation could hamper satisfactory nerve regeneration. The aim of this study was to simultaneously evaluate the effects of topographical, biological, and electrical cues on sciatic nerve regeneration. Accordingly, a series of multichannel nerve conduit was made using longitudinally-aligned laminin-coated poly (lactic-co-glycolic acid) (PLGA)/carbon nanotubes (CNT) nanofibers (NF, mean diameter: 455 ± 362 nm) in the lumen and randomly-oriented polycaprolactone (PCL) NF (mean diameter: 340 ± 200 nm) on the outer surface. In vitro studies revealed that the materials were nontoxic and able to promote cell attachment and proliferation on nanofibers and on fibrin gel. To determine the influence of laminin as biological and CNT as electrical cues on nerve regeneration, either of hollow PCL conduits, PLGA NF-embedded, PLGA/CNT NF-embedded or laminin-coated PLGA/CNT NF-embedded PCL conduits were implanted in rats. A new surgery method was utilized and results were compared with an autograft. The results of motor and sensory tests in addition to histopathological examination of the regenerated nerves demonstrated the formation of nerve fibers in laminin-coated PLGA/CNT NF-embedded PCL conduits. Results suggested that these conduits have the potential to improve sciatic nerve regeneration.Graphical abstract

Derivation of Stem Cell-like Cells From Spherical Culture of Astrocytes for Enhanced Neural Repair After Middle Cerebral Artery Occlusion.

Neural precursor cells (NPCs) tend to aggregate and develop into three-dimensional (3D) spheres, which in turn help maintain the stemness of the cells. This close relationship between spherical environments and cell stemness direct us to assume that 3D spheres of astrocytes (ASTs) may facilitate the acquisition of stem cell-like features and generate sufficient seed cells for the regeneration of neurons. In vitro results confirmed that mouse ASTs cultured on agarose surfaces spontaneously formed cell spheres and exhibited molecular features similar to stem cells, particularly capable of further differentiating into neurons and forming functional synaptic networks with synchronous burst activities. RNA-sequencing results revealed the similarity between AST-derived stem cells (A-iSCs) and NPCs in global gene expression profiles. The potency of A-iSCs in repairing neural injuries was evaluated in a mouse model of middle cerebral artery occlusion. It was observed that the transplanted A-iSCs expressed a series of markers related to neural differentiation, such as NeuN, Tuj1, and Map2, indicating the conversion of the transplanted A-iSCs into neurons in the scenario. We also found that the injured mice injected with A-iSCs exhibited significant improvements in sensorimotor functions after 8 weeks compared with the sham and control mice. Taken together, mouse ASTs form cell spheres on agarose surfaces and acquire stem cell-associated features; meanwhile, the derived A-iSCs possess the capacity to differentiate into neurons and facilitate the regeneration of damaged nerves.

Nerve Regeneration: Human Periodontal Ligament‐ and Gingiva‐derived Mesenchymal Stem Cells Promote Nerve Regeneration When Encapsulated in Alginate/Hyaluronic Acid 3D Scaffold (Adv. Healthcare Mater. 24/2017)

Sahar Ansari and co-workers fabricate a 3D delivery system based on alginate/hyaluronic hydrogel in article number 1700670. The Encapsulated Periodontal Ligament Stem Cells and Gingival Mesenchymal Stem Cells demonstrate positive β-tubulin III staining after four weeks of in vitro differentiation in the presence of recombinant human nerve growth factor (β-NGF) and high levels of gene expression related to neurogenic differentiation. This system has the potential to be used for cell therapy in the repair or regeneration of damaged nerves.

Human Periodontal Ligament- and Gingiva-derived Mesenchymal Stem Cells Promote Nerve Regeneration When Encapsulated in Alginate/Hyaluronic Acid 3D Scaffold.

Repair or regeneration of damaged nerves is still a challenging clinical task in reconstructive surgeries and regenerative medicine. Here, it is demonstrated that periodontal ligament stem cells (PDLSCs) and gingival mesenchymal stem cells (GMSCs) isolated from adult human periodontal and gingival tissues assume neuronal phenotype in vitro and in vivo via a subcutaneous transplantation model in nude mice. PDLSCs and GMSCs are encapsulated in a 3D scaffold based on alginate and hyaluronic acid hydrogels capable of sustained release of human nerve growth factor (NGF). The elasticity of the hydrogels affects the proliferation and differentiation of encapsulated MSCs within scaffolds. Moreover, it is observed that PDLSCs and GMSCs are stained positive for βIII-tubulin, while exhibiting high levels of gene expression related to neurogenic differentiation (βIII-tubulin and glial fibrillary acidic protein) via quantitative polymerase chain reaction (qPCR). Western blot analysis shows the importance of elasticity of the matrix and the presence of NGF in the neurogenic differentiation of encapsulated MSCs. In vivo, immunofluorescence staining for neurogenic specific protein markers confirms islands of dense positively stained structures inside transplanted hydrogels. As far as it is known, this study is the first demonstration of the application of PDLSCs and GMSCs as promising cell therapy candidates for nerve regeneration.

Effects of medium flow on axon growth with or without nerve growth factor

Axon growth is a crucial process in regeneration of damaged nerves. On the other hand, elongation of nerve fibers in the epidermis has been observed in skin of atopic dermatitis patients. Thus, regulation of nerve fiber extension might be an effective strategy to accelerate nerve regeneration and/or to reduce itching in pruritus dermatosis. We previously demonstrated that neurons and epidermal keratinocytes similarly contain multiple receptors that are activated by various environmental factors, and in particular, keratinocytes are influenced by shear stress. Thus, in the present study, we evaluated the effects of micro-flow of the medium on axon growth in the presence or absence of nerve growth factor (NGF), using cultured dorsal-root-ganglion (DRG) cells. The apparatus, AXIS™, consists of two chambers connected by a set of microgrooves, through which signaling molecules and axons, but not living cells, can pass. When DRG cells were present in chamber 1, NGF was present in chamber 2, and micro-flow was directed from chamber 1 to chamber 2, axon growth was significantly increased compared with other conditions. Acceleration of axon growth in the direction of the micro-flow was also observed in the absence of NGF. These results suggest that local micro-flow might significantly influence axon growth.

Alginate-matrigel microencapsulated Schwann cells for inducible secretion of glial cell line derived neurotrophic factor

Controlled expression of glial cell line derived neurotrophic factor (Gdnf) can be integrated in the development of a system for repair of injured peripheral nerves. This delivery strategy was demonstrated via inducible Gdnf from microencapsulated cells in barium alginate. The Schwann cell line RT4-D6P2T was initially modified utilizing an ecdysone-based stable transfection system to produce RT4-Gdnf cells. During construct preparation, it was found that C6 cells (where Gdnf cDNA was isolated) make three Gdnf transcript variants. Additionally, the importance of 5′ untranslated region to drive biologically-functional Gdnf synthesis was shown. Encapsulation of RT4-Gdnf in 1% alginate was then performed. It was determined that cells were able to survive at least 1 month in vitro using starting densities of 20, 200 and 2000 cells/capsule and barium ion concentrations of 10, 50, 100 and 200 mM. Most importantly, encapsulated cells secreted exogenous Gdnf upon ponasterone A induction. Mixture of basement membrane extract Matrigel™ to alginate promoted increased proliferation, cell spreading and Gdnf release. Finally, compression tests showed that cell-loaded microcapsules fractured at 75% diameter compression with 38 kPa of stress. Regulated Gdnf release from these microcapsules in vivo may potentially aid in the regeneration of damaged nerves.