| Peripheral nerve injury is a relatively common medical problem caused by trauma,tumor resection,or reconstructive surgery.Due to the complexity of the nervous system,restoring the function of injured nerves or repairing the damage associated with neurodegenerative diseases remains a major challenge in the field of biomedicine.Now,neurorrhaphy is considered the clinical gold standard in the treatment of nerve spaces smaller than 1cm,and autologous nerve transplantation is a common treatment for nerve injuries of exceeding 1 cm.However,the Limits of nerve transplantation,donor site morbidity,possible neuroma formation,and immune response are some of the key issues limiting autologous nerve grafts as a therapeutic method.Despite nerve tissue repair faces the inherent challenges,the effective therapeutic strategies are not unreachable.With the more realizing of nervous system,and the progress of engineering and materials science,which formed the basis of the field of neural tissue engineering.The strategies of neural tissue engineering utilize biomaterials to influence the physical and chemical environment at the site of nerve injury or disease.By controlling the spatiotemporal distribution of different types of signals,a regenerative environment can be constructed to restore the complex physical and biochemical structure of neural tissue.The inherent characteristic of neurons is wired to transmit electrochemical signals throughout the nervous system,and they are highly influenced by electrical stimulation.Therefore,the preparation of scaffold materials with appropriate chemical,mechanical and electrical activity is the key to the success of neural tissue engineering.Clinical electrical stimulation is a non-drug treatment method,which is safe,economical and convenient for the prevention,delay and treatment of diseases,and has important social and economic value.It has been widely used in pain relief,wound healing and neuromuscular retraining.In the field of neuroscience,it has been shown that electrical stimulation is an effective signal to stimulate the proliferation or differentiation of various cell types.And electrical impulses can stimulate the directional growth of neurites to fill the gaps of damaged nerve tissue.Compared with the chemical factor method,electric stimulation has incomparable advantages,such as no immune response,controllable parameters,minor damage,easy implementation,localization induction,and can be combined with other induction methods.However,most of the clinical or research uses commercial electrical stimulator as the electrical stimulation output device.These devices still use invasive electrodes,which increase the risk of wound pain and infection,and are not suitable for repairing tissues and organs in the body.Therefore,New electrical stimulation modes with implantable,low-cost,and non-invasive are urgently required for nerve regeneration devices.Above the discussion,in this thesis,we have studied the electroactive scaffold materials combined with three electrical stimulation modes regulate the fate of stem cell differentiation.This paper mainly includes the following four parts:1.Self-powered electrical stimulation for enhancing neural differentiation of r ADSCs on PBF flexible antimicrobial film.PBF flexible film with superior mechanical properties,transparent,excellent electrical conductivity,antibacterial and biocompatibility was prepared by vacuum filtration method as nerve scaffold material.A human-flap-driven self-powered triboelectric piezoelectric nanogenerators（TPNG）with an output of 100 V and+8.5μA was built as the electrical simulation power source to replace the large electric stimulation instrument,and the TPNG was established by using biodegradable piezoelectric fish gelatin and BC as two friction layers.A promising strategy for combining a biodegradable,low-cost,small-sized and large-output self-powered TPNG and electroconductivity-improved PBF flexible film to build a self-powered electrical simulation system for r ADSCS neural differentiation.Under TPNG electrical simulation,the q PCR result reveals that the expression of MAP2 and GFAP on the PBF-2 flexible film was respectively enhanced by 4.06-fold and 0.84-fold over that without electrical simulation.The results confirmed that the self-powered neural repair system could have achieve the directed differentiation of r ADSCs into neuro-like cells without any inducible factors.This work provides a promising application of a self-powered,wearable,electrical stimulation TPNG system to assist nerve regeneration.2.Piezoelectric PVDF nanopillar array induced neuron-like differentiation of stem cells via wireless electrical stimulation.Piezoelectric scaffold as a non-invasive mode of signal stimulation in response to demand.A PVDF nanopillar array with a diameter of 200 nm,a height of 500 nm,and a column spacing of 450 nm was prepared by hot pressing using AAO template.The piezoelectric properties of nanopillar array films are significantly higher than those of PVDF films by PFM and ultrasonic piezoelectric characterization.Therefore,nanopillar array was selected as a neural scaffold material to regulate the neural differentiation of rat bone marrow mesenchymal stem cells（rBMSCs）.Ultrasonic stimulation can produce deep penetration into biological tissue.Moreover,nanopillar array can be stimulated by ultrasonic stimulation to stimulate the neural differentiation of stem cells by offering a stronger piezoelectric electrical signal.Detection of gene expression and protein levels showed that the typical neuronal markers Tuj1 and MAP2 after 21 days culture of rBMSCs on nanopillar array were significantly upregulated under ultrasonic stimulation.In a certain range of ultrasonic stimulation,the higher ultrasonic wattage,the higher MAP2 expression.However,Nestin and GFAP were not detected in all the samples,which suggesting that rBMSCs on nanopillar array did not transform into neural stem cells（NSCs）and not conducive to glial differentiation under ultrasonic stimulation.This opens up new avenues for non-contact,controlled nerve regeneration therapy,and it also holds great promise for the development of non-invasive nerve regeneration devices.3.Degradable piezoelectric hydrogels promote neural differentiation of NSCsThe problem in the piezoelectric field is that high voltage responsive polymers are not easy to degrade,and the biodegradable piezoelectric materials themselves are not high piezoelectric.Based on this,we have adopted a novel and simple strategy to design and prepare different shapes of cellulose piezoelectric hydrogels（ACH）by simple pre-stretching and acid treatment process.The piezoelectric properties of ACH were verified by PFM and ultrasonic piezoelectric methods.And the piezoelectric properties of prestretched ACH（1 V）were significantly higher than that of unprestretched ACH（0.5 V）.In order to increase the biocompatibility and piezoelectric properties of ACH,PDA/ACH hydrogel was prepared by in situ polymerization of dopamine on the surface of ACH.This hydrogel has good piezoelectric property（the ultrasonic piezoelectric of pre-stretched PDA/ACH hydrogel is 1.5V）,soft surface and good biocompatibility.In addition,NSC was used as seed cells to explore the role of hydrogels in regulating the fate of stem cells.The electrical signals generated by ultrasonic deformation of PDA/ACH hydrogel were used to stimulate and promote cell differentiation.Only Tuj1 and GFAP genes were expressed after 7 days of culture on TCP under ultrasound and without ultrasound.However,after 3 days,Tuj1,MAP2 and GFAP were significantly expressed on PDA/ACH piezoelectric hydrogel.In particular,after 7 days of culture,compared with TCPs,the Tuj1 and MAP2 genes of cells cultured on prestretching PDA/ACH increased by 164-fold and 7.9-fold,under 400 W ultrasonic condition,respectively.Thus,this biodegradable piezoelectric hydrogel creates the possibility for the clinical application of non-invasive electrically stimulated nerve regeneration.4.Highly-specific differentiation of MSCs into neurons directed by local electrical stimuli wirelessly triggered by electromagnetic inductionTransdifferentiation of mesenchymal stem cells（MSCs）into neurons provides a practical way alternative to NSCs for neurodegenerative diseases,given the limited resources of autologous NSCs as well as the ethical and immunogenicity issues of allogeneic NSCs.However,it is challenging to get mature neurons through MSCs differentiation.In this work,a wirelessly-triggered local electrical stimulation system was established to specifically induce neurons differentiation from rBMSCs,by coupling a highly conductive and flexible MWCNT membrane and a rotating magnetic field.At absence of nerve inducing factors,the differentiation was realized by the localized electrical stimuli mediated from electromagnetic induction.A highly-specific neuronal differentiation was achieved without the presence of neuroglia cells.Functional neuronal firings were revealed by rapid spontaneous[Ca2+]i-transient peaks under the action of neurotransmitters.The potential of this discovery as a novel stem cell therapy for neurodegenerative disease was further demonstrated in vivo,by its implementation in the brain defects of Wistar rats,where the successful differentiation of exogenous rBMSCs to nerve tissue accelerated the brain recovery.In this study,electrical signals generated by electromagnetic induction was used as the source of electrical stimulation.Compared with ultrasonic stimulation,this method is softer,avoiding the damage of surrounding cells due to ultrasound,and achieving a good effect of nerve differentiation.In summary,this study proposes the regulation of neural differentiation in stem cells based on electrical signals mediated by outfield-driven nanostructures.The material construction,the construction of the field regulation device,especially the demonstration and characterization of the differentiation process were verified.This kind of stem cell regulation based on nanomaterials and radio signal stimulation will have an important prospect in the clinical treatment of neurodegenerative diseases. |
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