Construction Of Electroactive Hydrogels And Its Application In Nerve Injury Repair

The nervous system consists mainly of the central nervous system and the peripheral nervous system.It functions by transmitting electrical signals through a complex neural network to coordinate the activities of various parts of the body.Injuries to both the central nervous system and peripheral nervous system injuries may destroy the electrical signal conduction between the brain and the target tissue,resulting in the loss of patient function.Therefore,restoring the electrical signal conduction of nerve tissue and re-establishing signal transmission with the remote target tissue is critical for nerve injury repair.However,the importance of nerve conduction is often overlooked in current treatment strategies which include surgical treatment,drug therapy,cell transplantation,exosome therapy,and tissue engineering scaffold implantation,resulting in unsatisfactory therapeutic effects.To solve this problem,this study combined a conductive polymer（polypyrrole,ppy）with extracellular matrix hydrogel（methacrylated gelatin,GM）to construct a conductive electroactive hydrogel.This GM network conferred biocompatibility,tissue-like softness,degradability,and tissue and cell adhesion,while the ppy network provided an electroconductive electrical activity to the hydrogels.Here,we use this hydrogel to restore electrical signal conduction in diabetic peripheral neuropathy and spinal cord injury（SCI）.Besides,the molecular mechanism of the conductive electroactive hydrogel to promote the repair of peripheral nerve injury and the SCI was further studied.Although conductive electroactive hydrogels effectively enhance neuronal and axonal regeneration,their efficacy is compromised by host recognition and the subsequent foreign body immune responses,which cannot attenuate or even aggravate the early secondary inflammation after acute SCI.In response to this problem,this study has also loaded bone marrow stem cell（BMSC）-derived exosomes in conductive electroactive hydrogels to reduce the early inflammatory response induced by hydrogels.The immune regulation effect of BMSC-derived exosomes and the nerve regeneration effect of the conductive electroactive hydrogel can synergistically promote the repair of SCI.The specific research content is as follows:1.Conductive electroactive hydrogel dressing mediates tissue repair by promoting peripheral nerve regenerationWe mixed oxychondroitin sulfate-ppy（OCS-P）conductive particles with GM prepolymer to construct the GM-OCS-P conductive electroactive hydrogel dressing by ultraviolet light（UV）irradiation.GM-OCS-P conductive hydrogel dressing possesses a three-dimensional porous structure and skin tissue-like conductivity（2.5 m S/cm）.In vitro experiments have found that the conductive hydrogel can promote the migration of PC12 cells,axon-related genes and proteins（including neurofilament protein,NF;growth associated protein 43,GAP43;synapsin I,Syn-1;postsynaptic density protein 95;PSD95）expression,ultimately inducing axon growth.A diabetic skin wound model which is a type of difficult-to-heal wounds caused by severe peripheral neuropathy,was constructed for further in vivo study.Conductive hydrogel dressings can promote the healing of diabetic skin wounds on the 14th day.Histological examination found that the regeneration of nerves in the repaired skin is significantly increased in the conductive hydrogel group.Therefore,the conductive electroactive hydrogel dressing can mediate the rapid healing of diabetic wounds by promoting peripheral nerve regeneration.2.A study of the mechanism by which conductive electroactive hydrogels promotes peripheral nerve injury repair.We further studied the mechanism by which GM-OCS-P conductive hydrogels encourages peripheral nerve injury repair.Immunofluorescence images show that conductive hydrogel increases the intracellular Ca²⁺concentration by 3.73±1.62 times,which subsequently improves related protein phosphorylation in mitogen-activated proteinkinase kinase（MEK）/extracellular regulated protein kinases（ERK）and phosphatidylinositol3-kinase（PI3K）/protein kinase B（AKT）pathway,and ultimately promotes the expression of axon-related genes and proteins and axon growth.Therefore,we uncovered mechanistically that conductive hydrogel promotes nerve regeneration by regulating the Ca²⁺concentration,which then activates the downstream PI3K/AKT and MEK/ERK signaling pathways.3.Exosomes loaded conductive electroactive hydrogel promote SCI repairIn this study,BMSC-derived exosomes were immobilized in the TA-doped GM/PPy（GMP）hydrogel network to form a GM/PPy/Exos（GMPE）hydrogel,which is used for SCI repair.The hydrogel possesses suitable mechanical properties（1056.0±133.1 Pa）and excellent electrical properties(1.49×10^-3 S/cm)which are similar to that of central nervous tissue.Exosomes sustained release time for the GMPE hydrogel lasted up to 14 days.In vitro studies found that exosomes-loaded hydrogels can modulate microglia M2 polarization via the NF-κB pathway,and synergistically enhance neuronal and oligodendrocyte differentiation of neural stem cells（NSCs）while inhibiting astrocyte differentiation.Meanwhile,released exosomes and electrical characteristics of GMPE hydrogel can also increase axon outgrowth via the synergistic activation of the PTEN/PI3K/AKT/m TOR pathway.Furthermore,exosomes combined with electroconductive hydrogels significantly decrease the number of CD68-positive microglia,enhance local neurogenesis,and promote axonal regeneration,resulting in significant functional recovery at the early stage in an SCI mouse model.