Research Of Axon Stretch Growth For Peripheral Neural Repair And Neural Interface

Because of rising traumatic accidents and diseases,the number of patients suffer from peripheral nerve injury is increasing.If they fail to carry out effective neural repair and rehabilitation therapy,it will affect the patient's motor and sensory functions,and bring much trouble to their life.There are two kinds of treatments for peripheral nerve injury.If the patient's limbs are intact,the surgical procedure can be performed to suture the disconnected nerves to restore the natural neural function.As for amputees,neural interface technology can be used to splice nerves and electrical wires together in a way that allows them to control an artificial limb as if it was a natural extension of the body.Neural repair and neural interface technology based on tissue engineering have shown promising prospects in the treatment of nerve injury.However,both of the two treatments need autologous nerve to stimulate axonal regeneration and extension into target tissues,which are limited by the supply of donor nerves.In the dissertation,the main contributions were as follows:We had designed and fabricated a new miniaturized bioreactor system for stretch growth of integrated axon tracts.The device was composed of two parts:the control system and mechanical system.Neurons were firstly placed on two adjoining substrates and formed new synaptic connections between each other.Then the axon bundles across the border between the top and bottom membranes were stretched in a stepwise fashion by a microstepper motor system.After several days' stretch,the axon tracts could reach specific length that can be used to create living nervous tissue constructs.By measuring the linear displacement with 0.5 ?m,1 ?m and 2 ?m step,we found the axon stretch system taking 1?m every step was the most appropriate.The substrate of axon expansion chamber was made of Aclar film,which was a transparent film,suitable for axon stretch growth after surface modification.In order to get more appropriate neuronal culture and stretch conditions,the Aclar films were respectively treated with PLL,PDL,laminin,type 1 rat-tail collagen,both PDL and lamini,both PDL and collagen.Through the statistical analysis of the DRGs cultured in the different groups,we found the elongator substrates coated by PDL,both PDL and rat-tail collagen,or both PDL and laminin could maximize the number of axons bundles.And they were the best methods to modify the Aclar film for axon stretch growth.The DRGs were cultured in the bioreactor,and the bridging axons adapted to the stretch by increasing their length from 500 ?m to become about 6 mm long over 7 days of stretch growth.Immunocytochemical analysis confirmed that major cytoskeletal constituents were present along axons.Then we cultured DRGs in a 3 dimensional(3D)fashion and found the axons also grew very well in the bioreactor,which provided some new thoughts for the encapsulation of stretch growth nervous tissue.There is no direct electrophysiological examination devices for axons under extreme stretch growth conditions.In order to investigate the ability of DRG neurons to generate and convey action potentials after a period of rapid axon stretch growth in vitro,and what stimulating parameters can evoke action potential,we have used computational tools to simulate the stretch growth neurons.In the simulation,we had analyzed the spike amplitude and conduction velocity in both static culture neuron and stretch growth neuron.And we also analysed the sensitivity of spike propagation in the stretch growth neuron to different stimulating frequencies and stimulating voltages.Finally,we had investigated how t-junction influenced the spike propagation in neurons.It could provide some guidelines for the electrophysiological experiments in future.