Repair Of Adult Rat Spinal Cord Injury By Multichannel PLGA Scaffold Combined With ASCs And MSCs Transplant

Objectives: To investigate whether PLGA scaffold with smaller diameter micro-catheters promotes greater functional recovery over scaffold with larger diameter micro-catheters.To determine the therapeutic effects of a combinatorial strategy for the repair of complete spinal cord transection in adult rat.Methods: We fabricated two multiple-channel PLGA scaffolds with different 3D structures and implanted into a 3 mm-wide transection gap in the thoracic spinal cord of adult rats.After the transplantations,at different time point,the locomotor functional improvement was assessed using the BBB Scale.Eight weeks post the transplantations,the area of cystic cavity were measured among all groups.The number of axon regeneration and the number of survival neurons at the lesion/graft area were evaluated by immunohistochemistry for NF and MAP 2.Myelination of axons at the lesion/graft area were tested by immunohistochemistry for MBP and confirmed by electron microscopy.Multichannel polymer scaffolds seeded with activated Schwann cells(ASCs)and/or rat bone marrow-derived mesenchymal stem cells(MSCs)were grafted acutely into a 3 mm-wide transection gap in the thoracic spinal cord of adult rats.BBB scoring and climbing were tested at 3 days,and each week after the operation.Somatosensory evoked potential and motor evoked potential were tested for each group at 2,4,and 8 weeks after the operation.Four and eight weeks after the first surgery,HE and Nissl staining were performed to asseess the cystic cavity area and immunohistochemistry for neural cell special antigen were performed to detect the neural differentiation of MSCs,the regeneration of nerve tissue,the Myelination of new axons and the expression of neurotransmitters in the lesion area.Eight weeks after the first surgery,some rats in the AS+MS group received the retransection in the middle of the lesion/graft site.Results: Eight weeks after the scaffolds transplantation,according to the effect on the locomotor functional improvement,the number of regenerating axons,the number of survival neurons and the area of the cystic cavity at the lesion/graft area,scaffold 1 was more suitable for transplantation to promote the recovery of spinal cord injury.At 4 weeks post transplantation and thereafter,rats that had received scaffolds seeded with ASCs and MSCs showed significant recovery of nerve function as indicated by BBB score and electrophysiological test results.This was not observed in groups receiving the empty scaffold or lesion only.Immunohistochemistry(IHC)analysis at 4 and 8 weeks after transplantation revealed that the implanted MSCs at the lesion/graft site,survived and differentiated into neuron-like cells and formed contacts with host neurons.Robust bundles of regenerated fibers were observed in the lesion/graft site in ASCs and MSCs co-transplantation rats,and neurofilament 200(NF)immunostaining confirmed that these fibers are neural tissues.In addition,some myelin basic protein(MBP)-positive myelin sheathes were also observed at the lesion/graft site and confirmed by electron microscopy.Importantly,in addition to expressing mature neuronal markers,sparse MSC-derived neuron-like cells also expressed synapsin,in the injury area of ASCs and MSCs co-transplantation rats.Conclusions: The results suggest that PLGA scaffold integrated effectively with the host tissue of the spinal cord by bridging rostral and caudal stumps.Functional recovery was achieved in severe SCI rats through ASC-induced and MSC-derived neuron-like cells making contacts with host neurons and promoting regeneration of axons,which in overall created local neural network.This donor-derived restoration of neural connectivity was verified by retransection in the middle of the lesion/graft site at 8 weeks after the first operation.The BBB score of rats that received retransection dropped to complete SCI level.The multichannel PLGA scaffold effectively bridged the lesion gap in completely transected SCI rats,and co-implantation strategy promoted MSCs survival and differentiation into neuron-like cells,leading to enhanced regeneration of axons and functional recovery.