Clinical Outcome And Prognosis Of Chronic Cervical Spinal Cord Compression And Autophagy In Spinal Motor Neurons

ObjectiveAsymptomatic cervical degenerative spinal cord compression(A-SCCC)is quite common in the general population,and it may develop to cervical spondylotic myelopathy(CSM)which will seriously disturb patients' life and work,and will bring heavy burden to family and society.In this study,we tried to find out the risk factors of A-SCCC turning to CSM and to predict how it progresses.Under the guidance of syndrome differentiation theory in traditional Chinese medicine(TCM),we herein tried to establish a unique comprehensive treatment of TCM,to intensify the prevention efforts of this kind of person,and to control risk factors which are associated with CSM,thus reducing the incidence of CSM.Autophagy is a highly regulated,self-destructive process which plays important roles in maintaining cellular homeostasis and providing energies for cellular metabolism to help cells survive under physiological or pathological conditions.With the spinal cord injury(SCI)animal model and spinal motor neurons(SMNs)model established,we tried to explore the role of autophagy and the secondary response after injury in the SMNs,and to explore the related molecular mechanism,thus providing theoretical basis for clinical treatment.Methods1.The risk fators of A-SCCC and CSM,including cervical stability,high signal intensity of spinal cord in MRI,types of compression,compression ratio of the spinal cord,congenital cervical stenosis and numbers of compression,are retrospective collected in our study,and logistic regression are used to find out the risk factors of CSM;2.Under the principles of control dynamic compression and reinforcing kidney and activating blood circulation/Bu Shen Huo Xue,we performed cervical traction,decoction,exercise to treat A-SCCC,with the neck disability index(NDI)recorded during follow-up time,to observe whether A-SCCC will process to CSM;3.The SCI animal model was built by PinPointTM precision impactor,to validate the reliability and the replication of the model.Later,we used western blot,immunohistochemical staining and electron microscopic to detect whether LC3 protein,a autophagical marker,and autophagical structures exist in the spinal cord or not;4.By choosing proper gestational rats,optimizing the diseecting steps,and immunpanning procedure with p75 antibody,we tried to optimize the isolation and purification of SMNs;5.Simulated ischemia and anoxic conditions,we use the gas incubator to build a hypoxic and glucose-free SMNs model/oxygen glucose Deprivation model.Western blotting,immunohistochemical staining and electron microscopic are used to detect the trends of LC3 protein and autophagical structures;6.By adding the autophagical inhibitor,Bafilomycin A1,to intervent neurons during oxygen glucose deprivation,we later used CCK-8 to test the cell viability,so that we can judge the role of autophagy to be protective or detrimental.Later,we tested the role of isorhynchophylline in SMNs during oxygen glucose deprivation,and explore the possible mechanism of regulating autopahgy and the underlying mTor signaling pathways.Resulis1.Chronic cervical spinal cord compression can be found in different ages and there are no gender differences,but most of them are middle and elderly people.The onset sites of chronic cervical spinal cord compression segment are C5/6 and C4/5.Whether chronic cervical spinal cord compression will turn to CSM or not,is related to the cervical stability,high signal intensity of spinal cord,types of compression,but are not in correlation with compression ratio of the spinal cord,congenital cervical stenosis and numbers of compression number.No cervical instability and high signal intensity was protective factors for cord compression.2.Patients with A-SCCC under conservative treatment were most found single disc herniation in C5/6 segments,and risk fators,such as calcification or ossification,high signal intensity,cervical instability were in a low proportion.Under the principles of control dynamic compression and reinforcing kidney and activating blood circulation/Bu Shen Huo Xue,we used cervical traction,decoction,exercise to treat A-SCCC and had good effects.There was low recurrence and no one develop to CSM during the follow-up period.These results suggest that the chronic asymptomatic cervical degenerative spinal cord compression may only be a clinical phenomenon,but it still need to take a closely observation;3.Through the observation of the BBB score to evaluate the behavior of rats,and pathological histology to observe the pathological changes,PinpointTM instrument can copy a high precise,controllable and stabilized model of SCI,which enables us to study the pathological mechanism of SCI.At the same time,we found that autophagy activated in spinal cord tissue after SCI.4.We have successfully developed optimized methods for rapidly dissecting spinal cords and harvesting SMNs with high survival and purity from embryonic rats:14-16 days of fetal rats is preferred;immunopanning with p75 can effectively improve the purity of cultured SMNs in vitro.5.Successfully build a hypoxic glucose-free SMNs model,and the cell viability decreased or even death as time goes on;Autophagy is activated,and reach a peak in 5 hours;6.Autophagy protects SMNs;isorhynchophylline protects SMNs through activation of autophagy,which may be associated with mTor signaling pathways.Conclusion Whether chronic cervical spinal cord compression will develop to CSM or not is associated with cervical stability,high signal intensity of spinal cord in MRI,types of compression.The existence of the risk factors that could lead the spinal cord tissue to ischemia and hypoxia,eventually cause spinal cord injury and clinical symptoms.Under the principles of control dynamic compression and reinforcing kidney and activating blood circulation/Bu Shen Huo Xue,we used cervical traction,decoction,exercises to treat A-SCCC and had good effects.It suggested that maintaining the cervical stability and spinal canal environmental stability,improve spinal cord ischemia and oxygen deficit may be helpful to repair the spinal cord.Simulated pathology of spinal cord injury,under the condition of glucose-free and hypoxia or injury the spinal cord,cell viability was decreasing or even necrosis;under the pathological condition,autophagy plays a role of protection in SCI animal model and hypoxia SMNs model.Isorhynchophylline can activate autophagy,to maintain energy balance of the cell to resist low oxygen and glucose-free condition,which may be associated with the mTor signaling pathway,it also suggests that improving the cell low nutritional status may have protective effect on SMNs.