| [Objective]Observe the effect of Tuina therapy to nerve functional recovery of SNI rats through behavioristics,morphology. Then probe the biological mechanisms of Tuina therapy in SNI through the content of CGRP in Spinal cord to peripheral motor pathways.[Methods]Use SD rats as experimental animals which were divided into five groups, they were normal group, sham operation group, model group, model control group and tuina group,the last three groups were made the SNI model by nerve clamping method. Then rats of tuina group were treated by "Massage massage simulator"which treated by imitating Stiring,Pointing and Kneading Manipulation.Acupoints selected Yinmen, Chengshan and Yanglinquan, each point worked 1 minute.After the intervention, using the Plane test to evaluate the recovery of motor function of rat.The morphology of spinal cord, nerves and gastrocnemius were analyzed to find the massage for evidence of nerve repair;Finally, testing the content of CGRP in spinal cord-injured nerve-neuromuscular junction to promote the mechanism of regeneration and repairation after peripheral nerve injury.[Results]1 Results of behavioristics——Tuina can improve the angle of plate test in SNI rats.Results of Plane test:7 days after operation,the score of the Plane test significantly lower than the blank control group and sham-operated group (p< 0.05). After 20 treatments, scores of Tuina group, model group and model control group were all higher than before and the score of Tuina group was the highest, which higher than model group and model control group, but still significantly lower than blank control group (p< 0.05).2 Results of morphology——Tuina can improve the microstructure of rat spinal cord anterior horn, nerves and muscles.2.1 The result of HE staining of Spinal cord,Nissl staining and Nissl cell dyed number of Motor neurons.7 days of operation,HE staining showed that neurons of normal group and sham operation group are completion and former;While the quantity and form of the neuron in spinal cord of model group are abnormal, such as disordered arrangement, the number of neuron was decreased and cavitations were observed.The Nissl staining show that Nissl in normal group and sham operation group are distributed uniformly and dyed dark blue uniformly;While the motor neuron of model group are edema,The number of Nissl body were dyed light and reduced and even dissolved.Nissl dyeing cell number of Motor neurons of model group was much lower than normal group and sham-operated group (p<0.05).20 times after treatment, the normal group, sham operation group of HE staining is similar compared with that in 7 days after the operation.The pathological changes of the model control group is similar to those in model group.There are slightly irregular neurons in Tuina group, with swelling cell body, the cell nucleus is at the edge part of the cell, and some of the neurons are regeneration.Nissl dyed showed that morphological in normal group and sham-opertation group is similar to those in 7 days after;after the autologous recovery, Model group and control group had some improvement.Diameter of the motor nuron is slightly decreased than those of seven days, but solution is still visible, degeneration can be seen in the cytoplasm vacuoles.Most motor neuron in Tuina group was seen regeneration after Nissl body dissolved, dyed deep and uniform, diameter is slightly higher than those in normal group.Nissl dyeing cells number in motor neurons of model group rats were significantly lower than the normal group and sham-operation group (P< 0.05), The dyed cells number of Tuina group was obviously higher than those in model group and control group (P< 0.05), but still lower than normal group and sham-operation group (P< 0.05).2.2 HE staining and Masson staining results in sciatic nerve injuryafter 7 days of opertaion, HE staining in normal group and sham-operation group showed that nerve fiber and myelin sheath are complete;In Model group,the nerve fibers wew scattered and axon collapsed, schwann cells were proliferation.Masson staining showed that,in normal group and sham-operation group,the nerve fiber and myelin displayed neatly and tightly, schwann cell nucleus were formed normaly.The nerve membrane were arranged continuosly; In Model group, nerve fibers were scattered and axons were disintegration and part of the myelin sheath were vacuolated degeneration.Schwann cell nucleus were fragmented, neuron membrane is discontinuous.20 times after treatment, the normal group, sham-operation group, model group and control group of nerve fiber HE dyeing were similar than those in 7 days afteroperation; Nerve fiber arrangement of Tuina group is neat, the axon is clear, schwann cells had normal structure.Masson staining showed that nerve fibers in Tuina group is arranged neatly, such as the myelin sheath, schwann cells have normal structure and the same as the neuron membrane.2.3 HE staining of the gastrocnemius muscle, muscle cell diameter7 days after operation,the muscle cell in normal group and sham-operation wre dyeing uniformitly, diameter of all cells is consistent, arranged neatly, the gap between cells is small, muscle cell nucleus can be seen clearly.ln model group, muscle cell can be seen dyeing irregularity, cell diameters have different sizes, most diameter of cells are reduced, the gap between muscle cells is increased,cells arranged loosely, the size of cells were contraction, at the same time the number of nucleus is increased and less cleared.According to the diameter of muscle cell,model group rats were significantly less than normal group and sham-operation group (P< 0.05).20 times after treatment, the normal group and sham-opertaion group is similar to those in 7 days after operation;Gastrocnemius muscle cells dyeing were nonuniform,and the gap between muscle cells is decreased compared with 7 days and atrophied obviously.ln tuina group, the muscle is dyed neatly and the diameter of muscle cell is slightly lower than normal group,and the gap between muscles is increased than normal group and cell arrangement is neat, there are slight contraction phenomenon, nucleus can be seen clearly.According to Muscle cell diameter, the muscle cells of rats are bigger than the diameter of the model group and control group (P< 0.05), but still less than the normal group and sham-operation group (P<0.05).3 immunohistochemical results——Tuina can obviously increase the CGRP in the anterior horn of spinal cord, nerve muscle joints, while reducing the CGRP in injuried nerve.3.1 CGRP expression in spinal cord ventral hornlnAfter 7 days operation, CGRP in model group rats are higher than the normal group and sham-operated group (P<0.05).20 times after treatment, CGRP in Tuina group was obviously higher than that of the rest of the other four groups (P< 0.05), CGRP in model group and control group are still did high, compared with normal group and sham-operated group significantly (P< 0.05).3.2 nerve damage points of CGRP expressionAfter 7 days operation, The expression of CGRP in model group were significantly higher than the normal group and the sham-operated group (P< 0.05).20 times after treatment, The expression of CGRP in Tuina group is significantly lower than the model group and control group (P< 0.05), but still higher than normal group and sham-operated group (P< 0.05).3.3 CGRP expression in neuromuscular junctionAfter 7 days opertaion,The expression of CGRP in neuromuscular junction of model group were significantly lower than the normal group and the sham-operatedl group (P< 0.05).20 times after treatment, The expression of CGRP in neuromuscular junction of Tuina group were obviously higher than model group and control group (P< 0.05), but still lower than normal group and sham-operated group (P<0.05).[Conclusion]1 Tuina can promote the recovery of SNI rats in Plane test which stated that Tuina can improve motor function of SNI rats.2 Tuina intervention can relieve swelling degree of spinal anterior horn motor neurons, improve the function of of Nissl, regenerated the key parts of nerve-"neurons", such promoting the functional recovery.3 Tuina intervention can increase the expression of CGRP in the spinal cord ventral horn, enhance the recovery of axoplasmic transport, reduce the accumulation of CGRP in nerve damage, and regulate the neuromuscular joint function, so as to improve the function of spinal cord to peripheral motor pathways, which is the one of mechanism of Tuina promoting nerve injury repair. |
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