| Objective(1) To observe the expression of myelin-associated axon growth inhibitors after spinal cord been transected and in different injury degree Allen’s model in SD (Sprague-Dawley) rats. (2) To explore the optimal timepoint for preparing Spinal cord extract to simulate the microenvironment of myelin-associated axon growth inhibitors after SCI (spinal cord injury) in SD rats. Methods:(1) A total of 125 healthy adult female SD rats were selected and randomly divided into group A (as sham-operation group,25 rats), group B (hit potential 20g×5cm,25 rats), group C (hit potential 20g×10cm,25 rats), group D (hit potential 20g×15cm,25 rats), and group E (spinal cord compete transection group,25 rats).At 1d,5d,7d,14d, and 28d after successful modeling, the model and degree of spinal cord injury were identified by the BBB scores of motion function, MEP (Motor evoked potential,MEP) and pathological section, and the expression of myelin-associated axon growth inhibitors (Nogo-A, MAG, and OMgP) were test ed by Western Blot analysis and real-time fluorescent PCR (2) All the data were expressed as x± and analyzed by SPSS 19.0 software package and variance analysis. P<0.05 stands for significant difference. Results:There were totally 24 SD rats died in the experiment, among them,2 were in group B with a death rate of 8.33%,4 in group C, with a death rate of 16.67%; 8 in group D, including 2 SD rats died of urinary tract infection,1 suffered from blood urine with the death rate of 33.33%. In group E, there were totally 10 rats died,4 rats died of urinary tract infection, and 3 rats were suffered from postoperative blood urine with the death rate of 41.67%. The death rate in group E was higher than it in groupB,C,D (2) The BBB score of each group was decreased. The SD rats of the group A were only manifested by mild walking disability with poor limb concordance and recovered to normal after 5days. The SD rats of the group D and E were manifested by compelete paralysis.The BBB scores in every group were increased as time went on. At 1d,5d,7d,14d, and 28d,The BBB scores of the group A were 19.89±0.57, 20.51±0.49,21,21 and 21, It in group B were 2.94±0.57,5.88±0.84, 8.46±0.76,14.12±0.35, and 15.34±0.76, the group C were 1.72±2.12, 4.28±1.46,6.02±1.38,10.33±1.40, and 11.28±3.42, the group D were 0.00± 0.00,0.84±0.75,1.25±0.78,3.50±1.51, and 4.31±2.32, and the group E were 0.00±0.00,0.89±0.39,1.35±0.97,2.45±0.75, and 4.42±1.93 respectively. The BBB scores in group A was significant higher than it in group B, group C, group D and group E at each time-point (P<0.05), while there was no significant difference between group D and E at each time point (P> 0.05). (3) Neuronal edema ocurred in the early stage and there was no abnormal change of cell numbers at other time-point in group A. There was infiltration of inflammatory cells, obvious welling of neuron around the lesions focus in group B, C, D, E and neuron reduced at 24 hours after injury.There were formations of a large number of astrogliosis scar and syringomyelia at 28 days after injury. It was more serious in D, E compared with it in group B,C. (4) MEP latency (ms) of each group after injury were 2.24±0.15,2.28±0.14,2.35 ±0.07,2.33±0.12,2.41±0.04 in group A;3.83±0.16,3.51±0.15,2.33±0.14, 2.28±0.16 and 2.50±0.12 in group B;4.32±0.11,4.13±0.29,3.63±0.28,3.49± 0.17and 3.35±0.05 in group C; 4.74±0.22,4.44±0.06,4.48±0.25,4.61± 0.11 and 4.72±0.51 in group D;4.75±0.32,4.41±0.07,4.53±0.01,4.71±0.09 and 4.69±0.34 in group E at 1d,5d,7d,14d,28d respectively.The MEP latency in Group B, C, D,E were statistically higher than it in group A at corresponding time-point. There were significant difference within groups of the MEP latency at different time-point. There was no statistically significant difference between group D and group E at corresponding time-point. (5) The relative amount expression of MAG were 0.324±0.084,0.334±0.054,0.413 ±0.032,0.389±0.032,0.404±0.043 in group A.0.413±0.092,0.494±0.132, 0.599±0.134,0.494±0.133 and 0.396±0.098 in group B; 0.436±0.123,0.496± 0.136,0.602±0.145,0.598±0.034 and 0.401±0.143 in group C; 0.514±0.137, 0.594±0.184,0.798±0.139,0.710±0.032 and 0.405±0.039 in group D; 0.503± 0.157,0.589±0.096,0.801±0.147,0.765±0.098 and 0.423±0.119 in group E at 1d,5d,7d,14d, and 28d respectively. The relative amount expression of Nogo-A were 0.334±0.076,0.367±0.045,0.423±0.031,0.387±0.041,0.463± 0.037 in group A.0.401±0.184,0.501±0.134,0.593±0.118,0.398±0.123 and 0.384±0.124 in group B; 0.424±0.163,0.474±0.125,0.674±0.136,0.584± 0.032 and 0.414±0.025 in group C; 0.513±0.141,0.574±0.164,0.794±0.129, 0.714±0.105 and 0.494±0.098 in group D; 0.515±0.098,0.585±0.174,0.807± 0.105,0.736±0.038 and 0.485±0.032 in group E at Id,7d,14d, and 28d respectively. The relative amount expression of OMgP were 0.303±0.084, 0.312±0.105,0.323±0.132,0.337±0.021,0.298± 0.134 in group A.0.473± 0.132,0.678±0.142,0.584± 0.104,0.403±0.098 and 0.394±0.132 in group B; 0.485±0.174,0.732±0.103,0.641±0.098,0.492±0.132 and 0.401 ±v0.132 in group C;0.532±0.134,0.814±0.093,0.703±0.032,0.501±0.134 and 0.432± 0.125 in group D;0.541±0.133,0.796±0.113,0.705±0.141,0.534±0.132 and 0.395±0.023 in group E at 1d,5d,7d,14d,28d respectively. The expression of Nogo-A, MAQ OMgP could be tested 1 day after spinal cord injury in group B,C,D,E. The expression of Nogo-A began to increase at 5 days after SCI with a highest expression at 7days and gradually reduced after 14 days while they in group D,E were more remarkable. The expression of MAG peaked at 7 days and restored gradually to the level of sham-operation group after 28 days after SCI. The expression of OMgP was observed one day, peaked after 5 days, and restored to sham-operation group level gradually at 28 days after SCI. (6) The relative amount of mRNANogo-A in group A were 1.0054±0.0042,1.0049 ±0.0038,1.0019±0.0040,1.0023±0.0037,1.0040±0.0028.0.8600±0.0034,1.2557 ±0.0029,1.3097±0.0042,1.2708±0.0028and 0.8693±0.0018 in group B, 0.8958±0.0013,1.3177±0.0016,1.4817±0.0021,1.3351±0.0019 and 0.8961± 0.0011 in group C,0.9451±0.0010,1.4253±0.0012,1.5452±0.0023,1.3933± 0.0019 and 0.9394±0.0023 in group D,1.0567±0.0021,1.5925±0.0023,1.7734 ±0.0017,1.6650±0.0025 and 1.0063±0.0026 in group E at 1d,5d,7d,14d,28d respectively. The relative amount of mRNAMAG were 1.0045±0.0015, 1.0001±0.0017,1.0047±0.0021,1.0017±0.0023,1.0019±0.0024 in group A,0.9651±0.0019,1.0455±0.0023,1.0903±0.0018,1.0143±0.0026 and 0.9986 ±0.0025 in group B,1.0233±0.0017,1.1732±0.0019,1.2075±0.0018,1.1843 ±0.0023 and 1.0360±0.0027 in group C; 1.0537±0.0019,1.2395±0.0025, 1.4250±0.0028,1.243±0.0019 and 1.0744±0.0026 in group D; 1.0780 ±0.0016,1.3170±0.0024,1.5045±0.0028,1.3239±0.0029 and 1.1280±0.0034 in group E at 1,5,7,14,28d respectively. The relative amount of mRNAOmgPwere0.5001±0.0018,0.5093±0.0021,0.5016±0.0020,0.5012± 0.0018,0.5013±0.0017 in group A,0.5335±0.0021,0.5653±0.0018,0.5638± 0.0021,0.5400±0.0023 and 0.5324±0.0024 in group B; 0.5919±0.0018, 0.7059±0.0021,0.6425±0.0020,0.6104±0.0029 and 0.5882±0.0028 in group C;0.6488±0.0018,0.8545±0.0030,0.8085±0.0019,0.6705±0.0034, and 0.6340±0.0019in group D; 0.6787±0.0015,0.9030±0.0021,0.8675±0.0026, 0.7269±0.0023 and 0.6661±0.0024 in group E at 1,5,7,14,28d respectively. There was statistically significant difference (P<0.05) for relative expression amount of mRNA MAG and mRNA Nogo-A in group B.C.D and E at 5 days compared with it at other time-point within groups. There was no significantly difference between group D and group E(P>0.05)at the same time point. The relative expression amount of mRNA OmgP at 7 days was significant higher than (P<0.05) it at other time-point within groups. There was no significantly difference between group D and E (P>0.05) at the same time point. The expression of Nogo-A, MAG were mainly upregulated at lesion focus and it could be detected at the first day, peaked (Nogo-A and MAG) 7 days after SCI while the expression of OMgP peaked at 5 days,restored to the level of sham-operation group gradually after 28 days after SCI. The expression of OMGP was more remarkable in group D and group E. Conclusion:(1) The expression of myelin-associated axon growth inhibitors (Nogo-A, MAQ OMgP) was fifferent under 20g×5cm,20g×10cm,20g×15cm hit potential in Allen animal modle of SD rats after SCI.Modified Allen’s model is more fit forbasic research for SCI compared with Spinal cord transection model does. (2) At 5-7 days after SCI, the spinal cord extract may simulates axonal growth inhibition microenvironment about myelin-associated axon growth inhibitors... |
PDF Full Text Request