| Abstract:Objective:To explore the effect of promoted axonal regeneration and functional recovery with GSK-3βinhibitors in vivo after spinal cord injury in rats.Methods:①A total of 115 healthly female Sprague-Dawley (SD) rats (weight 250±30)were randomly allocated into TDZD-8 group,PBS group,SCI group and Sham operated group with 10 rats in Sham operated group and 35 in other groups。The laminectomy was performed and a weight-dropoing impact at spinal levels of T9 to cause complete paraplegia。TDZD-8 or PBS were injected at the dose of (1mg/kg/Qd) or (60μl/Qd) through subarachnoid one hour after SCI and continued for three weeks。②The rats were killed by perfusion with 4%paraformaldehyde at 8h,24h,7d after SCI(2 rats at each time phase),then the frozen sections were stained with TUNEL method to measure the degree of apopotic。③The rats were killed by perfusion with 4%paraformaldehyde at 24h,7d,14d after SCI(2 rats at each time phase),then the frozen sections were stained with SP immunohistochemical method to measuring the expression of GAP-43。④Rats were used to make BBB scale in order to evaluate lower limb function recovery at 3w,6w,8w after SCI(5 rats at each time phase)。⑤Six weeks after SCI, three rats were injected FITC fluorescent tracer from sensorimotor cortex in each group。Two weeks after FITC inject, the animals were killed by perfusion with 4% paraformaldehyde, then the frozen sections were observed by Confocal microscopy in order to know the FITC fluorescent tracer passing condition。⑥Three weeks and Eight weeks after SCI, five rats were choosen to detect amplitude and latency changes of somatosensory evoked potential in each group。Results:①At the eight hours after SCI,the apoptosis positive cells were observed。The positive cells of TDZD-8 group were significantly lower than PBS group and SCI group, but higher than the Sham operated group.Eight hours after SCI,the positive cells of each group are 50.27±4.74,58.93±2.46,58.47±2.09,8.27±1.44 in turns。Twenty-four hours after SCI,the positive cells of each group are 45.73±4.95,55.40±3.94,54.53±3.11,8.27±1.44 in turns。Seven days after SCI,the positive cells of each group are 10.67±1.50,15.67±1.49,15.60±1.68,8.27±1.44 in turns。②After SCI, the GAP-43 positive reaction was increased gradually,and rose to its peak on the 14th day, which in TDZD-8 group was remarkably superior to that in other groups in different time point (P<0.05)。One day after SCI,the positive area of each group are 79.2±5.31,43.3±3.99,42.3±4.77,6.3±2.31 in turns,Seven day after SCI,the positive area of each group are:107.3±4.43,70.1±3.57,68.2±3.34,6.3±2.31 in turns,Two weeks after SCI,the positive area of each group are:156.7±4.35,94.5±3.91,96.1±3.56,6.3±2.31 in turns。③The structure of regenetation axonals are significantly disordered in lesion areas,TDZD-8 group have more regenetation axonals extend into caudal while the PBS group and SCI group have little regenetation axonal extend into caudal。The areas of regenetation axonals was followed:82.0±1.7,35.2±1.7,33.74±1.3,138±2.2。④Somatosensory evoked potentials examination postopertative 3,8 week:Compared TDZD-8 group with PBS group and SCI group,The waveform of somatosensory evoked potentials was significantly short, and the amplitude was significantly high。⑤BBB locomotor scale:All the BBB scores have arised, The mean of BBB scores of four group at 8 weeks after SCI were 9.4±1.14,8.2±0.45,8.0±0.71,21±0.00。Among the groups,there was significant difference between TDZD-8 group with PBS group and SCI group(p<0.05)。Conclusion:①TDZD-8 can inhibit neuronal apoptosis after spinal cord injury and reduce secondary spinal cord injury。②TDZD-8 can increase the growth associated protein 43 expression, reduce latency of somatosensory evoked potentials and improve the amplitude,promote the survival neuron sprouting or promote axonal regenaration,increase the plasticity of neurons,promote functional recovery。...
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