曲古霉素A对脊髓损伤大鼠神经行为学及组织学保护作用观察及机制探讨

Observation and mechanism on the neuroethology and histological protection of trichostatin A in rats with spinal cord injury

目的观察曲古霉素A(TSA)对脊髓损伤(SCI)大鼠神经行为学及组织学保护作用,并探讨其保护机制。方法 60只大鼠其中40只建立急性SCI模型后分为模型(M)组、低剂量(LD)组、中剂量(MD)组及高剂量(HD)组,术后腹腔注射50μg/ml、100μg/ml、200μg/ml TSA生理盐水溶液;另取10只仅做椎板切除术为假手术(S)组,剩余10只不做处理为对照(C)组,注射生理盐水溶液。术后评估神经行为学变化;处死后观察其脊髓组织病理学变化,并对比脊髓细胞凋亡情况;对比脊髓组织中增殖细胞核抗原(PCNA)、半胱氨酸天冬氨酸蛋白3(Caspase3) mRNA和蛋白表达情况。结果 BBB运动评分及斜板试验最大角度比较,C组、S组最高,MD组其次,LD组、HD组稍低,M组最低,除LD组与HD组比较、C组与S组比较差异无显著性(P> 0. 05),其他每两组间比较差异均有显著性(P <0. 05),M组、LD组、MD组、HD组随时间的延长呈显著升高趋势(P <0. 05),C组、S组随时间的延长变化不显著(P> 0. 05)。病理学观察发现,M组组织结构严重破坏、细胞病变明显; 3剂量组均不同程度改善,MD组改善最为明显。脊髓细胞凋亡率、Caspase3 mRNA和蛋白相对表达量组间比较,C组、S组最低,MD组其次,LD组、HD组稍高,M组最高,除LD组与HD组比较、C组与S组比较差异无显著性(P> 0. 05),其他每两组间比较差异均有显著性(P <0. 05)。PCNA mRNA和蛋白相对表达量比较,MD组最高,LD组、HD组其次,M组稍高,C组、S组最低,除LD组与HD组比较、C组与S组比较差异无显著性(P> 0. 05),其他每两组间比较差异均有显著性(P <0. 05)。结论 TSA对SCI大鼠神经行为学及组织学具有保护作用,其中100μg/ml保护作用最佳,可能与上调PCNA mRNA和蛋白、下调Caspase3 mRNA和蛋白表达有关。

Objective To observe the neuroethology and histological protection of trichostatin A(TSA)in rats with spinal cord injury(SCI)and to explore its protective mechanism.Methods 60 rats were selected,40 of them were made into acute SCI model by Allen method,they were randomly divided into model(M)group,low dose(LD)group,medium dose(MD)group and high dose(HD)group,intraperitoneal injection(50μg/ml,100μg/ml,200μg/ml)TSA physiological saline solution.Another 10 who underwent laminectomy were sham-operated(S)group,and the remaining 10 were not treated as control(C)group injection with physiological saline solution.The neurobehavioral changes were compared.All rats were sacrificed,and the pathological changes of spinal cord tissue were observed.The apoptosis of spinal cord cells were compared.The expressions of proliferating cell nuclear antigen(PCNA)and cysteine aspartate protein 3(caspase 3)mRNA and protein in spinal cord tissues were measured.Results The BBB motor function scores and oblique pull test maximum angles were compared between each 2 groups,and the results showed that the C and S groups were the highest.The MD group the second,the LD and HD groups slightly lower,the M group lowest,and there were significant differences between each 2 groups(P<0.05),without differences between the LD and HD groups,C and S groups(P>0.05).The above indicators of the M,LD,MD,HD groups showed a significant increase with time(P<0.05),while there were no significant differences in the C and S groups with time(P>0.05).Histopathological observation revealed that the tissue structure of group M was severely damaged,cytopathic effect was obvious.The 3 dose groups were improved to different extents,and the improvement of the MD group was the most obvious.The spinal cell apoptosis rates,relative expressions of Caspase3 mRNA and protein were compared between each 2 groups,and the results showed that the C and S groups were the lowest,the MD group the second,the LD and HD groups slightly higher,the M group highest,and there were significant differences between each 2 groups(P<0.05),without differences between the LD and HD groups,C and S groups(P>0.05).The relative expressions of PCNA mRNA and protein were compared between each 2 groups,and the results showed that the MD group were the lowest,the LD and HD groups the second,the M groups slightly higher,the C and S groups highest,and there were significant differences between each 2 groups(P<0.05),without differences between the LD and HD groups,C and S groups(P>0.05).Conclusion TSA has protective effects on neuroethology and histology of SCI rats,and 100μg/ml has the best protective effect,which may be related to up-regulation of PCNA mRNA and protein,down-regulation of Caspase3 mRNA and protein expression.