大鼠脊髓横断性损伤后细胞周期蛋白D3的表达变化

Expression of Cyclin D3 following transected spinal cord injury in rats

目的探讨细胞周期蛋白 D3(Cyclin D3)在横断性脊髓损伤(tSCI)后的表达变化以及定位情况。方法将48只成年 SD 大鼠随机分为8组:正常对照组,T9横断伤2、8 h、1、3、5、7和14 d 组,每组6只。采用 Western blot 测定损伤后各时间段 Cyclin D3蛋白水平在脊髓中的表达变化。采用免疫组织化学方法检测 Cyclin D3在正常以及损伤后脊髓中的分布和定位。结果 West-ern blot 显示,Cyclin D3蛋白水平在 tSCI 后头、尾段均呈现先升高后下降的趋势,尾段明显:CyclinD3的表达于损伤后8 h 开始逐渐升高,3d 达到高峰,一直持续到第5天,之后逐渐下降。免疫组织化学表明 Cyclin D3在正常脊髓中均匀分布,损伤后3 d,Cyclin D3在脊髓白质和灰质中表达明显增强;免疫荧光双标记表明 Cyclin D3与神经元的标记物 neuronal nucleus(NeuN)、少突胶质细胞标记物 cyclic nucleotide 3′phosphohydrolase(CNPase)有明显共定位,与星形胶质细胞标记物 glial fibril-lary acidic protein(GFAP)和小胶质细胞标记物 OX-42也存在部分共定位。结论脊髓损伤后 Cyc-lin D3蛋白水平呈现明显的时相变化,并且与神经元、少突胶质细胞、星形胶质细胞和小胶质细胞存在共定位,提示 Cyclin D3参与了脊髓损伤后的病理生理过程。

Objective To explore the expression and distribution of Cychn D3 after transected spinal cord injury （tSCI）. Methods tSCI was performed at T9 in adult Sprague-Dawley （SD） rats. The rats were sacrificed on the postoperative h 2 and 8 h, and day 1,3,5,7,14 respectively. Western blotting was used to detect the changes of Cychn D3 protein expression after tSCI. The distribution and localization of Cychn D3 in spinal cord was investigated by immunohistochemical staining and immunofluorescence double staining. Results Western blotting analysis showed the expression of Cyclin D3 was increased firstly and then had a decrease trend in both broken ends of transected spinal cord, especially the caudal parts. Cychn D3 was enhanced gradually at 8th h and reached its peak at 3rd day. This high level was kept until 5th day, and then decreased. Immunohistochemistry staining indicated that Cyclin D3 was distributed uniformly in normal spinal cord. At 3rd day, the staining was elevated obviously in both wltite and grey matter. Immunofluorescence double staining suggested that Cyclin D3 was co-localized with NeuN, CNPase, GFAP and OX-42. Conclusion The expression of Cyclin D3 undergoes spafiotemporal changes after tSCI. These alterations may be involved in secondary spinal cord lesion such as neuronal apoptosis and proliferation of glial ceils.