小鼠局灶性脑缺血后原蛋白转化酶1及神经肽Y的表达

摘要

目的：观察小鼠局灶性脑缺血后脑皮质神经细胞内原蛋白转化酶1（PC1）及其作用底物神经肽Y（NPY）的表达变化，探讨PC1在神经细胞缺血损伤中的作用。方法将24只雄性C57小鼠用计算机随机法分为假手术组和缺血－再灌注4、24 h组，每组各8只。采用线栓法制备小鼠大脑中动脉阻塞模型，阻塞1 h再灌注。采用Western Blot法、实时荧光定量核酸扩增检测小鼠脑皮质神经细胞中PC1及NPY蛋白、mRNA的表达变化。结果（1）与假手术组比较，缺血侧皮质脑组织PC1 mRNA缺血－再灌注4 h组表达增加（2.66±0.24），缺血－再灌注24 h组增加（2.07±0.23），差异均有统计学意义（均P＜0.05）。与假手术组比较，缺血－再灌注4 h组PC1前体蛋白水平明显增加（P＜0.05），24 h组差异无统计学意义（P＞0.05）。（2）与假手术组比较，缺血－再灌注4 h组前体NPY前体蛋白水平及mRNA增加（P＜0.05），mRNA表达为2.31±0.27；24 h组蛋白前体水平增加（P＜0.05）， NPY mRNA表达差异无统计学意义（P＞0.05）。结论小鼠脑缺血－再灌注后PC1前体表达增多，从而影响PC1的加工活性，导致PC1的作用底物NPY蛋白以前体形式堆积，可能为神经细胞缺血损伤的内在机制之一。%Objectives Tostudytheexpressionchangesofproproteinconvertase1(PC1)incerebral cortex nerve cells and its substrate neuropeptide Y (NPY)after focal cerebral ischemia in mice and to investigatetheeffectofPC1inneuronalischemicinjury.Methods Twenty-fourmaleC57micewere randomly allocated into a sham-operation group,an ischemia-reperfusion 4-or 24-hour group with computer (n=8 in each group). A rat model of middle cerebral artery occlusion was induced by the intraluminal suture method. Western blot and real-time quantitative nucleic acid amplification were used to detect the expression changes of PC1,NPY,and mRNA in mouse cortical neurons. Results (1)Compared with the sham operation group,the expression of PC1 mRNA of ischemic cortex brain tissue at ischemic side in the ischemia-reperfusion 4-hour group increased 2. 66 ± 0. 24 and in the ischemia-reperfusion 24-hour group expressed 2. 07 ± 0. 23 (all P<0. 05). Compared with the sham operation group,the PC1 precursor protein level increased significantly at 4 hours (P<0. 05). There was no significant difference in the 24-hour group (P >0. 05 ). (2 )Compared with the sham operation group,the preproNPY mRNA and protein level increased significantly after reperfusion in the ischemia-reperfusion 4-hour group (P < 0. 05 ),the mRNA expressed 2. 31 ± 0. 27,and the increase of precursor protein level continued until 24 hours. Conclusion TheexpressionofprecursorPC1increasedaftercerebralischemia-reperfusioninmice, thus affecting the processing activity of PC1 ,and resulting in NPY protein,an active substrate of PC1 accumulated with the form of precursors,which may be one of the underlying mechanisms of neuronal ischemic injury.