短暂脑缺血后蛋白伴侣hsp40变化对延迟性神经元死亡的影响

Alterations of chaperone hsp40 and its influence on the CAl neurons death after transient cerebral ischemia

目的探讨短暂脑缺血后海马CAl区神经元内蛋白伴侣hsp40的变化及对延迟性神经元死亡的影响。方法采用20min全脑缺血大鼠模型。28只Wistar大鼠按照再灌注时间分为假手术组，4h恢复组，24h恢复组，72h恢复组，每组7只。采用免疫组织化学法结合激光扫描共聚焦显微镜观察短暂脑缺血后蛋白伴侣hsp40在神经元内分布的变化，差速离心结合蛋白印迹分析短暂脑缺血后蛋白伴侣hsp40在细胞内数量的变化。结果免疫组化激光扫描共聚焦显微镜研究显示，短暂脑缺血后胞浆内的hsp40先开始减少，然后胞核内的hsp40再减少，直至神经元全部死亡。差速离心和蛋白印迹显示短暂脑缺血后胞浆内的hsp40从1．00±0．21逐渐减少到再灌注24h后的0．23±0．13，P〈0．01；蛋白聚集物中hsp40的含量则从1．00±0．18升高到再灌注24h后的8．61±1．89，P〈0．01。结论短暂脑缺血-再灌注后海马CAl区神经元内蛋白伴侣hsp40的显著减少是导致蛋白聚集物形成的一个重要因素，进而导致延迟性神经元死亡。

Objective To investigate the alteration of chaperone hsp40 and its effects on the dealyed neuron death in the CAl neurons after transient cerebral ischemia. Method Twenty-minute transient global ischemia rat model was used. Following different reperfusion period, all the 28 wistar rats were divided into sham-operation group,4-hour recovery group , 24- hour recovery group and 72-hour recovery group,7 ratsin in each group. Immunochemistry and laser scanning confocal microscopy were used to observe the distributional alteration of hsp40 in the neurons. Differential centrifuge and westemblot analysis were used to analyze the quantitative alteration of hsp40 and its redistribution in the neurons. Results Immunochemistry and laser scanning confocal microscopy showed the reduction of hsp40 first in cytosol, then in the nucleus until all the neurons in the CAl region died. Differential centrifuge and westemblot analysis showed the quantity of hsp40 decreased from （ 1.00 ± 0.21 ） to （0.23 ± 0.13） （P 〈 0.01 ） after 24-hour reperfusion; the quantity of hsp40 in the protein aggregates increased from （1.00 ± 0.18） to （8.61 ± 1.89） （P 〈 0.01） after 24-hour reperfnsion. Conclusions The reduction of hsp40 in the neurons of hippocampns CAl region is an important factor resulting in protein aggregates formation.