Dkk-1 siRNA处理对大鼠脑出血损伤的保护作用

Effects of Dkk-1 RNA interference on neuroprotection after intracranial hemorrhage in rats

目的：通过构建Dkk-1的靶向小干扰RNA（small interfering RNA,siDkk-1）,探讨其是否通过去抑制Wnt通路,进而改善血脑屏障（blood brain barrier,BBB）,对大鼠脑出血损伤起保护作用。方法：体重270～300 g成年SD雄性大鼠共144只,随机分为四组：假手术组（sham）,脑出血组,假干预组,siDkk-1干预组。分别在大鼠脑出血后24 h、72 h用real-time PCR检测Wnt-1的基因表达变化,用干湿重法检测脑含水量,伊文蓝（EB）检测血脑屏障（blood brain barrier,BBB）通透性变化。不同组别大鼠处死前均采用前肢协调实验进行行为学检测。结果：大鼠脑出血后24 h和72 h,Wnt-1基因表达水平分别较sham组显著降低（P〈0.001）,而经siDkk-1干预后24h和72 h,Wnt-1的表达水平较假干预组均显著升高（P〈0.05）。大鼠脑出血24 h和72 h术侧基底节区脑组织含水量及EB含量均显著增加（P〈0.001）,而经siDkk-1干预后较假干预组脑含水量及EB含量均显著下降（P〈0.01）,同时伴有行为学改善。结论：siDkk-1能够减轻大鼠脑出血后脑水肿,改善行为学功能,其保护机制可能与激活Wnt通路有关。

Objective：The goal of present study was to explore whether small interfering RNA targeting Dkk-1 gene（siDkk- 1） could provide neuroprotective effects after intracranial hemorrhage（ICH） lesion in rats and whether its mechanism was associated with disinhibition of Wnt pathway and improving the blood brain barrier（BBB） disruption.Methods：One hundred forty-four male Sprague-Dawley Rats（weighing 270- 300 g） were randomly divided into 4groups：sham,ICH,ICH ＋ vehicle（treatment with vehicle after ICH）,ICH ＋ siDkk-1（treatment with siDkk-1 after ICH）.All rats were sacrificed at 24 h and 72 h after ICH respectively.The gene expression of Wnt-1 was evaluated by real-time PCR.Brain water content was counted by detecting the wet weight and dry weight of brain,BBB disruption was detected by evans blue（EB） leakage content,behavioral test was examined by the forelimb use asymmetry test in different groups.Results：The expression level of Wnt-1 was decreased at 24 h and 72 h after ICH compared with the sham group（P 〈0.001）.There was up-regulated expression of Wnt-1 after treatment with siDkk-L compared with vehicletreated group on 24 h and 72 h respectively（P〈 0.05）.Brain water content and EB content in the ipsilateral basal ganglia increased significantly at 24 h and 72 h after ICH respectively（P 〈0.001）.Decreased brain water content and improved BBB disruption as well as ameliorated neurofunctional score were observed after treatment with siDkk-1 in this study（P〈 0.05）.Conclusion：siDkk-1 treatment could provide neuroprotective against brain edema following ICH and improve behavioral function,the mechanisms of neuroprotection may be associated with activation of Wnt signaling pathway.