circRNA和lncRNA在GSK-3β调控脑白质疏松症中的作用及分子机制

Aging has become a serious social problem in China, which led to leukoaraiosis (LA). However, the mechanism of pathogenesis of LA remains unclear. It have been reported that cognitive impairment, dementia, and stroke were all closely related to LA, and these diseases were all related with GSK-3β signaling pathway. Our preliminary study also found that circRNA may regulate the occurrence and development of LA by mediating the GSK-3β signaling pathway. Recent experiments suggest that lncRNA is also involved in the regulation of GSK-3 beta pathway. Therefore, on the basis of previous studies, this project will verify the regulation of non coding RNA (circRNA, lncRNA), GSK-3 beta pathway and LA in patients with LA. Therefore, this project will verify the LA patients with non-coding RNA (circRNA, lncRNA), GSK-3β pathway and LA regulation relationship. In this study, LA animal model was established and non-coding RNA interference vector was constructed. This study will investigate the regulation and molecular mechanism of ncRNA in GSK-3β promoting occurs and development of LA though circRNA interference, GSK-3 beta inhibition and functional verification. It is expected to provide a new basis for the early diagnosis and evaluation of LA, and to explore a new theoretical method for the prevention and treatment of LA.

老龄化已成为中国面临的严重社会问题，随之日趋增多的脑白质疏松症(LA)也备受关注。然而，目前关于LA的发病机制仍未定论。研究发现，LA与认知功能障碍、痴呆、卒中密切相关，而这些疾病均与GSK-3β通路有关。我们前期试验也表明，circRNA可介导GSK-3β调控LA；近期预实验提示lncRNA参与GSK-3β通路调控，故在此基础上，本项目首先将验证LA患者中非编码RNA(circRNA、lncRNA)、GSK-3β通路与LA的调控关系。其次，在筛选出差异非编码RNA前提下，建立LA动物模型，构建非编码RNA干扰载体，通过非编码RNA干涉、GSK-3β抑制及功能验证等方法，探讨非编码RNA在GSK-3β促进LA发生发展中的调控作用及分子机制，明确非编码RNA和GSK-3β的相互作用。预期为LA早期诊断、病情评估提供新的依据，为临床开展LA的预防与诊治提供新的理论方法。

（1）我们纳入110名受试者，其中脑白质疏松组50名，对照组60名。首先对来自脑白质疏松组的4个样本和来自对照组的3个样本进行了转录组测序和定量PCR。对差异表达的lncRNA和mRNA进行深入的生物信息学分析。另外，通过RT-qPCR鉴定高血压人群中脑白质疏松诊断的潜在生物标志物，并对lncRNA进行ROC分析。一种lncRNA AC020928.1被证明是高血压人群中脑白质疏松诊断的潜在生物标志物。本研究结果提示lncRNA可能在脑白质疏松发病机制中发挥重要作用，并可能成为进一步研究的新靶点。（2）为了进一步验证lncRNA在高血压脑白质病变中的作用机制，我们探索创建高血压脑白质病变动物模型。首先采用双肾两夹法建造了易卒中肾血管性高血压大鼠模型。LFB染色观察脑白质改变，电镜观察血脑屏障改变。发现该模型大鼠电镜下可以观察到血脑屏障轻度损害，但其脑白质病变并不明显。考虑小鼠与人类同源性及保守性较佳，后采用皮下植入含有血管紧张素II液体的渗透微型泵的高血压小鼠模型，LFB染色显示其脑白质病变同样不明显。提示该两种模型均不是较好的高血压脑白质病变动物模型。（3）为了丰富研究内容，我们收集和评估了华东地区脑卒中筛查人群的基本信息，并比较男性和女性短暂性脑缺血发作人群危险因素的分布差异，提示高血压、房颤、吸烟、糖尿病和超重的危险因素存在显著的性别差异。这对于面向不同性别群体制定有针对性的卒中防治策略有重要的意义。（4）我们采用自发性高血压大鼠为动物模型，予以小剂量长期喂食TMAO，通过影像学及病理染色评估，发现TMAO可以加重自发性高血压大鼠胼胝体区的白质病变。同时在体外培养的少突胶质细胞中予以TMAO及通路抑制剂干预，验证了TMAO通过ROS/NLRP3通路诱导少突胶质细胞焦亡，从而加重脑白质病变的发生。上述研究结果对于高血压所致的脑白质病变的发病机制提供了新的思路和方向。