我们的前期研究证实，敲除rnf213基因2号外显子致移码突变使该基因完整敲除，成功构建了与人类烟雾病高度相似的、能稳定遗传的斑马鱼模型，目前已获得F1代。比较敲减rnf213基因的环指和ATP酶功能区的斑马鱼，其异常血管较敲除2号外显子的轻，说明完整敲除更能模拟人类MMD。而国内外单纯敲除Rnf213重要功能区构建MMD的小鼠模型均未成功。在现有工作基础上，拟用CRISPR/Cas9对单基因遗传MMS的靶基因NF1、ACTA2、BRCC3、GUCY1A3以及MMD的主要易感基因RNF213进行完整敲除，并分别构建基因修饰性的斑马鱼和小鼠模型，叠加干扰素和/或颈动脉末端银夹致慢性缺血后，对两种模型的血管表型（血管狭窄/闭塞、增生）、病理（组织病理和电镜）及其相应的遗传特征（敲除基因测序）进行鉴定和比较，通过与野生型的斑马鱼或小鼠回交后，获得能稳定遗传的F1代，高度模拟人类MMD/MMS的模型

Our previous studies have verified that frameshift mutation knockout from exon 2 of rnf213 gene resultes in loss of its function, and that such mutant zebrafish model highly mimics human moyamoya disease(MMD) and is hereditarily stable. Generation F1 is now available. The vasculature abnormality in rnf213-mutant zebrafish with knockout of ring-finger domain and AAA+ ATPase domain of rnf213 gene is less prominent than that with knockout from exon 2, indicating better suitability of rnf213-mutant MMD zebrafish model with loss of function. Nevertheless, both abroad and domestic attempts to induce mice MMD models by knocking out functional domains of rnf213 had failed. Based on current works, it is attempted to fully knockout target genes of monogenic Moyamoya Syndrome (MMS), namely gene NF1, ACTA2, BRCC3 and GUY1A3, and major susceptibility gene RNF213 of MMD by means of CRISPR/Cas9, to establish gene-modified zebrafish and mice models respectively. INF(interferon) and/or chronic ischemia by mechanic occlusion of terminal internal carotid arteries using silver clips will be plus. Then it is to be compare the vascular phenotype (stenosis/occlusion and moyamoya vessels), histopathologic characteristics under both light and electron microscopes, and respective genetic features (genotype verified by sequencing knockout genes) of both models. Finally, through hybridization with wild-type zebrafish or mice, it is aim to generate genetically stable F1 that highly simulate human MMD/MMS.