小肠血管畸形治疗新靶点——SNORA31抑制核仁素细胞膜表达的机制研究

Small bowel vascular malformation (SBVM) is the main cause of intestinal bleeding. The pathogenesis of SBVM was not clear yet, and target therapy was not available. Our previous study on HIF-2α/VEGF pathway accidentally found expression of Nucleolin (NCL) was up-regulated with increased angiogenesis process. And NCL expressing on surface of endothelial cell mediated by VEGF was a key process during angiogenesis. Further studies suggested SNORA31, one of non-coding RNAs, showed overexpression under thalidomide interference, and could bind NCL in high-intensity by bioinformatics analysis. Higher cell-surface NCL expression in abnormal intestinal vascular malformation lesions of SBVM patients was also found. Another preliminary experiments showed functional fragments of SNORA31 could inhibit the expression of cell-surface NCL. Therefore, the present project will explore the binding site and intensity between SNORA31 and NCL, and how SNORA31 regulates cell-surface NCL by protein microarray, deletion mutant and RNA-CHIP techniques. Cell transfection, CRISPR/Cas9 and mouse Sponge-Matrigel model will be used to investigate how SNORA31/NCL interaction regulates angiogenesis process. It is of great significance to clarify the mechanism of SNORA31/NCL in the occurrence and development of SBVM, and lay a theoretical foundation for the treatment of SBVM as a new target.

小肠血管畸形（SBVM）是小肠出血的主要原因，发病机制尚不明晰、缺乏针对性强的治疗手段。我们前期研究发现血管生成增加时核仁素（NCL）表达上调，而NCL在细胞膜表达是促进血管生成的重要过程；进一步研究发现沙利度胺干预下SNORA31表达显著升高；CatRapid分析提示SNORA31能够和NCL高强度结合；SBVM患者畸形肠道血管内皮细胞的细胞膜NCL表达增高；且SNORA31片段能够抑制NCL在HUVEC细胞膜的表达。本项目拟借助蛋白质微阵列芯片，缺失突变体，RNA免疫共沉淀等技术深入研究SNORA31和NCL的结合强度和位点，并通过小鼠海绵-基质模型探究上述分子对血管生成的调控作用，证明SNORA31对NCL细胞膜表达的抑制作用，揭示SNORA31/NCL参与血管生成过程的作用机制，对阐明SBVM的发生发展机制有重要意义，并为治疗SBVM探索新靶点奠定理论基础。

胃肠道血管畸形（GIVM）是小肠出血的最主要原因，发病机制尚不明晰、缺乏针对性强的治疗手段。我们在国家自然科学基金（81670505）的资助下研究了EGFL6及其下游信号通路和GIVM发生发展之间关系。实验结果证明：1）EGFL6在SBVM患者血清及手术切除标本中高表达；2) 细胞和斑马鱼实验证明EGFL6促进血管形成；3）EGFL6主要通过其下游分子PAX6的调控从而达到促进血管形成的作用；4）沙利度胺的作用机制之一是通过抑制EGFL6水平从而达到抑制血管生成的作用；5）沙利度胺对EGFL6抑制作用表现的具体机制是促进EGFL6降解；6）沙利度胺主要通过E3泛素连接酶CRBN促进EGFL6降解。以上结果使我们对GIVM的发病机制以及沙利度胺的治疗效果有了更深一步的认识，对进一步寻找治疗GIVM的靶点提供了理论基础。