lncDdah1对坏死性小肠结肠炎肠上皮细胞损伤与修复的调控与机制研究

Necrotizing enterocolitis (NEC) is not a rare disease that still threatens premature infant lives. Imbalance between injury and renewal in intestinal epithelial cells is one of the key mechanisms that contribute to NEC. In previous studies, we set up a NEC neonatal murine model. We further screened a number of lncRNAs associated with NEC pathogenesis by RNA-seq, and identified that lncDdah1 was the most differentially expressed gene and it located at the sense strand of Ddah1. Ddah1 regulates nitric oxidesynthase (NOS) activity and nitric oxide（NO）level which are risk factors of NEC. Therefore, We hypothesized that lncDdah1 regulates the transcription of Ddah1 and its downstream gene, which result in intestinal epithelial cell injury and repair in NEC. In current study, characterization of lncDdah1 will be further identified. Effects and regulation mechanisms of lncDdah1 on intestinal epithelial cell injury and repair in NEC newborn murine will be also investigated by multiple biological techniques including CHIP and RIP. Our study will explore the relationship between lncDdah1 and NEC as well as its underlying mechanisms. It will be helpful to NEC effective intervention and target therapy.

坏死性小肠结肠炎(NEC)严重威胁早产儿生命健康。肠上皮细胞受损及自我修复障碍促使NEC进展恶化。近年发现长链非编码RNA（lncRNA）可调控肠上皮细胞生长与凋亡，但机制不明。申请人建立NEC新生小鼠模型，通过RNA高通量测序筛选与NEC肠上皮细胞损伤修复相关lncRNA，其中lncDdah1表达差异最明显，且位于Ddah1基因正义链上，后者参与调节一氧化氮合酶（NOS）活性及一氧化氮（NO）合成，而NOS活性及NO水平异常是诱发NEC的重要因素之一。从而推论lncDdah1可能与Ddah1相互作用参与NEC发病进展。本课题拟进一步明确lncDdah1细胞定位，时空表达；观察lncDdah1对NEC肠上皮细胞损伤及损伤后修复的影响；运用CHIP，RIP等分析lncDdah1与Ddah1相互作用及调控下游靶基因表达机制。揭示NEC转录水平发病机制，为寻找NEC分子靶向治疗的潜在靶点提供依据

肠上皮细胞损伤和自我修复障碍是坏死性小肠结肠炎（NEC）发生和进展的重要机制之一，长链非编码RNA（lncRNA）对肠上皮细胞生长和凋亡有潜在的调控作用。本研究发现，lncDdah1主要位于细胞核内，在NEC小鼠的发病过程中表达较为稳定。lncDdah1沉默后，NEC小鼠的肠道损伤减轻，炎症因子表达减少，肠道干细胞增殖能力略有增强。根据RNA pulldown结果，结合GO数据库、KEGG和PPI富集分析，初步明确了lncDdah1调控肠上皮损伤修复的分子机制，lncDdah1可能通过与Nogo-B蛋白结合，下调Nogo-B活性，从而促进NEC小鼠肠上皮细胞的炎症损伤。本研究可加深对坏死性小肠结肠炎肠上皮细胞损伤与修复的调控机制的认识，为寻找新生儿坏死性小肠结肠炎生物治疗的潜在分子靶点提供依据。