Feng Yong, Zhou Chonggao, Zhao Fan, Ma Tidong, Xiao Yong, Peng Kun, Xia Renpeng
Department of Fetal and Neonatal Surgery, Hunan Children's Hospital, Changsha 410007, China.
Department of Fetal and Neonatal Surgery, Hunan Children's Hospital, Changsha 410007, China.
Gene. 2024 Jun 20;912:148365. doi: 10.1016/j.gene.2024.148365. Epub 2024 Mar 12.
Hirschsprung's-associated enterocolitis (HAEC) is a prevalent complication of Hirschsprung's disease (HSCR). Zinc finger E-box binding homeobox 2 (ZEB2) and Notch-1/Jagged-2 are dysregulated in HSCR, but their role in HAEC progression remains poorly understood. We aimed to explore the role and underlying mechanism of enteric neural precursor cells (ENPCs) and the ZEB2/Notch-1/Jagged-2 pathway in HAEC development.
Colon tissues were collected from HSCR and HAEC patients. ENPCs were isolated from the HAEC group and stimulated by lipopolysaccharide (LPS). The expressions of ZEB2/Notch-1/Jagged-2 were measured using RT-qPCR and Western blot. Immunofluorescence and cell counting kit-8 assays were performed to assess the differentiation and proliferation of ENPCs. Inflammatory factors were measured by ELISA kits. Co-immunoprecipitation and bioinformatic analysis were used to explore the interaction between ZEB2 and Notch-1. Small interfering RNA and overexpression vectors were used to investigate the role and mechanism of ZEB2 and Notch-1 in regulating ENPCs' proliferation and differentiation during HAEC progression.
We observed increased LPS in the colon tissues of HAEC, with downregulated ZEB2 expression and upregulated Notch-1/Jagged-2 expression. ZEB2 interacts with Notch-1. LPS treatment downregulated ZEB2 expression, upregulated Notch-1/Jagged-2 expression, and induced proliferation and differentiation disorders in ENPCs, which were reversed by the knockdown of Notch-1. Furthermore, overexpression of ZEB2 inhibited Notch-1/Jagged-2 signaling and ameliorated inflammation and dysfunction in LPS-induced ENPCs. Notch-1 overexpression enhanced LPS-induced dysfunction, but this effect was antagonized by the overexpression of ZEB2.
Overexpression of ZEB2 ameliorates LPS-induced ENPCs' dysfunction via the Notch-1/Jagged-2 pathway, thus playing a role in HAEC.
先天性巨结肠相关小肠结肠炎（HAEC）是先天性巨结肠病（HSCR）的一种常见并发症。锌指E盒结合同源框2（ZEB2）和Notch-1/Jagged-2在HSCR中表达失调，但其在HAEC进展中的作用仍知之甚少。我们旨在探讨肠神经前体细胞（ENPCs）和ZEB2/Notch-1/Jagged-2通路在HAEC发生发展中的作用及潜在机制。
收集HSCR和HAEC患者的结肠组织。从HAEC组分离出ENPCs，并用脂多糖（LPS）刺激。采用逆转录定量聚合酶链反应（RT-qPCR）和蛋白质免疫印迹法检测ZEB2/Notch-1/Jagged-2的表达。进行免疫荧光和细胞计数试剂盒-8检测以评估ENPCs的分化和增殖。通过酶联免疫吸附测定试剂盒检测炎症因子。采用免疫共沉淀和生物信息学分析探讨ZEB2与Notch-1之间的相互作用。使用小干扰RNA和过表达载体研究ZEB2和Notch-1在HAEC进展过程中调节ENPCs增殖和分化的作用及机制。
我们观察到HAEC患者结肠组织中LPS增加，ZEB2表达下调，Notch-1/Jagged-2表达上调。ZEB2与Notch-1相互作用。LPS处理下调ZEB2表达，上调Notch-1/Jagged-2表达，并诱导ENPCs增殖和分化紊乱，而Notch-1基因敲低可逆转这种情况。此外，ZEB2过表达抑制Notch-1/Jagged-2信号传导，并改善LPS诱导的ENPCs炎症和功能障碍。Notch-1过表达增强LPS诱导的功能障碍，但ZEB2过表达可拮抗这种作用。
ZEB2过表达通过Notch-1/Jagged-2通路改善LPS诱导的ENPCs功能障碍，从而在HAEC中发挥作用。
