Department of Pediatric Surgery, the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, PR China.
Department of Endoscopy, Shaanxi Nuclear Industry 215 Hospital, Xianyang, Shaanxi, PR China.
Exp Physiol. 2020 Nov;105(11):1939-1949. doi: 10.1113/EP088780. Epub 2020 Oct 7.
What is the central question of this study? Long non-coding RNAs (lncRNAs) are widely involved in the progression of Hirschsprung's disease (HSCR), but the role of actin filament associated protein 1 antisense RNA1 (AFAP1-AS1), an lncRNA, in HSCR has not been explored before. What is the main finding and its importance? Downregulation of AFAP1-AS1 blocks enteric neural crest stem cell proliferation, differentiation, migration and invasion and promotes the occurrence of HSCR via the miR-195/E2F3 axis, indicating thatAFAP1-AS might be a potential biomarker for HSCR patients.
Long non-coding RNAs (lncRNAs) are involved in several human disorders. Nevertheless, it remains unclear whether they are implicated in the phenotypes of enteric neural crest stem cells (ENCSCs) in Hirschsprung's disease (HSCR). Therefore, we designed this study to explore the pathogenicity of AFAP1-AS1 for HSCR. Microarray analysis and bioinformatic tools were used to screen out the differentially lncRNAs and microRNAs (miRNAs) in patients with HSCR. Small interference RNA transfection was applied to carry out functional experiments in ENCSCs. Cellular activities were detected by cell counting kit-8, 5-ethynyl-2'-deoxyuridine, Transwell assays and flow cytometry. Finally, rescue experiments were performed to examine the cofunction of AFAP1-AS1 and miR-195 and of miR-195 and E2F transcription factor 3 (E2F3). AFAP1-AS1 was reduced in HSCR patients. Meanwhile, knockdown of AFAP1-AS1 reduced the cell migratory and proliferative capacities and facilitated cell apoptosis along with G0/G1 phase arrest. E2F3 was diminished when miR-195 was upregulated, and AFAP1-AS1 inhibition reduced its ability to bind to miR-195. Altogether, AFAP1-AS1 silencing acts as an endogenous RNA by interacting with miR-195 to alter E2F3 expression, thus conferring repressive effects on ENCSC activity and promoting HSCR progression.
本研究的核心问题是什么？长链非编码 RNA（lncRNA）广泛参与先天性巨结肠（HSCR）的进展，但 actin filament associated protein 1 antisense RNA1（AFAP1-AS1）这种 lncRNA 在 HSCR 中的作用尚未被探索。主要发现及其重要性是什么？下调 AFAP1-AS1 可通过 miR-195/E2F3 轴阻断肠神经嵴干细胞增殖、分化、迁移和侵袭，促进 HSCR 的发生，表明 AFAP1-AS 可能是 HSCR 患者的潜在生物标志物。
长链非编码 RNA（lncRNA）参与多种人类疾病。然而，它们是否参与先天性巨结肠（HSCR）肠神经嵴干细胞（ENCSC）的表型尚不清楚。因此，我们设计了这项研究以探讨 AFAP1-AS1 对 HSCR 的致病作用。通过微阵列分析和生物信息学工具筛选出 HSCR 患者中差异表达的 lncRNA 和 microRNAs（miRNA）。用小干扰 RNA 转染在 ENCSC 中进行功能实验。通过细胞计数试剂盒-8、5-乙炔基-2'-脱氧尿苷、Transwell 测定和流式细胞术检测细胞活性。最后，进行了挽救实验以检查 AFAP1-AS1 和 miR-195 以及 miR-195 和 E2F 转录因子 3（E2F3）的共同作用。在 HSCR 患者中发现 AFAP1-AS1 减少。同时，下调 AFAP1-AS1 降低了细胞迁移和增殖能力，并促进细胞凋亡以及 G0/G1 期阻滞。当 miR-195 上调时，E2F3 减少，AFAP1-AS1 抑制降低了其与 miR-195 结合的能力。总之，AFAP1-AS1 沉默通过与 miR-195 相互作用作为内源性 RNA 发挥作用，改变 E2F3 的表达，从而对 ENCSC 活性产生抑制作用，并促进 HSCR 的进展。
