METTL14 aggravates endothelial inflammation and atherosclerosis by increasing FOXO1 N6-methyladeosine modifications.

Aims: The N6-methyladenosine (m6A) modification plays an important role in various biological processes, but its role in atherosclerosis remains unknown. The aim of this study was to investigate the role and mechanism of m6A modification in endothelial cell inflammation and its influence on atherosclerosis development. Methods: We constructed a stable TNF-α-induced endothelial cell inflammation model and assessed the changes in the expression of m6A modification-related proteins to identify the major factors involved in this process. The m6A-modified mRNAs were identified by methylated RNA immunoprecipitation (RIP) sequencing and forkhead box O1 (FOXO1) was selected as a potential target. Through cytological experiments, we verified whether methyltransferase-like 14 (METTL14) regulates FOXO1 expression by regulating m6A-dependent mRNA and protein interaction. The effect of METTL14 on atherosclerosis development in vivo was verified using METTL14 knockout mice. Results: These findings confirmed that METTL14 plays major roles in TNF-α-induced endothelial cell inflammation. During endothelial inflammation, m6A modification of FOXO1, an important transcription factor, was remarkably increased. Moreover, METTL14 knockdown significantly decreased TNF-α-induced FOXO1 expression. RIP assay confirmed that METTL14 directly binds to FOXO1 mRNA, increases its m6A modification, and enhances its translation through subsequent YTH N6-methyladenosine RNA binding protein 1 recognition. Furthermore, METTL14 was shown to interact with FOXO1 and act directly on the promoter regions of VCAM-1 and ICAM-1 to promote their transcription, thus mediating endothelial cell inflammatory response. In vivo experiments showed that METTL14 gene knockout significantly reduced the development of atherosclerotic plaques. Conclusion: METTL14 promotes FOXO1 expression by enhancing its m6A modification and inducing endothelial cell inflammatory response as well as atherosclerotic plaque formation. Decreased expression of METTL14 can inhibit endothelial inflammation and atherosclerosis development. Therefore, METTL14 may serve as a potential target for the clinical treatment of atherosclerosis.

Down-regulation of lncRNA-NEF indicates poor prognosis in intrahepatic cholangiocarcinoma.

LncRNA-NEF is a tumor suppressor lncRNA in liver cancer. The present study aimed to investigate the role of lncRNA-NEF in intrahepatic cholangiocarcinoma (IHCC), which is second most common type of primary cancer of the hepatobiliary system that causes high mortality rate. In the present study we found that lncRNA-NEF was down-regulated, while Runt-related transcription factor 1 (RUNX1) was up-regulated in tumor tissues than in adjacent healthy tissues of IHCC patients. Expression levels of lncRNA-NEF and RUNX1 were significantly and reversely correlated in tumor tissues but not in adjacent healthy tissues. Plasma levels of lncRNA-NEF were significantly lower in IHCC patients than in healthy controls. Down-regulation of lncRNA-NEF effectively distinguished stage I and II IHCC patients from healthy controls. Patients were followed up for 5 years, patients with high plasma levels of lncRNA-NEF showed significantly better survival conditions compared with patients with low expression levels of lncRNA-NEF. LncRNA-NEF overexpression led to inhibited expression of RUNX1 in cells of IHCC cell lines and inhibited cancer cell migration and invasion. In contrast, RUNX1 overexpression showed no significant effects on lncRNA-NEF expression, but attenuated the effects of lncRNA-NEF overexpression on cancer cell migration and invasion. We therefore concluded that lncRNA-NEF participated in IHCC possibly by interacting with RUNX1.