Knockdown of mitotically associated long non‐coding RNA dysregulates endothelial functions, transcriptome and proteome

Preeclampsia (PE) is a hypertensive disorder during pregnancy, which is characterized by a wide spectrum of endothelial dysfunction. However, the mechanisms underlying PE-associated endothelial dysfunction remain poorly understood. Our previous RNA-sequencing analysis showed that PE down-regulated mitotically-associated long non-coding RNA (MANCR) in human umbilical vein endothelial cells (HUVECs). Herein, we tested the hypothesis that knockdown of MANCR dysregulates endothelial functions via disrupting the transcriptome and proteome using HUVECs as a cell model. Reverse transcription quantitative PCR (RT-qPCR) was conducted to confirm the down-regulation of MANCR in PE-HUVECs. MANCR was knocked down in HUVECs from normotensive pregnancy using antisense oligonucleotides (ASOs), followed by functional assays, RNA-sequencing and proteomics. RT-qPCR revealed that PE decreased MANCR in male and female HUVECs. Compared with scrambled ASOs, MANCR ASOs reduced MANCR by 71% and 74% in male and female HUVECs, respectively. Functionally, MANCR ASOs blocked fibroblast growth factor-2 (FGF2)- and vascular endothelial growth factor-A (VEGFA)-stimulated cell proliferation, whereas they did not affect cell chemotaxis and tube formation in responses to FGF2 and VEGFA. MANCR ASOs dysregulated the expression of 37 and 39 genes as well as 97 and 15 proteins in male and female cells, respectively. Bioinformatics analysis predicted that MANCR-regulated genes and proteins in male and female HUVECs were differently associated with diseases and biological functions, many of which are relevant to cell mitosis and hypertension. Thus, down-regulation of MANCR impairs endothelial functions and causes sex-specific disruption of transcriptome and proteome, possibly partially contributing to PE-associated fetal endothelial dysfunction. KEY POINTS: Preeclampsia causes widespread endothelial dysfunction, but the roles of long non-coding RNAs in endothelial cells remain poorly understood. Mitotically associated long non-coding RNA (MANCR) is down-regulated in human umbilical vein endothelial cells (HUVECs) obtained from preeclampsia. We provide the first evidence that down-regulation of MANCR blocks endothelial cell proliferation stimulated by fibroblast growth factor 2 and vascular endothelial growth factor A. We also demonstrate that down-regulation of MANCR disrupts transcriptome and proteome as well as pathways associated with cell division, cardiovascular disease-related processes and immune responses. These findings identify MANCR as a previously unrecognized regulator of endothelial cell function.