Role of microRNAs 99b, 181a, and 181b in the differentiation of human embryonic stem cells to vascular endothelial cells.
MicroRNAs (miRNAs) are short noncoding RNAs, which post-transcriptionally regulate gene expression. miRNAs are transcribed as precursors and matured to active forms by a series of enzymes, including Dicer. miRNAs are important in governing cell differentiation, development, and disease. We have recently developed a feeder- and serum-free protocol for direct derivation of endothelial cells (ECs) from human embryonic stem cells (hESCs) and provided evidence of increases in angiogenesis-associated miRNAs (miR-126 and -210) during the process. However, the functional role of miRNAs in hESC differentiation to vascular EC remains to be fully interrogated. Here, we show that the reduction of miRNA maturation induced by Dicer knockdown suppressed hES-EC differentiation. A miRNA microarray was performed to quantify hES-EC miRNA profiles during defined stages of endothelial differentiation. miR-99b, -181a, and -181b were identified as increasing in a time- and differentiation-dependent manner to peak in mature hESC-ECs and adult ECs. Augmentation of miR-99b, -181a, and -181b levels by lentiviral-mediated transfer potentiated the mRNA and protein expression of EC-specific markers, Pecam1 and VE Cadherin, increased nitric oxide production, and improved hES-EC-induced therapeutic neovascularization in vivo. Conversely, knockdown did not impact endothelial differentiation. Our results suggest that miR-99b, -181a, and -181b comprise a component of an endothelial-miRNA signature and are capable of potentiating EC differentiation from pluripotent hESCs.
British Heart Foundation Glasgow Cardiovascular Research Centre, Institute of Cardiovascular and Medical Sciences, University of Glasgow, Glasgow, UK.
Nicole M
NM
Kane
Lynsey
L
Howard
Betty
B
Descamps
Marco
M
Meloni
John
J
McClure
Ruifang
R
Lu
Angela
A
McCahill
Christopher
C
Breen
Ruth M
RM
Mackenzie
Christian
C
Delles
Joanne C
JC
Mountford
Graeme
G
Milligan
Costanza
C
Emanueli
Andrew H
AH
Baker
D000328:Adult; D015415:Biomarkers / Q000378:metabolism; D002454:Cell Differentiation / Q000235:genetics*; D002460:Cell Line; D019070:Cell Lineage / Q000235:genetics; D053487:DEAD-box RNA Helicases / Q000378:metabolism; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism*; D042783:Endothelial Cells / Q000166:cytology* / Q000378:metabolism; D020869:Gene Expression Profiling; D055785:Gene Knockdown Techniques; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D018919:Neovascularization, Physiologic / Q000235:genetics; D009569:Nitric Oxide / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D010641:Phenotype; D015203:Reproducibility of Results; D043244:Ribonuclease III / Q000378:metabolism; D059467:Transcriptome / Q000235:genetics
D015415:Biomarkers; C561679:MIRN99 microRNA, human; C540193:MIrn181 microRNA, human; D035683:MicroRNAs; D009569:Nitric Oxide; C408087:DICER1 protein, human; D043244:Ribonuclease III; D053487:DEAD-box RNA Helicases