miR‐342‐5p Is a Notch Downstream Molecule and Regulates Multiple Angiogenic Pathways Including Notch, Vascular Endothelial Growth Factor and Transforming Growth Factor β Signaling

Abstract

BackgroundEndothelial cells (ECs) form blood vessels through angiogenesis that is regulated by coordination of vascular endothelial growth factor (VEGF), Notch, transforming growth factor β, and other signals, but the detailed molecular mechanisms remain unclear.Methods and ResultsSmall RNA sequencing initially identified miR‐342‐5p as a novel downstream molecule of Notch signaling in ECs. Reporter assay, quantitative reverse transcription polymerase chain reaction and Western blot analysis indicated that miR‐342‐5p targeted endoglin and modulated transforming growth factor β signaling by repressing SMAD1/5 phosphorylation in ECs. Transfection of miR‐342‐5p inhibited EC proliferation and lumen formation and reduced angiogenesis in vitro and in vivo, as assayed by using a fibrin beads–based sprouting assay, mouse aortic ring culture, and intravitreal injection of miR‐342‐5p agomir in P3 pups. Moreover, miR‐342‐5p promoted the migration of ECs, accompanied by reduced endothelial markers and increased mesenchymal markers, indicative of increased endothelial–mesenchymal transition. Transfection of endoglin at least partially reversed endothelial–mesenchymal transition induced by miR‐342‐5p. The expression of miR‐342‐5p was upregulated by transforming growth factor β, and inhibition of miR‐342‐5p attenuated the inhibitory effects of transforming growth factor β on lumen formation and sprouting by ECs. In addition, VEGF repressed miR‐342‐5p expression, and transfection of miR‐342‐5p repressed VEGFR2 and VEGFR3 expression and VEGF‐triggered Akt phosphorylation in ECs. miR‐342‐5p repressed angiogenesis in a laser‐induced choroidal neovascularization model in mice, highlighting its clinical potential.ConclusionsmiR‐342‐5p acts as a multifunctional angiogenic repressor mediating the effects and interaction among angiogenic pathways.

Differences in proliferation rate between CADASIL and control vascular smooth muscle cells are related to increased TGFβ expression

“…EC function in blood vessels is regulated by coordination of vascular VEGF, Notch and TGFβ 47. Studies have shown that signalling of TGFβ/ALK1–Smad1/5 stimulates EC migration and proliferation 48.…”

Circulating microRNA as a marker for predicting liver disease progression in patients with chronic hepatitis B

“…For example, it has been demonstrated that mir-342 acts as a tumor suppressor gene; thus, its expression is increased during tumorigenesis (42) . It has also been identified that tumorigenesis depends on angiogenesis, the process of new blood vessel formation from an existing vasculature (43) .…”

“…The eventual result is an alteration in the net balance between negative and positive regulators. Accordingly, it has been reported that miRNA-342 regulates angiogenesis likely through the modulation of TGF-β signaling as mediated by VEGFR and endoglin, a co-receptor of the TGF-β receptor signaling pathway (42) . Together, these finding suggest that miRNA-342 suppresses angiogenesis.…”

SNAI1, an endothelial–mesenchymal transition transcription factor, promotes the early phase of ocular neovascularization

“…Assessment of tube formation ability of HUVECs was performed as described previously [31]. HUVECs (1 × 10 5 cells per well) were loaded onto 200 μL matrigel (1:1, BD Biosciences) which was pre-coated in 48-well plates and incubated at 37 °C for 4 h. The number of loops and total length of cell cords in the enclosed lumen structures were determined.…”