Linalool inhibits the angiogenic activity of endothelial cells by downregulating intracellular ATP levels and activating TRPM8

Abstract

Angiogenesis crucially contributes to various diseases, such as cancer and diabetic retinopathy. Hence, anti-angiogenic therapy is considered as a powerful strategy against these diseases. Previous studies reported that the acyclic monoterpene linalool exhibits anticancer, anti-inflammatory and anti-oxidative activity. However, the effects of linalool on angiogenesis still remain elusive. Therefore, we investigated the action of (3R)-(−)-linalool, a main enantiomer of linalool, on the angiogenic activity of human dermal microvascular endothelial cells (HDMECs) by a panel of angiogenesis assays. Non-cytotoxic doses of linalool significantly inhibited HDMEC proliferation, migration, tube formation and spheroid sprouting. Linalool also suppressed the vascular sprouting from rat aortic rings. In addition, Matrigel plugs containing linalool exhibited a significantly reduced microvessel density 7 days after implantation into BALB/c mice. Mechanistic analyses revealed that linalool promotes the phosphorylation of extracellular signal-regulated kinase (ERK), downregulates the intracellular level of adenosine triphosphate (ATP) and activates the transient receptor potential cation channel subfamily M (melastatin) member (TRPM)8 in HDMECs. Inhibition of ERK signaling, supplementation of ATP and blockade of TRPM8 significantly counteracted linalool-suppressed HDMEC spheroid sprouting. Moreover, ATP supplementation completely reversed linalool-induced ERK phosphorylation. In addition, linalool-induced ERK phosphorylation inhibited the expression of bone morphogenetic protein (BMP)-2 and linalool-induced TRPM8 activation caused the inhibition of β1 integrin/focal adhesion kinase (FAK) signaling. These findings indicate an anti-angiogenic effect of linalool, which is mediated by downregulating intracellular ATP levels and activating TRPM8.

Hypoxia-induced downregulation of microRNA-186-5p in endothelial cells promotes non-small cell lung cancer angiogenesis by upregulating protein kinase C alpha

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Brassinin Promotes the Degradation of Tie2 and FGFR1 in Endothelial Cells and Inhibits Triple-Negative Breast Cancer Angiogenesis

“…In this study, the sample size chosen for each assay was based on previous publications [ 14 , 15 , 16 , 17 ]. For in-vitro assays, at least three independent experiments with at least three biological replicates (i.e., independent cell cultures) were performed.…”

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Replacement in angiogenesis research: Studying mechanisms of blood vessel development by animal-free in vitro, in vivo and in silico approaches

“…A commonly detected enzyme, which leaks from the cytoplasm of dead cells into the culture medium, is lactate dehydrogenase (LDH) ( Becker et al, 2021 ). LDH catalyzes the conversion of pyruvate to lactate, which is associated with the conversion of NAD + to NADH.…”

“…For this purpose, the radioactive compound 3H-thymidine in combination with liquid scintillation counting has been classically used ( Lou et al, 1997 ; Nishizuka et al, 2001 ). More recently, non-radioactive nucleotide analogs, such as 5-bromo-2′-deoxyuridine (BrdU) or 5-ethynyl-2′-deoxyuridine (EdU), offer more environmentally friendly alternatives, which can be detected by flow cytometry, enzyme-linked immunosorbent assay (ELISA) or immunohistochemistry ( Becker et al, 2021 ; Hillenmayer et al, 2022 ; Zhang et al, 2022 ).…”