Dickkopf-1 Promotes Angiogenesis by Upregulating VEGF Receptor 2-mediated mTOR/p70S6K Signaling in Hepatocellular Carcinoma

Abstract

Background: The expression of Dickkopf-1 (DKK1), a negative regulator of the Wnt/β-catenin signaling pathway, is upregulated in hepatocellular carcinoma (HCC). Here, we investigated the tumorigenic and angiogenic potential of DKK1 in HCC.Methods: Stable cell lines were established using the clustered regularly interspaced short palindromic repeats (CRISPR)-associated nuclease 9 (CRISPR/Cas9)-based DKK1 knock-out system in Hep3B cells and the tetracycline-based DKK1 inducible system in Huh7 cells. Multicellular tumor spheroids (MCTSs) were cultured using Hep3B stable cells. We also employed xenografts generated using Hep3B stable cells and transgenic mouse models established using hydrodynamic tail vein injection.Results: The angiogenic potential increased in HUVECs treated with CM from Huh7 stable cells with high DKK1 expression and Hep3B wild-type cells. There was an increase in the phosphorylation of vascular endothelial growth factor receptor 2 (VEGFR2) and p70 S6 kinase (p70S6K), which are downstream proteins of VEGFR2-mediated mTOR/p70S6K signaling. MCTSs generated using Hep3B wild-type cells promoted compact spheroid formation and raised expression of CD31 and epithelial-mesenchymal transition (EMT) markers, compared to the controls (all P<0.01). Xenograft tumors generated using Hep3B cells with DKK1 knock-out (n=10) exhibited slower growth than, the controls (n=10) and the expression of Ki-67, VEGFR2, CD31 and EMT markers decreased (all P<0.05). In addition, forced DKK1 expression with HRAS in transgenic mouse livers (n=5) resulted in the formation of more tumors and increased expression of Ki-67, VEGFR2, CD31, and fibronectin, compared to the controls (n=5) (all P<0.05).Conclusions: Our findings indicate that DKK1 facilitates angiogenesis and tumorigenesis by upregulating VEGFR2-mediated mTOR/p70S6K signaling in HCC.

Immunohistochemical and Morphometric Assessment on the Biological Function and Vascular Endothelial Cells in the Initial Process of Cortical Porosity in Mice With PTH Administration

“…Among these genes, we noted that Dkk1 is an osteocyte-derived molecule, which was reportedly an inhibitor of Wnt signaling and an angiogenesis activator. [31][32][33] Because Ckap4 (a Dkk1 receptor) 34,35 and Plvap (plasmalemma vesicle-associated protein, an angiogenetic factor) 36 expressions were also elevated, we postulated that osteocyte-derived molecules, such as Dkk1, affected vascular endothelial cells to induce angiogenesis. This reaction potentially enables invasion into the cortical bone with osteoclasts in the PTH group.…”

Updating the Clinical Application of Blood Biomarkers and Their Algorithms in the Diagnosis and Surveillance of Hepatocellular Carcinoma: A Critical Review

“…However, by altering the tumour microenvironment and causing inflammation, DKK1 seems to promote tumour invasion and migration via TGF-β1 [87]. Additionally, as recently shown, DKK1 stimulates HCC angiogenesis and tumorigenesis via VEGFR2mediated mTOR/p70S6K signalling [88].…”

mTOR (Mammalian Target of Rapamycin): Hitting the Bull’s Eye for Enhancing Neurogenesis After Cerebral Ischemia?

“…[76][77][78] Numerous studies have demonstrated a positive correlation between mTOR signaling and angiogenesis in ischemic brain injury. 79,80 It has been reported that COMP-Ang1 induced prominent angiogenesis through an AKT/mTOR interaction, leading to enhanced migration of human cord blood-derived endothelial progenitor cells, tube formation, and vascular morphogenesis. Besides, the combined treatments of endothelial progenitor cells and COMP-Ang1 effectively mitigated ischemic brain injury by increasing angiogenesis.…”