Vascular endothelial-derived Von Willebrand factor inhibits lung cancer progression through the αvß3/ERK1/2 axis.

Lung cancer remains a common malignant tumor causing death due to the rapid industrialization and serious pollution of the environment. The Von Willebrand Factor (vWF) protein is an endothelial marker and is widely used to diagnose cancer and other inflammations, however its exact mechanism of action remains largely unexplored. In particular, how it plays two opposing roles in tumor development is not clear. Our study aimed to the impact of endothelial-derived vWF on tumor development by co-culturing human umbilical vein endothelial cells (HUVECs) with lung cancer cells (95D and A549). A knockdown of endothelial-derived vWF assisted lung cancer cell in proliferation, migration and inhibited apoptosis in vitro, while overexpression of endothelial-derived vWF inhibited the proliferation, migration and induced apoptosis of lung cancer cells. The results of further experiments indicated that the vWF secreted by endothelial cells could affect lung cancer cell migration and apoptosis via its binding to integrin αvß3 on the surface of lung cancer cells. Furthermore, a novel finding was the fact that endothelial-derived vWF inhibited lung cancer cell apoptosis by phosphorylating ERK1/2. At the same time, we established experimental lung metastasis model and xenograft model in normal mice and vWF-/- mice, and found that knockout of vWF in mice significantly promoted lung cancer growth and metastasis. In conclusion, our research found that endothelial-derived vWF could directly combine to αvß3 on the exterior of A549 and 95D, thereby mediating lung cancer proliferation, migration and apoptosis and inhibiting the development of lung cancer.

Hypoxia promotes the expression of Von Willebrand factor in breast cancer cells by up-regulating the transcription factor YY1 and down-regulating the hsa-miR-424.

Von Willebrand factor (VWF), a large glycoprotein with hemostatic properties, is mainly synthesized by megakaryocytes and endothelial cells (ECs). In recent years, studies have found that tumor cells also can produce VWF de novo. Tumor growth is usually accompanied by hypoxic environment, and whether hypoxia will influence von Willebrand factor production in tumor cells is still unknown. In this research, we demonstrated that hypoxia could induce the production of VWF in breast cancer cells (MCF-7 and MDA-MB-231 cell lines), and promoted cell migration as well as angiogenesis. Notably, VWF is a key factor for hypoxia to promote breast cancer cell migration and angiogenesis, and knocking down VWF can attenuate the effects of hypoxia. Further study was conducted on the molecular mechanism to clarify why hypoxia can promote VWF synthesis in breast cancer cells. We found that Yin-Yang 1 (YY1, a transcription factor) had a binding site to the promoter region of VWF, and acted as a transcriptional activator of VWF. Meanwhile, hsa-miR-424 inhibited VWF production by associating with the 3'-UTR of VWF mRNA. Here, we proved that hypoxia up-regulated the transcription factor YY1 and down-regulated hsa-miR-424 to increase the expression level of VWF. Additionally, knockdown of transcription factor YY1 and transfection of hsa-miR-424 mimics had a synergistic effect in reducing hypoxia-induced VWF production of breast cancer cells, cell migration and angiogenesis in vitro.

Lung adenocarcinoma-derived vWF promotes tumor metastasis by regulating PHKG1-mediated glycogen metabolism.

Tumor metastasis is a series of complicated biological events. Hematogenous metastasis mediated by von Willebrand factor (vWF) is critical in tumor metastasis. However, the source of vWF and its role in tumor metastasis are controversial, and the further mechanism involved in mediating tumor metastasis is still unclear. In this study, we first demonstrated that lung adenocarcinoma cells could express vWF de novo and promotes tumor metastasis. Through the analysis of transcriptome sequencing, the metastasis promotion effect of vWF may be related to phosphorylase kinase subunit G1 (PHKG1), a catalytic subtype of phosphorylase kinase (PhK). PHKG1 was highly expressed in lung adenocarcinoma patients and led to poor prognosis. Further experiments found that lung adenocarcinoma-derived vWF induced the upregulation of PHKG1 through the PI3K/AKT pathway to promote glycogenolysis. Glycogen was funneled into glycolysis, leading to increased metastasis. Tumor metastasis assayed in vitro and in vivo showed that knockdown of PHKG1 or synergistic injection of phosphorylase inhibition based on the overexpression of vWF could inhibit metastasis. In summary, our research proved that lung adenocarcinoma-derived vWF may mediate tumor metastasis by regulating PHKG1 to promote glycogen metabolism and suggested potential targets for inhibition of lung adenocarcinoma metastasis.

Breast cancer cells-derived Von Willebrand Factor promotes VEGF-A-related angiogenesis through PI3K/Akt-miR-205-5p signaling pathway.

The metastasis and angiogenesis of breast cancer has always been a difficult problem for treatment. It has recently been discovered that Von Willebrand Factor (vWF), in addition to hemostasis, also plays a role in tumor metastasis and angiogenesis. We noticed that besides endothelial cells, breast cancer cells (MDA-MB-231 and MCF-7) could also express vWF. In vitro experiments showed that knocking down vWF inhibited breast cancer cell metastasis. And we found that overexpression of vWF significantly promoted VEGF-A-dependent vascular proliferation in vitro by activating the PI3K/Akt signaling pathway. Further studies indicated that inhibition of PI3K/Akt signaling pathway up-regulated the expression of miR-205-5p, and miR-205-5p could bind to the 3'UTR region of VEGF-A to hinder the production of VEGF-A. Furthermore, when a spontaneous lung metastasis model was established in Balb/c female mice, knockdown of vWF in 4 T1 cells resulted in a decrease in tumor blood vessel density and effectively inhibited lung metastasis, accompanied by a decrease in the expression level of VEGF-A and an increase in the expression level of miR-205-5p. In summary, our findings provide experimental evidence that overexpression of vWF in breast cancer cells down-regulates the expression of miR-205-5p and up-regulates the expression of VEGF-A through the PI3K/Akt signaling pathway, thereby promoting tumor angiogenesis and metastasis.

Both endogenous and exogenous miR-139-5p inhibit Fusobacterium nucleatum-related colorectal cancer development.

Colorectal cancer (CRC) is one of the most common cancers worldwide. Colorectal carcinogenesis represents a heterogeneous process which influenced by diet, environmental and microbial exposures. Microbes in the gut might take up microRNAs (miRNAs) and these miRNAs might affect microbes in turn. Our previous work identified miR-139-5p as a tumor suppressor gene down-regulated in CRC. At present, the regulatory role and mechanism of miR-139-5p between Fusobacterium nucleatum and CRC are unclear. In this study, after co-incubating Fusobacterium nucleatum with CRC cells, MTT assay, colony formation assay and wound-healing assay showed that Fusobacterium nucleatum could stimulate cell proliferation and migration. After knocking down the expression of c-met in cells, western blot assay proved that knocking down c-met could weaken this stimulation. C-met is one of the target genes of miR-139-5p. Experimented with miR-139-5p overexpressed CRC cell lines, we found the same results as knocking down c-met, which means that endogenous miR-139-5p can reduce the stimulation. Next, by co-incubating the exogenous miR-139-5p mimics with Fusobacterium nucleatum, we proved that exogenous miR-139-5p could inhibit the proliferation of Fusobacterium nucleatum. After treating CRC cells with Fusobacterium nucleatum, which incubated with miR-139-5p mimics in advance, MTT assay indicated that the stimulation of Fusobacterium nucleatum was weakened. Besides, we speculated the binding site between miR-139-5p and Fusobacterium nucleatum. In sum, our study suggests a new prospect for the treatment of CRC, and the combination of Fusobacterium nucleatum and miR-139-5p could be used as a more valuable comprehensive biomarker for CRC prognosis.

Contribution of glutaminases to glutamine metabolism and acid resistance in Lactobacillus reuteri and other vertebrate host adapted lactobacilli.

The bacterial conversion of glutamine to glutamate is catalyzed by glutamine-amidotransferases or glutaminases. Glutamine deamination contributes to the formation of the bioactive metabolites glutamate, γ-aminobutyrate (GABA) and γ-glutamyl peptides, and to acid resistance. This study aimed to investigate the distribution of glutaminase(s) in lactobacilli, and to evaluate their contribution in L. reuteri to amino acid metabolism and acid resistance. Phylogenetic analysis of the glutaminases gls1, gls2 and gls3 in the genus Lactobacillus demonstrated that glutaminase is exclusively present in host-adapted species of lactobacilli. The disruption gls1, gls2 and gls3 in L. reuteri 100-23 had only a limited effect on the conversion of glutamine to glutamate, GABA, or γ-glutamyl peptides in sourdough. The disruption of all glutaminases in L. reuteri 100-23Δgls1Δgls2Δgls3 but not disruption of gls2 and gls3 eliminated the protective effect of glutamine on the survival of the strain at pH 2.5. Glutamine also enhanced acid resistance of L. reuteri 100-23ΔgadB and L. taiwanensis 107q, strains without glutamate decarboxylase activity. Taken together, the study demonstrates that glutaminases of lactobacilli do not contribute substantially to glutamine metabolism but enhance acid resistance. Their exclusive presence in host-adapted lactobacilli provides an additional link between the adaptation of lactobacilli to specific habitats and their functionality when used as probiotics and starter cultures.