Moon-Kyoung Bae

Visfatin enhances ICAM-1 and VCAM-1 expression through ROS-dependent NF-κB activation in endothelial cells

Visfatin has recently been identified as a novel visceral adipokine which may be involved in obesity-related vascular disorders. However, it is not known whether visfatin directly contributes to endothelial dysfunction. Here, we investigated the effect of visfatin on vascular inflammation, a key step in a variety of vascular diseases. Visfatin induced leukocyte adhesion to endothelial cells and the aortic endothelium by induction of the cell adhesion molecules, ICAM-1 and VCAM-1. Promoter analysis revealed that visfatin-mediated induction of CAMs is mainly regulated by nuclear factor-kappaB (NF-kappaB). Visfatin stimulated IkappaBalpha phosphorylation, nuclear translocation of the p65 subunit of NF-kappaB, and NF-kappaB DNA binding activity in HMECs. Furthermore, visfatin increased ROS generation, and visfatin-induced CAMs expression and NF-kappaB activation were abrogated in the presence of the direct scavenger of ROS. Taken together, our results demonstrate that visfatin is a vascular inflammatory molecule that increases expression of the inflammatory CAMs, ICAM-1 and VCAM-1, through ROS-dependent NF-kappaB activation in endothelial cells.

Hypoxic induction of human visfatin gene is directly mediated by hypoxia-inducible factor-1

Visfatin has been originally identified as a growth factor for early stage B cells and recently known as an adipokine. Here, we report that hypoxia induces the visfatin mRNA and protein levels in MCF7 breast cancer cells. We also demonstrate that induction of visfatin gene is regulated by hypoxia‐inducible factor‐1α (HIF‐1α). Moreover, 5′‐flanking promoter region of human visfatin gene contains two functional HIF responsive elements (HREs), activating the expression of visfatin. Mutation of these HREs in the visfatin promoter abrogates activation of a luciferase reporter gene driven by visfatin promoter under hypoxia. Taken together, our results demonstrate that visfatin is a new hypoxia‐inducible gene of which expression is stimulated through the interaction of HIF‐1 with HRE sites in its promoter region.

Resveratrol inhibits Porphyromonas gingivalis lipopolysaccharide-induced endothelial adhesion molecule expression by suppressing NF-κB activation

P. gingivalis is a major pathogen that is involved in the onset and progression of periodontal disease. This study investigated the effect of resveratrol, a naturally occurring polyphenol, on P. gingivalis LPS-accelerated vascular inflammation, a key step in the progression of periodontitis. Resveratrol significantly inhibited the P. gingivalis LPS-induced adhesion of leukocytes to endothelial cells and to the aortic endothelium by down-regulating the cell adhesion molecules, ICAM-1 and VCAM-1. Moreover, the inhibition of the P. gingivalis LPS-induced cell adhesion molecules by resveratrol was mainly mediated by nuclear factor-κB (NF-κB). Resveratrol suppressed P. gingivalis LPS-stimulated IκBα phosphorylation and nuclear translocation of the p65 subunit of NF-κB in HMECs. Overall, these findings suggest that resveratrol significantly attenuates the P. gingivalis LPS-induced monocyte adhesion to the endothelium by suppressing the expression of the NF-κB-dependent cell adhesion molecules, suggesting its therapeutic role in periodontal pathogen-induced vascular inflammation.

Upregulation of fibroblast growth factor-2 by visfatin that promotes endothelial angiogenesis.

Adipokines have been known to act as angiogenic regulators in the process of angiogenesis. Recently, we have demonstrated that visfatin, a novel adipokine, has angiogenic activity. However, little has been reported on the underlying mechanism of visfatin-induced angiogenesis. In this study, we report that visfatin-induced angiogenesis is mediated by endothelial fibroblast growth factor-2 (FGF-2). Visfatin increased the levels of FGF-2 mRNA and protein in human endothelial cells. The enhancement in FGF-2 expression was prevented by an inhibitor of the extracellular signal-regulated kinase 1/2 (Erk1/2) pathway. Furthermore, visfatin-induced angiogenesis was reduced by inhibition of FGF-2 receptor kinase or by neutralization of FGF-2 function. Taken together, our results indicate that visfatin-induced endothelial angiogenesis is composed largely of two sequential steps: the induction of Erk1/2-dependent FGF-2 gene expression by visfatin and the subsequent FGF-2-induced angiogenesis. These data further suggest an integral role for visfatin-FGF-2 signaling axis in modulating endothelial angiogenesis.

Curcumin Down-Regulates Visfatin Expression and Inhibits Breast Cancer Cell Invasion

Obesity is frequently associated with breast cancer. Such associations are possibly mediated by adipokines. Visfatin, an adipokine, has recently been shown to be related to the development and progression of breast cancer. Therefore, the down-regulation of visfatin may be a novel strategy for breast cancer therapy. Curcumin has anticancer activities by modulating multiple signaling pathways and genes. The purpose of this study was to investigate whether visfatin gene expression is affected by curcumin in human breast cancer cells and to characterize the functional role of visfatin in breast cancer. We found that the mRNA and protein levels of visfatin were down-regulated by curcumin in MDA-MB-231, MDA-MB-468, and MCF-7 breast cancer cells, along with decreased activity of constitutive nuclear factor (NF)-κB. We confirmed the repressive effect of curcumin on visfatin transcription by performing a visfatin promoter-driven reporter assay and identified two putative NF-κB-binding sites on visfatin promoter that are important for this effect. EMSA and chromatin immunoprecipitation analysis indicated the binding of p65 to the visfatin promoter, which was effectively blocked by curcumin. Enforced expression of p65 protein increased visfatin promoter activity, whereas blocking NF-κB signaling suppressed visfatin gene expression. Visfatin could enhance the invasion of MDA-MB-231 cells and also attenuate curcumin-induced inhibition of cell invasion; on the other hand, visfatin knockdown by small interfering RNA led to the reduction of cell invasion. Our data demonstrate, for the first time, that curcumin down-regulates visfatin gene expression in human breast cancer cells by a mechanism that is, at least in part, NF-κB dependent and suggest that visfatin may contribute to breast cancer cell invasion and link obesity to breast cancer development and progression.

Aspirin induces apoptosis through the blockade of IL-6-STAT3 signaling pathway in human glioblastoma A172 cells

Aspirin has been reported to induce apoptosis in various cancer cell lines. However, the apoptotic effects of aspirin on human brain tumor cells are not well understood. Here, we have assessed the effect of aspirin on human gliobalstoma cell line A172 and found that aspirin induced the apoptosis of A172 cells, as determined by TUNEL assay, FACS analysis, and Hoechst staining. The underlying mechanism of this effect consists of reduction in the level of phosphorylated STAT3 (Tyr705), a transcription factor required for survival of A172 cells. Moreover, the expression of STAT3 target genes such as Cyclin D1, XIAP, and Bcl-2 that are essential for cell growth and survival was apparently attenuated after aspirin treatment. We also showed that the expression and secretion of interleukin-6 (IL-6), leading to STAT3 phosphorylation, was inhibited by aspirin. When administered exogenous IL-6 to aspirin-treated A172 cells, the phosphorylation of STAT3 and cellular apoptosis were restrained compared to aspirin only-treated cells. Taken together, our results indicate that aspirin causes apoptosis via down-regulation of IL-6-dependent STAT3 signaling, suggesting that aspirin could be therapeutically useful for a potential anti-glioblastoma therapeutic approach.

Visfatin through STAT3 activation enhances IL-6 expression that promotes endothelial angiogenesis

Signal transducer and activator of transcription 3 (STAT3) acts as a mediator and biomarker in endothelial activation. We have recently shown that a novel adipokine visfatin promotes endothelial angiogenesis. The present study was to determine whether visfatin affects STAT3 activity and to explore the potential target gene(s). Here, we found that visfatin induced the activation of STAT3, as characterized by increased tyrosine phosphorylation, nuclear translocation, and DNA-binding activity in human endothelial cells. In addition, visfatin significantly upregulated mRNA and protein levels of endothelial interleukin-6 (IL-6), which was blocked by a specific inhibitor of STAT3 signaling and by the transfection of siRNA specific for STAT3. Furthermore, visfatin-induced angiogenesis was reduced by the inhibition of STAT3 signaling or neutralization of IL-6 function, as measured by tube formation, rat aortic ring assay, and mouse Matrigel plug assay. Taken together, our results provide the first example of STAT3-dependent endothelial IL-6 induction by visfatin and of the role of IL-6 in mediating visfatin-induced angiogenesis.

Tid-1 Interacts with the von Hippel-Lindau Protein and Modulates Angiogenesis by Destabilization of HIF-1α

The von Hippel-Lindau protein (pVHL) is a major tumor suppressor protein and also associated with the inhibition of angiogenesis via HIF-1alpha ubiquitination and proteasomal degradation. To further elucidate the biological activity of pVHL in angiogenesis, pVHL-interacting proteins were screened using the yeast two-hybrid system. We found that a mouse homologue of the long form of Drosophila tumor suppressor l(2)tid, Tid-1(L), directly interacts with pVHL in vitro and in vivo. Furthermore, Tid-1(L) protein; enhanced the interaction between HIF-1alpha and pVHL, leading to the destabilization of HIF-1alpha protein; therefore, Tid-1(L) protein decreased vascular endothelial growth factor expression and inhibited angiogenesis in vivo and in vitro. These findings propose that Tid-1(L) may play a critical role in pVHL-mediated tumor suppression by modulating the pVHL-dependent HIF-1alpha stability.

Regulation of Egr-1 by association with the proteasome component C8

Primary response transcription factor, Egr-1, is rapidly activated by a variety of extracellular stimuli. Activation of Egr-1 is shown to function as a master switch activated by ischemia to trigger expression of pivotal regulators of inflammation, coagulation and vascular hyperpermeability. Egr-1 is a short-lived protein, but the mechanism that regulates its stability has not yet been clarified. In this study, the yeast two-hybrid screening revealed that Egr-1 interacts significantly with PRC8 (proteasome component C8) and the specific interaction was confirmed by GST pull-down assay and coimmunoprecipitation. Interestingly, we found that the PRC8-mediated regulation of Egr-1 activity is associated with the proteasome pathway and PRC8 inhibits the transcriptional activity of Egr-1. In addition, Egr-1 protein was specifically multiubiquitinated by ubiquitin. These data strongly imply that Egr-1 protein is targeted for proteolysis by the ubiquitin-dependent proteasome pathway.

Neuromedin B induces angiogenesis via activation of ERK and Akt in endothelial cells

Neuromedin B (NMB) is one of the bombesin-like peptides in mammals. Recently, bombesin-like peptides have been characterized as growth factors in highly vascularized tumors. In this study, we report that NMB potently stimulates in vivo neovascularization in a mouse Matrigel plug and the sprouting of endothelial cells ex vivo in rat aortic rings. In addition, NMB increases the migration and tube formation in human umbilical vein endothelial cells (HUVECs). Moreover, treatment of HUVECs with NMB activates the extracellular signal-regulated kinase 1/2 (ERK(1/2)), Akt, and endothelial nitric oxide synthase (eNOS) and increases the level of NO production in a dose- and time-dependent manner. Furthermore, ERK activation and angiogenic sprouting in response to NMB are significantly blocked by the MEK inhibitor. Inhibition of phosphatidylinositol 3-kinase (PI3K) suppresses the NMB-stimulated tubular formation of HUVECs, along with reduction in the phosphorylation of Akt and eNOS. Taken together, these results indicate that NMB is a novel angiogenic peptide, and its angiogenic activity is mediated by activating the MEK/ERK- and PI3K/Akt/eNOS-dependent pathways. This study suggests that NMB may play important roles in mediating a variety of pathophysiological angiogenesis.