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Title: Kruppel-like factor 2 inhibit the angiogenesis of cultured human liver sinusoidal endothelial cells through the ERK1/2 signaling pathway

Abstract

Kruppel-like factor 2 (KLF2) is a crucial anti-angiogenic factor. However, its precise role in hepatic angiogenesis induced by liver sinusoidal endothelial cells (LSECs) remain unclear. This study was aimed to evaluate the effect of KLF2 on angiogenesis of LSECs and to explore the corresponding mechanism. Cultured human LSECs were infected with different lentiviruses to overexpress or suppress KLF2 expression. The CCK-8 assay, transwell migration assay and tube formation test, were used to investigate the roles of KLF2 in the proliferation, migration and vessel tube formation of LSECs, respectively. The expression and phosphorylation of ERK1/2 were detected by western blot. We discovered that the up-regulation of KLF2 expression dramatically inhibited proliferation, migration and tube formation in treated LSECs. Correspondingly, down-regulation of KLF2 expression significantly promoted proliferation, migration and tube formation in treated LSECs. Additionally, KLF2 inhibited the phosphorylation of ERK1/2 pathway, followed by the function of KLF2 in the angiogenesis of LSECs disrupted. In conclusion, KLF2 suppressed the angiogenesis of LSECs through inhibition of cell proliferation, migration, and vessel tube formation. These functions of KLF2 may be mediated through the ERK1/2 signaling pathway. - Highlights: • Overexpression of KLF2 inhibits the proliferation and migration of LSECs. • Overexpression of KLF2 inhibitsmore » the angiogenesis of LSECs. • ERK1/2 signaling pathway involved in the anti-angiogenic process of KLF2 on LSECs.« less

Authors:
 [1];  [1];  [1];  [1];  [1];  [1];  [1];  [1];  [1];  [1];  [2]
  1. Department of Gastroenterology of Zhongshan Hospital, Fudan University, Shanghai (China)
  2. (China)
Publication Date:
OSTI Identifier:
22462244
Resource Type:
Journal Article
Resource Relation:
Journal Name: Biochemical and Biophysical Research Communications; Journal Volume: 464; Journal Issue: 4; Other Information: Copyright (c) 2015 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved.; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; ANGIOGENESIS; CELL PROLIFERATION; HUMAN POPULATIONS; HYPERTENSION; INHIBITION; LIVER; MIGRATION; PHOSPHORYLATION; SIGNALS; TUBES

Citation Formats

Zeng, Xiao-Qing, E-mail: zeng.xiaoqing@zs-hospital.sh.cn, Li, Na, E-mail: Linala.2009@163.com, Pan, Du-Yi, E-mail: lasikesmi@hotmail.com, Miao, Qing, E-mail: sadsadvenus@163.com, Ma, Gui-Fen, E-mail: ma.guifen@zs-hospital.sh.cn, Liu, Yi-Mei, E-mail: liuyimei1988@163.com, Tseng, Yu-Jen, E-mail: dianatseng14@gmail.com, Li, Feng, E-mail: li.feng2@zs-hospital.sh.cn, Xu, Li-Li, E-mail: xu.lili3@zs-hospital.sh.cn, Chen, Shi-Yao, E-mail: chen.shiyao@zs-hospital.sh.cn, and Institute of Endoscopic Research of Zhongshan Hospital, Fudan University, Shanghai. Kruppel-like factor 2 inhibit the angiogenesis of cultured human liver sinusoidal endothelial cells through the ERK1/2 signaling pathway. United States: N. p., 2015. Web. doi:10.1016/J.BBRC.2015.07.113.
Zeng, Xiao-Qing, E-mail: zeng.xiaoqing@zs-hospital.sh.cn, Li, Na, E-mail: Linala.2009@163.com, Pan, Du-Yi, E-mail: lasikesmi@hotmail.com, Miao, Qing, E-mail: sadsadvenus@163.com, Ma, Gui-Fen, E-mail: ma.guifen@zs-hospital.sh.cn, Liu, Yi-Mei, E-mail: liuyimei1988@163.com, Tseng, Yu-Jen, E-mail: dianatseng14@gmail.com, Li, Feng, E-mail: li.feng2@zs-hospital.sh.cn, Xu, Li-Li, E-mail: xu.lili3@zs-hospital.sh.cn, Chen, Shi-Yao, E-mail: chen.shiyao@zs-hospital.sh.cn, & Institute of Endoscopic Research of Zhongshan Hospital, Fudan University, Shanghai. Kruppel-like factor 2 inhibit the angiogenesis of cultured human liver sinusoidal endothelial cells through the ERK1/2 signaling pathway. United States. doi:10.1016/J.BBRC.2015.07.113.
Zeng, Xiao-Qing, E-mail: zeng.xiaoqing@zs-hospital.sh.cn, Li, Na, E-mail: Linala.2009@163.com, Pan, Du-Yi, E-mail: lasikesmi@hotmail.com, Miao, Qing, E-mail: sadsadvenus@163.com, Ma, Gui-Fen, E-mail: ma.guifen@zs-hospital.sh.cn, Liu, Yi-Mei, E-mail: liuyimei1988@163.com, Tseng, Yu-Jen, E-mail: dianatseng14@gmail.com, Li, Feng, E-mail: li.feng2@zs-hospital.sh.cn, Xu, Li-Li, E-mail: xu.lili3@zs-hospital.sh.cn, Chen, Shi-Yao, E-mail: chen.shiyao@zs-hospital.sh.cn, and Institute of Endoscopic Research of Zhongshan Hospital, Fudan University, Shanghai. Fri . "Kruppel-like factor 2 inhibit the angiogenesis of cultured human liver sinusoidal endothelial cells through the ERK1/2 signaling pathway". United States. doi:10.1016/J.BBRC.2015.07.113.
@article{osti_22462244,
title = {Kruppel-like factor 2 inhibit the angiogenesis of cultured human liver sinusoidal endothelial cells through the ERK1/2 signaling pathway},
author = {Zeng, Xiao-Qing, E-mail: zeng.xiaoqing@zs-hospital.sh.cn and Li, Na, E-mail: Linala.2009@163.com and Pan, Du-Yi, E-mail: lasikesmi@hotmail.com and Miao, Qing, E-mail: sadsadvenus@163.com and Ma, Gui-Fen, E-mail: ma.guifen@zs-hospital.sh.cn and Liu, Yi-Mei, E-mail: liuyimei1988@163.com and Tseng, Yu-Jen, E-mail: dianatseng14@gmail.com and Li, Feng, E-mail: li.feng2@zs-hospital.sh.cn and Xu, Li-Li, E-mail: xu.lili3@zs-hospital.sh.cn and Chen, Shi-Yao, E-mail: chen.shiyao@zs-hospital.sh.cn and Institute of Endoscopic Research of Zhongshan Hospital, Fudan University, Shanghai},
abstractNote = {Kruppel-like factor 2 (KLF2) is a crucial anti-angiogenic factor. However, its precise role in hepatic angiogenesis induced by liver sinusoidal endothelial cells (LSECs) remain unclear. This study was aimed to evaluate the effect of KLF2 on angiogenesis of LSECs and to explore the corresponding mechanism. Cultured human LSECs were infected with different lentiviruses to overexpress or suppress KLF2 expression. The CCK-8 assay, transwell migration assay and tube formation test, were used to investigate the roles of KLF2 in the proliferation, migration and vessel tube formation of LSECs, respectively. The expression and phosphorylation of ERK1/2 were detected by western blot. We discovered that the up-regulation of KLF2 expression dramatically inhibited proliferation, migration and tube formation in treated LSECs. Correspondingly, down-regulation of KLF2 expression significantly promoted proliferation, migration and tube formation in treated LSECs. Additionally, KLF2 inhibited the phosphorylation of ERK1/2 pathway, followed by the function of KLF2 in the angiogenesis of LSECs disrupted. In conclusion, KLF2 suppressed the angiogenesis of LSECs through inhibition of cell proliferation, migration, and vessel tube formation. These functions of KLF2 may be mediated through the ERK1/2 signaling pathway. - Highlights: • Overexpression of KLF2 inhibits the proliferation and migration of LSECs. • Overexpression of KLF2 inhibits the angiogenesis of LSECs. • ERK1/2 signaling pathway involved in the anti-angiogenic process of KLF2 on LSECs.},
doi = {10.1016/J.BBRC.2015.07.113},
journal = {Biochemical and Biophysical Research Communications},
number = 4,
volume = 464,
place = {United States},
year = {Fri Sep 04 00:00:00 EDT 2015},
month = {Fri Sep 04 00:00:00 EDT 2015}
}
  • Background: Angiogenesis plays a major role in the pathogenesis of inflammatory bowel disease (IBD). Placental growth factor (PlGF) is a specific regulator of pathological angiogenesis and is upregulated in the sera of IBD patients. Therefore, the role of PlGF in IBD angiogenesis was investigated here using HIMECs. Methods: The expression of PlGF and its receptors in human intestinal microvascular endothelial cells (HIMECs) and inflamed mucosa of IBD patients were examined using quantitative PCR and western blot analysis and the role of PlGF in IBD HIMECs was further explored using small interfering RNA (siRNA). The induction of pro-inflammatory cytokine by PlGFmore » in HIMECs was confirmed by ELISA. The capacity of PlGF to induce angiogenesis in HIMECs was tested through proliferation, cell-migration, matrigel tubule-formation assays and its underlying signaling pathway were explored by western blot analysis of ERK1/2 and PI3K/Akt phosphorylation. Results: mRNA and protein expression of PlGF and its receptor NRP-1 were significantly increased in IBD HIMECs. Inflamed mucosa of IBD patients also displayed higher expression of PIGF. The production of IL-6 and TNF-α in culture supernatant of HIMECs treated with exogenous recombinant human PlGF-1 (rhPlGF-1) were increased. Furthermore, rhPlGF-1 significantly induced HIMECs migration and tube formation in a dose-dependent manner and knockdown of endogenous PlGF in IBD HIMECs using siRNA substantially reduced these angiogenesis activities. PlGF induced PI3K/Akt phosphorylation in HIMECs and pretreatment of PlGF-stimulated HIMECs with PI3K inhibitor (LY294002) significantly inhibited the PlGF-induced cell migration and tube formation. Conclusion: Our results demonstrated the pro-inflammatory and angiogenic effects of PlGF on HIMECs in IBD through activation of PI3K/Akt signaling pathway. PlGF/PI3K/Akt signaling may serve as a potential therapeutic target for IBD. - Highlights: • Expression of PlGF and its receptor NRP-1 were significantly increased in IBD HIMECs. • Exogenous rhPlGF-1 treatment significantly induced HIMECs migration and tube formation. • Knockdown of endogenous PlGF in IBD HIMECs using siRNA substantially reduced cell angiogenesis activities. • PlGF induced PI3K/Akt phosphorylation in HIMECs which is required for PIGF-induced cell migration and tube formation.« less
  • Arsenic exposure represents a major health concern increasing cancer risks, yet the mechanism of arsenic carcinogenesis has not been elucidated. We and others recently reported that cell malignant transformation by arsenic is accompanied by epithelial to mesenchymal transition (EMT). However, the role of EMT in arsenic carcinogenesis is not well understood. Although previous studies showed that short term exposure of endothelial cells to arsenic stimulated angiogenesis, it remains to be determined whether cells that were malignantly transformed by long term arsenic exposure have a pro-angiogenic effect. The objective of this study was to investigate the effect of arsenic-transformed human bronchialmore » epithelial cells that underwent EMT on angiogenesis and the underlying mechanism. It was found that the conditioned medium from arsenic-transformed cells strongly stimulated tube formation by human umbilical vein endothelial cells (HUVECs). Moreover, enhanced angiogenesis was detected in mouse xenograft tumor tissues resulting from inoculation of arsenic-transformed cells. Mechanistic studies revealed that β-catenin was activated in arsenic-transformed cells up-regulating its target gene expression including angiogenic-stimulating vascular endothelial growth factor (VEGF). Stably expressing microRNA-200b in arsenic-transformed cells that reversed EMT inhibited β-catenin activation, decreased VEGF expression and reduced tube formation by HUVECs. SiRNA knockdown β-catenin decreased VEGF expression. Adding a VEGF neutralizing antibody into the conditioned medium from arsenic-transformed cells impaired tube formation by HUVECs. Reverse transcriptase-PCR analysis revealed that the mRNA levels of canonical Wnt ligands were not increased in arsenic-transformed cells. These findings suggest that EMT in arsenic-transformed cells promotes angiogenesis through activating β-catenin–VEGF pathway. - Highlights: • Arsenic-transformed cells that underwent EMT displayed a pro-angiogenic effect. • EMT in arsenic-transformed cells activates β-catenin. • β-Catenin activation increases VEGF expression in arsenic-transformed cells. • β-Catenin activation is likely independent of canonical Wnt signaling. • EMT in arsenic-transformed cells promotes angiogenesis via β-catenin–VEGF pathway.« less
  • Highlights: •H{sub 2}O{sub 2} inhibits TGF-β1-induced cell cycle arrest. •H{sub 2}O{sub 2} induces Smad3 linker phosphorylation through Akt-ERK1/2 pathway. •H{sub 2}O{sub 2}-mediated suppression of TGF-β signal requires Smad3 linker phosphorylation. •This is a first report about interplay between H{sub 2}O{sub 2} and growth inhibition pathway. -- Abstract: Hydrogen peroxide (H{sub 2}O{sub 2}) functions as a second messenger in growth factor receptor-mediated intracellular signaling cascade and is tumorigenic by virtue of its ability to promote cell proliferation; however, the mechanisms underlying the growth stimulatory action of H{sub 2}O{sub 2} are less understood. Here we report an important mechanism for antagonistic effectsmore » of H{sub 2}O{sub 2} on growth inhibitory response to transforming growth factor-β1 (TGF-β1). In Mv1Lu and HepG2 cells, pretreatment of H{sub 2}O{sub 2} (0.05–0.2 mM) completely blocked TGF-β1-mediated induction of p15{sup INK4B} expression and increase of its promoter activity. Interestingly, H{sub 2}O{sub 2} selectively suppressed the transcriptional activation potential of Smad3, not Smad2, in the absence of effects on TGF-β1-induced phosphorylation of the COOH-tail SSXS motif of Smad3 and its nuclear translocation. Mechanism studies showed that H{sub 2}O{sub 2} increases the phosphorylation of Smad3 at the middle linker region in a concentration- and time-dependent manner and this effect is mediated by activation of extracellular signal-activated kinase 1/2 through Akt. Furthermore, expression of a mutant Smad3 in which linker phosphorylation sites were ablated significantly abrogated the inhibitory effects of H{sub 2}O{sub 2} on TGF-β1-induced increase of p15{sup INK4B}-Luc reporter activity and blockade of cell cycle progression from G1 to S phase. These findings for the first time define H{sub 2}O{sub 2} as a signaling molecule that modulate Smad3 linker phosphorylation and its transcriptional activity, thus providing a potential mechanism whereby H{sub 2}O{sub 2} antagonizes the cytostatic function of TGF-β1.« less
  • Research highlights: {yields} Low-concentration oxidized LDL enhances angiogenesis through nitric oxide (NO). {yields} Oxidized LDL increases intracellular NO levels via eNOS phosphorylation. {yields} Akt/PI3K signaling mediates oxidized LDL-induced eNOS phosphorylation. -- Abstract: It has long been considered that oxidized low-density lipoprotein (oxLDL) causes endothelial dysfunction and is remarkably related to the development of atherosclerosis. However, the effect of oxLDL at very low concentration (<10 {mu}g/ml) on the endothelial cells remains speculative. Nitric oxide (NO) has a crucial role in the endothelial cell function. In this study, we investigated the effect of oxLDL at low concentration on NO production and proliferation,more » migration, tube formation of the human coronary artery endothelial cells (HCAEC). Results showed that oxLDL at 5 {mu}g/ml enhanced HCAEC proliferation, migration and tube formation. These phenomena were accompanied by an increased intracellular NO production. L-NAME (a NOS inhibitor), LY294002 and wortmannin (PI3K inhibitors) could abolish oxLDL-induced angiogenic effects and prevent NO production in the HCAEC. The phosphorylation of Akt, PI3K and eNOS were up-regulated by oxLDL, which was attenuated by LY294002. Our results suggested that oxLDL at low concentration could promote in-vitro angiogenesis and activate nitric oxide synthesis through PI3K/Akt/eNOS pathway in HCAEC.« less
  • Aim: The objective of this study was to investigate the effects of urotensin II (UII) treatment on the proliferation and collagen synthesis of cultured rat pulmonary arterial smooth muscle cells (PASMCs) and to explore whether these effects are mediated by mitogen-activated protein kinase (MAPK) signaling pathways and early growth response 1 (Egr-1). Methods: The proliferation of cultured PASMCs stimulated with different doses of UII was detected by BrdU incorporation. The mRNA expression levels of procollagen I (procol I), procollagen III (procol III), extracellular regulated protein kinase 1/2 (ERK1/2), stress-stimulated protein kinase (Sapk), p38 MAPK (p38), and Egr-1 mRNA in culturedmore » PASMCs after treatment with UII, the UII-specific antagonist urantide, and the ERK1/2 inhibitor PD98059 were detected by real-time polymerase chain reaction (PCR), and the protein expression levels of procol I, procol III, phosphorylated (p)-ERK1/2, p-Sapk, p-p38, and Egr-1 were detected by Western blotting. Results: Treatment with UII increased the proliferation of cultured PASMCs in a dose-dependent manner (P < 0.05). However, treatment with urantide and PD98059 inhibited the promoting effect of UII on PASMC proliferation (P < 0.05). Real-time PCR analysis showed that UII up-regulated the expression of procol I, procol III, ERK1/2, Sapk, and Egr-1 mRNA (P < 0.05), but not p38 mRNA. However, the up-regulating effect of UII was inhibited by PD98059 and urantide. Western blotting analysis showed that UII increased the synthesis of collagen I, collagen III, p-ERK1/2, p-Sapk, and Egr-1, and these effects also were inhibited by PD98059 and urantide (P < 0.05). Conclusions: Egr-1 participates in the UII-mediated proliferation and collagen synthesis of cultured rat PASMCs via activation of the ERK1/2 signaling pathway.« less