Abstract
The potential role of hepatocyte growth factor (HGF) in the regulation of angiogenesis factors in hepatoma cells is not widely appreciated. We investigated the role of HGF-induced activation of a transcription factor, Egr-1, in the expression of pro-angiogenic factors. Genes associated with angiogenesis induced by HGF were screened by using cDNA microarray technology in hepatocellular carcinoma cell lines, HepG2 and Hep3B. Expression levels of Egr-1, vascular endothelial growth factor (VEGF), and interleukin (IL)-8 were further confirmed by real time RT-PCR and Western blot analysis. Roles of Egr-1 in the levels of HGF-induced up-regulations of VEGF and IL-8 were measured by knockdown of Egr-1 with Egr-1 shRNA and chromatin immunoprecipitation assay. The levels of Egr-1, VEGF and IL-8 were up-regulated in cells treated with HGF. HGF-induced up-regulations of Egr-1, VEGF, and IL-8 were inhibited by the pretreatment with an MEK inhibitor, PD098059. HGF-induced up-regulation of VEGF and IL-8 were repressed by Egr-1 knockdown. HGF enhanced the binding activity of Egr-1 to the VEGF promoter in control cells, but not in the Egr-1-shRNA cells. No constitutive and inducible Egr-1 binding activities to the IL-8 promoter were observed in control and Egr-1-shRNA cells. Egr-1 knockdown reduced the luciferase activities increased by HGF not in the IL-8 promoter, but in the VEGF promoter. Egr-1 might play an important role in the up-regulation of VEGF and IL-8 induced by HGF and contribute to HGF-mediated angiogenesis, which might be promising targets for hepatocellular carcinoma therapy.
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Abbreviations
- HGF:
-
Hepatocyte growth factor
- VEGF:
-
Vascular endothelial growth factor
- Egr-1:
-
Early growth response gene-1
References
Jeffers M, Rong S, Vande GF (1996) Enhanced tumorigenicity and invasion—metastasis by hepatocyte growth factor/scatter factor-met signaling in human cells concomitant with induction of the urokinase proteolysis network. Mol Cell Biol 16:1115–1125
Comoglio PM, Boccaccio C (2001) Scatter factors and invasive growth. Semin Cancer Biol 11:153–165. doi:10.1006/scbi.2000.0366
Vejchapipat P, Tangkijvanich P, Theamboonlers A, Chongsrisawat V, Chittmittrapap S, Poovorawan Y (2004) Association between serum hepatocyte growth factor and survival in untreated hepatocellular carcinoma. J Gastroenterol 39:1182–1188. doi:10.1007/s00535-004-1469-8
Brian W, Xin PY, Tin LL, Lorena B, Ning TY, Joshua GC, Carter VW, Zhong C (2005) Hepatocyte growth factor/scatter factor differentially regulates expression of proangiogenic factors through Egr-1 in head and neck squamous cell carcinoma. Cancer Res 65(16):7071–7080. doi:10.1158/0008-5472.CAN-04-0989
Ninck S, Reisser C, Dyckhoff G, Helmke B, Bauer H, Herold-Mende C (2003) Expression profiles of angiogenic growth factors in squamous cell carcinomas of the head and neck. Int J Cancer 106:34–44. doi:10.1002/ijc.11188
Riedel F, Gotte K, Schwalb J, Wirtz H, Bergler W, Hormann K (2000) Serum levels off vascular endothelial growth factor in patients with head and neck cancer. Eur Arch Otorhinolaryngol 257:332–336. doi:10.1007/s004059900208
Dong G, Chen Z, Li ZY, Yeh NT, Bancroft CC, Van WC (2001) Hepatocyte growth factor/scatter factor-induced activation of MEK and PI3 K signal pathways contributes to expression of proangiogenic cytokines interleukin-8 and vascular endothelial growth factor in head and neck squamous cell carcinoma. Cancer Res 61:5911–5918
Zhang YW, Su Y, Volpert OV, Vande Woude GF (2003) Hepatocyte growth factor/scatter factor mediates angiogenesis through positive VEGF and negative thrombospondin 1 regulation. Proc Natl Acad Sci USA 100:12718–12723. doi:10.1073/pnas.2135113100
Bancroft CC, Chen Z, Yeh J et al (2002) Effects of pharmacologic antagonists of epidermal growth factor receptor, PI3 K and MEK signal kinases on NF-kB and AP-1 activation and IL-8 and VEGF expression in human head and neck squamous cell carcinoma lines. Int J Cancer 99:538–548. doi:10.1002/ijc.10398
Bancroft CC, Chen Z, Dong G et al (2001) Coexpression of proangiogenic factors IL-8 and VEGF by human head and neck squamous cell carcinoma involves coactivation by MEK-MAPK and IKK-NF-kB signal pathways. Clin Cancer Res 7:435–442
Sukhatme VP, Cao X, Chang LC, Tsai-Morris C-H, Stamenkovich D, Ferreira PCP, Cohen DR, Edwards SA, Shows TB, Curran T, Le Beau MM, Adamson ED (1988) A zinc finger-encoding gene coregulated with c-fos during growth and differentiation, and after cellular depolarization. Cell 53:37–43. doi:10.1016/0092-8674(88)90485-0
Chavrier P, Zerial M, Lemaire P, Almendral J, Bravo R, Charnay P (1988) A gene encoding a protein with zinc fingers is activated during G0/G1 transition in cultured cells. EMBO J 7:29–35
Baek SJ, Kim JS, Moore SM, Lee SH, Martinez J, Eling TE (2005) Cyclooxygenase inhibitors induce the expression of the tumor suppressor gene EGR-1, which results in the up-regulation of NAG-1, an antitumorigenic protein. Mol Pharmacol 67:356–364. doi:10.1124/mol.104.005108
Fahmy RG, Dass CR, Sun LQ, Chesterman CN, Khachigian LM (2003) Transcription Factor Egr-1 supports FGF-dependent angiogenesis during neovascularization and tumor growth. Nat Med 9:1026–1032. doi:10.1038/nm905
Yang SH, Kim JS, Oh TJ, Kim MS, Lee SW, Woo SK, Cho HS, Choi YH, Kim YH, Rha SY, Chung HC, An SW (2003) Genome-scale analysis of resveratrol-induced gene expression profile in human ovarian cancer cells using a cDNA microarray. Int J Oncol 22:741–750
Kyung HL, Eun YC, Min KK, Byung IJ, Tae NK, Sang WK, Sun KS, Jung HK, Jae RK (2006) Regulation of hepatocyte growth factor-mediated urokinase plasminogen activator secretion by MEK/ERK activation in human stomach cancer cell lines. Exp Mol Med 38(1):27–35
Schug J, Overton GC (1977) TESS: transcription element search software. Technical Report CBIL-TR-1997-1001-v0.0 of the Computational Biology and Informatics Laboratory, School of Medicine, University of Pennsylania
Bussolino F, Di Renzo MF, Ziche M, Bocchietto E, Olivero M, Naldini L, Gaudino G, Tamagnone L, Coffer A, Comoglio PM (1992) Hepatocyte growth factor is a potent angiogenic factor which stimulates endothelial cell motility and growth. J Cell Biol 119:629–641. doi:10.1083/jcb.119.3.629
Grant DS, Kleinman HK, Goldberg ID, Bhargava MM, Nickoloff BJ, Kinsella JL, Polverini P, Rosen EM (1993) Scatter factor induces blood vessel formation in vivo. Proc Natl Acad Sci USA 90:1937–1941. doi:10.1073/pnas.90.5.1937
Lim RW, Varnum BC, Herschman HR (1987) Cloning of tetradecanoyl phorbol ester-induced ‘primary response’ sequences and their expression in density-arrested Swiss 3T3 cells and a TPA non-proliferative variant. Oncogene 1(3):263–270
Cao XM, Koski RA, Gashler A et al (1990) Identification and characterization of the Egr-1 gene product, a DNA-binding zinc finger protein induced by differentiation and growth signals. Mol Cell Biol 10:1931–1939
Herrlich P, Morrison H, Sleeman J, Orian-Rousseau V, Konin H, Weg-Remers S, Ponta H (2000) CD44 acts both as a growth-and invasiveness-promoting molecule and as a tumor-suppressing cofactor. Ann N Y Acad Sci 910:106–120
Hyun AH, Gu L, Hee JK, Yonh GK, Sung HB, Jae LL, Kyung HL, Dong SK (2003) Overexpression of c-Met protein in gastric cancer and role of uPAR as a therapeutic target. Cancer Res Treat 35(1):9–15
Juan AR, Glenn M (2003) Hepatocyte growth factor/scatter factor induces feedback up-regulation of CD44v6 in melanoma cells through Egr-1. Cancer Res 63:1576–1582
Tolnay E, Kuhnen C, Wiethege T, Konig JE, Voss B, Mmuller KM (1998) Hepatocyte growth factor/scatter factor and its receptor c-met are overexpressed and associated with an increased microvessel density in malignant pleural mesothelioma. J Cancer Res Clin Oncol 124:291–296. doi:10.1007/s004320050171
Schmidt NO, Westphal M, Hagel C, Ergun S, Stavrou D, Rosen EM, Lamszus K (1999) Levels of vascular endothelial growth factor, hepatocyte growth factor/scatter factor and vasic fibroblast growth factor in human gliomas and their relation to angiogenesis. Int J Cancer 84:10–18. doi:10.1002/(SICI)1097-0215(19990219)84:1<10::AID-IJC3>3.0.CO;2-L
Khachigian LM, Lindner V, Williams AJ, Collins T (1996) Egr-1 induced endothelial gene expression: a common theme in vascular injury. Science 271:1427–1431. doi:10.1126/science.271.5254.1427
Silverman ES, Khachigian LM, Lindner V, Williams AJ, Collins T (1997) Inducible PDGF A-chain transcription in smooth muscle cells is mediated by Egr-1 displacement of Sp1 and Sp3. Am J Physiol 273:H1415–H1426
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“This work was supported by the Korea Science and Engineering Foundation(KOSEF) NCRC grant funded by the Korea government(MEST) (No. R15-2004-033-05001-0)”
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Lee, K.H., Kim, JR. Hepatocyte growth factor induced up-regulations of VEGF through Egr-1 in hepatocellular carcinoma cells. Clin Exp Metastasis 26, 685–692 (2009). https://doi.org/10.1007/s10585-009-9266-7
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DOI: https://doi.org/10.1007/s10585-009-9266-7