Mechanism of transforming growth factor-beta1-induced expression of vascular endothelial growth factor in murine osteoblastic MC3T3-E1 cells.
Transforming growth factor-beta1 (TGF-beta1), an abundant growth factor in bone matrix, has been shown to be involved in bone formation and fracture healing. The mechanism of action of the osteogenic effect of TGF-beta1 is not clearly understood. In this study, we found that the addition of TGF-beta1 to murine osteoblastic MC3T3-E1 cells induced vascular endothelial growth factor (VEGF) mRNA production. VEGF mRNA levels reached a plateau within 2 h after the addition of TGF-beta1. The induction was superinduced by cycloheximide and blocked by actinomycin D. Ro 31-8220, a protein kinase C inhibitor, abrogated the induction. In addition, curcumin, an inhibitor for transcription factor AP-1, also blocked the induction. Electrophoretic mobility shift assay revealed an enhanced binding of transcription factors AP-1 and NF-kappaB. Transient transfection experiment showed that VEGF promoter activity increased 3.6-fold upon TGF-beta1 stimulation. Immunoblot analysis showed that the amount of secreted VEGF was elevated in the medium 4 h after TGF-beta1 stimulation. Our results therefore suggest that at least part of the osteogenic activity of TGF-beta1 may be attributed to the production of VEGF.
Chua
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Cecile Cox Quillen Laboratory of Geriatric Research, James Quillen School of Medicine, East Tennessee State University, Johnson City, TN 37614, USA. chuab@xtn.net
Hamdy
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Chua
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D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D003474:Curcumin / Q000494:pharmacology; D003513:Cycloheximide / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D016228:Endothelial Growth Factors / Q000235:genetics* / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D007211:Indoles / Q000494:pharmacology; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008222:Lymphokines / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D016328:NF-kappa B / Q000187:drug effects / Q000378:metabolism; D010006:Osteoblasts / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D012333:RNA, Messenger / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D018808:Transcription Factor AP-1 / Q000187:drug effects / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D016228:Endothelial Growth Factors; D004791:Enzyme Inhibitors; D007211:Indoles; D008222:Lymphokines; D016328:NF-kappa B; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D018808:Transcription Factor AP-1; D016212:Transforming Growth Factor beta; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D003609:Dactinomycin; D003513:Cycloheximide; D008156:Luciferases; D011493:Protein Kinase C; D003474:Curcumin; C064758:Ro 31-8220