Growth factor-stimulated phospholipase D (PLD) catalyzes the hydrolysis of phosphatidylcholine (PC), generating phosphatidic acid (PA) which may act as a second messenger during cell proliferation and survival. Therefore, PLD is believed to play an important role in tumorigenesis. In this study, a potential mechanism for PLD-mediated tumorigenesis was explored. Ectopic expression of PLD1 or PLD2 in human glioma U87 cells increased the expression of hypoxia-inducible factor-1alpha (HIF-1alpha) protein. PLD-induced HIF-1 activation led to the secretion of vascular endothelial growth factor (VEGF), a HIF-1 target gene involved in tumorigenesis. PLD induction of HIF-1alpha was significantly attenuated by 1-butanol which blocks PA production by PLD, and PA per se was able to elevate HIF-1alpha protein level. Inhibition of mTOR, a PA-responsive kinase, reduced the levels of HIF-1alpha and VEGF in PLD-overexpressed cells. Epidermal growth factor activated PLD and increased the levels of HIF-1alpha and VEGF in U87 cells. A specific PLD inhibitor abolished expression of HIF-1alpha and secretion of VEGF. PLD may utilize HIF-1-VEGF pathway for PLD-mediated tumor cell proliferation and survival.
Cell Line, Tumor ; Epidermal Growth Factor/metabolism ; Gene Expression Regulation, Neoplastic ; Glioma/genetics/*metabolism ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit/genetics/metabolism ; Phosphatidic Acids/*metabolism ; Phospholipase D/genetics/*metabolism ; *Signal Transduction ; Transfection ; Vascular Endothelial Growth Factor A/*metabolism

A variety of benzylidenethiazole analogs have been demonstrated to inhibit 5-lipoxygenase (5-LOX). Here we report the anti-atherogenic potential of 5-(4-hydroxy-2,3,5-trimethylbenzylidene) thiazolidin-2,4-dione (HMB-TZD), a benzylidenethiazole analog, and its potential mechanism of action in LDL receptor-deficient (Ldlr-/-) mice. HMB-TZD Treatment reduced leukotriene B4 (LTB4) production significantly in RAW264.7 macrophages and SVEC4-10 endothelial cells. Macrophages or endothelial cells pre-incubated with HMB-TZD for 2 h and then stimulated with lipopolysaccharide or tumor necrosis factor-alpha (TNF-alpha) displayed reduced cytokine production. Also, HMB-TZD reduced cell migration and adhesion in accordance with decreased proinflammatory molecule production in vitro and ex vivo. HMB-TZD treatment of 8-week-old male Ldlr-/- mice resulted in significantly reduced atherosclerotic lesions without a change to plasma lipid profiles. Moreover, aortic expression of pro-atherogenic molecules involved in the recruitment of monocytes to the aortic wall, including TNF-alpha , MCP-1, and VCAM-1, was downregulated. HMB-TZD also reduced macrophage infiltration into atherosclerotic lesions. In conclusion, HMB-TZD ameliorates atherosclerotic lesion formation possibly by reducing the expression of proinflammatory molecules and monocyte/macrophage recruitment to the lesion. These results suggest that HMB-TZD, and benzylidenethiazole analogs in general, may have therapeutic potential as treatments for atherosclerosis.
Animals ; Atherosclerosis/*drug therapy ; Cell Adhesion/drug effects ; Cell Line ; Cell Movement/drug effects ; Chemokine CCL2/metabolism ; Dinoprostone/metabolism ; Enzyme-Linked Immunosorbent Assay ; Humans ; Leukotriene B4/metabolism ; Macrophages/cytology/drug effects ; Male ; Mice ; Monocytes/cytology/*drug effects ; Random Allocation ; Receptors, LDL/deficiency/genetics ; Thiazolidinediones/*therapeutic use ; Tumor Necrosis Factor-alpha/pharmacology