The -460T/+405C haplotype in the VEGF gene, which is associated with lower promoter activity, was significantly less common in women with endometriosis than in controls. These data suggest that the +405G allele may influence the likelihood of a woman developing the disease.
The redox state plays an important role in gene regulation. Thiols maintain the intracellular redox homeostasis. To understand the role of thiols in redox signaling, we have studied the effect of thiol alkylation on platelet-derived growth factor-BB (PDGF-BB)-induced cell survival events in vascular smooth muscle cells. PDGF-BB stimulated Akt phosphorylation predominantly at Ser-473. N-Ethylmaleimide (NEM), a thiol alkylating agent, blocked PDGF-BB-induced Akt phosphorylation without affecting its upstream phosphatidylinositol 3-kinase (PI3K). On the other hand, LY294002 and wortmannin, specific inhibitors of PI3K, prevented PDGF-BB-induced phosphorylation of Akt and its downstream effector molecules, p70S6K, ribosomal protein S6, 4E-BP1, and eIF4E. NEM also abrogated the phosphorylation of p70S6K, ribosomal protein S6, 4E-BP1, and eIF4E induced by PDGF-BB, suggesting that thiol alkylation interferes with the PI3K/Akt pathway at the level of Akt. In addition, NEM blocked PDGF-BBinduced phosphorylation of BAD and forkhead transcription factor FKHR-L1, and these events correlated with increased apoptosis. NEM alone and in concert with PDGF-BB increased reactive oxygen species (ROS) production and protein phosphatase 2A (PP2A) activity in VSMC. The inhibition of PDGF-BB-induced Akt phosphorylation by NEM was completely reversed by PP2A inhibitors fostriecin and okadaic acid, ceramide synthase inhibitor fumonisin B1, and ROS scavenger Nacetylcysteine (NAC). NAC also attenuated the apoptosis induced by NEM, alone or in combination with PDGF-BB. Together, these findings demonstrate for the first time that PP2A mediates thiol alkylation-dependent redox regulation of Akt and cell survival.The cellular redox state plays an important role in the regulation of gene expression in prokaryotes and eukaryotes (1-4). The following observations support this notion: 1) Oxidants regulate the activities of several transcription factors, including activator protein-1, nuclear factor kappa B, and p53 (5-7); 2) Oxidants are capable of activating several early response events, including stimulation of protein tyrosine phosphorylation, activation of mitogen-activated protein kinases and induction of expression of proto-oncogenes (8 -11); 3) Oxidants are produced acutely in response to various agents, including growth factors and cytokines in several cell types (12, 13), and a requirement for their production in the mitogenic effects of receptor tyrosine kinase and G protein-coupled receptor agonists has been demonstrated (14, 15); and 4) In addition to producing oxidants, cells also possess enzymatic and non-enzymatic mechanisms for their removal (16 -18), and this feature attests to the role of oxidants as second messenger molecules (19). Despite the growing body of information on the role of oxidants in the regulation of gene expression, the mechanisms by which these molecules transmit the extracellular signals from the plasma membrane to the nucleus are less clear. Thiols play a critical role in the reduction/oxidation reactions as w...
Objective-Migration of vascular smooth muscle cells (VSMCs) from media to intima is a key event in the pathophysiology of atherosclerosis and restenosis. The lipoxygenase products of polyunsaturated fatty acids (PUFA) were shown to play a role in these diseases. cAMP response element binding protein (CREB) has been implicated in the regulation of VSMC growth and motility in response to thrombin and angiotensin II. The aim of the present study was to test the role of CREB in an oxidized lipid molecule, 15(S)-HETE-induced VSMC migration and neointima formation. Methods and Results-15(S)-HETE stimulated VSMC migration in CREB-dependent manner, as measured by the modified Boyden chamber method. Blockade of MEK1, JNK1, or p38MAPK inhibited 15(S)-HETE-induced CREB phosphorylation and VSMC migration. 15(S)-HETE induced expression and secretion of interleukin-6 (IL-6), as analyzed by RT-PCR and ELISA, respectively. Neutralizing anti-IL-6 antibodies blocked 15(S)-HETE-induced VSMC migration. Dominant-negative mutant-mediated blockade of ERK1/2, JNK1, p38MAPK, or CREB suppressed 15(S)-HETE-induced IL-6 expression in VSMCs. Serial 5Ј deletions and site-directed mutagenesis of IL-6 promoter along with chromatin immunoprecipitation using anti-CREB antibodies showed that cAMP response element is essential for 15(S)-HETE-induced IL-6 expression. Dominant-negative CREB also suppressed balloon injury-induced IL-6 expression, SMC migration from media to intimal region, and neointima formation. Adenovirus-mediated transduction of 15-lipoxygenase 2 (15-LOX2) caused increased production of 15-HETE in VSMCs and enhanced IL-6 expression, SMC migration from media to intimal region, and neointima formation in response to arterial injury. Key Words: cAMP response element binding protein Ⅲ hydroxyeicosatetraenoic acid Ⅲ interleukin-6 Ⅲ vascular smooth muscle cell migration V SMC migration from media to intima plays a determinant role in atherosclerosis and restenosis. [1][2][3] Arachidonic acid (AA) and its oxygenative metabolites, known as eicosanoids, are involved in the maintenance of vascular tone. 4,5 Lipoxygenases (LOXs) are nonheme iron dioxygenases that stereospecifically introduce molecular oxygen into polyunsaturated fatty acids (PUFA) such as AA, resulting in the formation of hydroperoxyeicosatetraenoic acids (HPETEs) which are further converted to hydroxyeicosatetraenoic acids (HETEs). Two LOXs in particular, 15-LOX1 in humans and its closely related ortholog, 12/15-LOX, in mice, as well as 5-LOX that convert AA to HETEs are the prime candidates implicated in atherosclerosis and restenosis. 6 -8 It is known that oxidation of low-density lipoprotein (LDL) is a contributing factor in the pathogenesis of atherosclerosis. 9 -11 Many studies have shown that 15-LOX1 and 12/15-LOX are involved in the oxidation of LDL, and thereby in the pathogenesis of atherosclerosis. 10,11 It was also demonstrated that atherosclerotic arteries express increased levels of 15-LOX1 and its AA product, 15-HETE in rabbits. 12,13 In addition, recently ...
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