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Biomedical subjects

Douglas E Vaughan

Publications and source records attributed to Douglas E Vaughan.

At least 19 recordsLinked to original sources
PubMed2007

Epistatic effects of polymorphisms in genes from the renin-angiotensin, bradykinin, and fibrinolytic systems on plasma t-PA and PAI-1 levels.

Tissue plasminogen activator (t-PA) and plasminogen activator inhibitor 1 (PAI-1) directly influence thrombus formation and degradation and thereby risk for arterial thrombosis. Activation of the renin-angiotensin system has been linked to the production of PAI-1 expression via the angiotensin II type 1 receptor (AT1R). In addition, bradykinin can induce the release of t-PA through a B2 receptor mechanism. In the present study, we aimed to investigate the epistatic effects of polymorphisms in genes from the renin-angiotensin, bradykinin, and fibrinolytic systems on plasma t-PA and PAI-1 levels in a large population-based sample (n=2527). We demonstrated a strong significant interaction within genetic variations of the bradykinin B2 gene (P=0.002) and between ACE and bradykinin B2 (p=0.003) polymorphisms on t-PA levels in females. In males, polymorphisms in the bradykinin B2 and AT1R gene showed the most strong effect on t-PA levels (P=0.006). In both females and males, the bradykinin B2 gene interacted with AT1R gene on plasma PAI-1 levels (P=0.026 and P=0.039, respectively). In addition, the current study found a borderline significant interaction between PAI 4G5G and ACE I/D on plasma t-PA and PAI-1 levels. These results support the idea that the interplay between the renin-angiotensin, bradykinin, and fibrinolytic systems might play an important role in t-PA and PAI-1 biology.

Bradykinin↗
PubMed2006

Antiproliferative agents alter vascular plasminogen activator inhibitor-1 expression: a potential prothrombotic mechanism of drug-eluting stents.

OBJECTIVE: Drug eluting stents (DES) reduce the incidence of restenosis after coronary angioplasty. Enthusiasm has been tempered by a possible increased risk of in-stent thrombosis. We examined the effects of paclitaxel and rapamycin on the endothelial transcriptome to identify alterations in gene expression associated with thrombosis. METHODS AND RESULTS: Gene expression profiling was performed on human coronary artery endothelial cells treated with rapamycin or paclitaxel. Plasminogen activator inhibitor-1 (PAI-1) was the most consistently induced transcript in rapamycin-treated human coronary artery endothelial cells. RT-PCR and ELISA were performed to confirm positive findings. Transgenic mice engineered to express enhanced green fluorescent protein under control of the human PAI-1 promoter were also treated. Rapamycin and paclitaxel treated endothelial cells produced dose-dependent increases in PAI-1. There was a variable effect on endothelial tissue-type plasminogen activator (t-PA) expression. Enhanced expression of PAI-1 and enhanced green fluorescent protein were detected in coronary arteries, the aorta, and kidney of the mice. CONCLUSIONS: Antiproliferative agents stimulate the expression of prothrombotic genes. PAI-1 expression may also play a role in the prevention of restenosis through an antimigratory mechanism. The effects of antiproliferatives on vascular gene expression deserve further scrutiny in view of the increasing utilization of drug-eluting stents.

Animals↗
PubMed2006

Angiotensin II induces interleukin-6 in humans through a mineralocorticoid receptor-dependent mechanism.

This study tested the hypothesis that angiotensin promotes oxidative stress and inflammation in humans via aldosterone and the mineralocorticoid receptor. We measured the effect of intravenous aldosterone (0.7 mug/kg per hour for 10 hours followed by 0.9 mug/kg per hour for 4 hours) and vehicle in a randomized, double-blind crossover study in 11 sodium-restricted normotensive subjects. Aldosterone increased interleukin (IL)-6 (from 4.7+/-4.9 to 9.4+/-7.1 pg/mL; F=4.94; P=0.04) but did not affect blood pressure, serum potassium, or high-sensitivity C-reactive protein. We next conducted a randomized, double-blind, placebo-controlled, crossover study to measure the effect of 3-hour infusion of angiotensin II (2 ng/kg per minute) and norepinephrine (30 ng/kg per minute) on separate days after 2 weeks of placebo or spironolactone (50 mg per day) in 14 salt-replete normotensive subjects. Angiotensin II increased blood pressure (increase in systolic pressure: 13.7+/-7.5 and 15.2+/-9.4 mm Hg during placebo and spironolactone, respectively; P<0.001 for angiotensin II) and decreased renal plasma flow (-202+/-73 and -167+/-112 mL/min/1.73 kg/m(2); P<0.001 for angiotensin II effect) similarly during placebo and spironolactone. Spironolactone enhanced the aldosterone response to angiotensin II (increase of 17.0+/-10.6 versus 9.0+/-5.7 ng/dL; P=0.002). Angiotensin II transiently increased free plasma F(2)-isoprostanes similarly during placebo and spironolactone. Angiotensin II increased serum IL-6 concentrations during placebo (from 1.8+/-1.1 to 2.4+/-1.4 pg/mL; F=4.5; P=0.04) but spironolactone prevented this effect (F=6.4; P=0.03 for spironolactone effect). Norepinephrine increased blood pressure and F(2)-isoprostanes but not aldosterone or IL-6. Aldosterone increases IL-6 in humans. These data suggest that angiotensin II induces IL-6 through a mineralocorticoid receptor-dependent mechanism in humans. In contrast, angiotensin II-induced oxidative stress, as measured by F(2)-isoprostanes, is mineralocorticoid receptor independent and may be pressor dependent.

Adult↗
PubMed2006

Bone marrow plasminogen activator inhibitor-1 influences the development of obesity.

Plasma levels of plasminogen activator inhibitor-1 (PAI-1) are elevated in obesity and correlate with body mass index. The increase in PAI-1 associated with obesity likely contributes to increased cardiovascular risk and may predict the development of type 2 diabetes mellitus. Although adipocytes are capable of synthesizing PAI-1, the bulk of evidence indicates that cells residing in the stromal fraction of visceral fat are the primary source of PAI-1. We hypothesized that bone marrow-derived PAI-1, e.g. derived from macrophages located in visceral fat, contributes to the development of diet-induced obesity. To test this hypothesis, male C57BL/6 wild-type mice and C57BL/6 PAI-1 deficient mice were transplanted with either PAI-1(-/-), PAI-1(+/-), or PAI-1(+/+) bone marrow. The transplanted animals were subsequently fed a high fat diet for 24 weeks. Our findings show that only the complete absence of PAI-1 protects from the development of diet-induced obesity, whereas the absence of bone marrow-derived PAI-1 protects against expansion of the visceral fat mass. Remarkably, there is a link between the PAI-1 levels, the degree of inflammation in adipose tissue, and the development of obesity. Based on these findings we suggest that bone marrow-derived PAI-1 has an effect on the development of obesity through its effect on inflammation.

Adiponectin↗
PubMed2006

Calmodulin kinase II inhibition protects against myocardial cell apoptosis in vivo.

Inhibition of the multifunctional Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) or depletion of sarcoplasmic reticulum (SR) Ca(2+) stores protects against apoptosis from excessive isoproterenol (Iso) stimulation in cultured ventricular myocytes, suggesting that CaMKII inhibition could be a novel approach to reducing cell death in conditions of increased adrenergic tone, such as myocardial infarction (MI), in vivo. We used mice with genetic myocardial CaMKII inhibition due to transgenic expression of a highly specific CaMKII inhibitory peptide (AC3-I) to test whether CaMKII was important for apoptosis in vivo. A second line of mice expressed a scrambled, inactive form of AC3-I (AC3-C). AC3-C and wild-type (WT) littermates were used as controls. AC3-I mice have reduced SR Ca(2+) content and are resistant to Iso- and MI-induced apoptosis compared with AC3-C and WT mice. Phospholamban (PLN) is a target for modulation of SR Ca(2+) content by CaMKII. PLN(-/-) mice have increased susceptibility to Iso-induced apoptosis. Verapamil pretreatment prevented Iso-induced apoptosis in PLN(-/-) mice, indicating the involvement of a Ca(2+)-dependent pathway. AC3-I and AC3-C mice were bred into a PLN(-/-) background. Loss of PLN increased and equalized SR Ca(2+) content in AC3-I, AC3-C, and WT mice and abolished the resistance to apoptosis in AC3-I mice after MI. There was a trend (P = 0.07) for increased Iso-induced apoptosis in AC3-I mice lacking PLN compared with AC3-I mice with PLN. These findings indicate CaMKII is proapoptotic in vivo and suggest that regulation of SR Ca(2+) content by PLN contributes to the antiapoptotic mechanism of CaMKII inhibition.

Adrenergic beta-Agonists↗
PubMed2006

Bradykinin and its metabolite bradykinin 1-5 inhibit thrombin-induced platelet aggregation in humans.

Bradykinin 1-5 is a major stable metabolite of bradykinin, formed by the proteolytic action of angiotensin-converting enzyme. In vitro and animal studies suggest that bradykinin 1-5 possesses biological activity. This study tests the hypothesis that bradykinin 1-5 affects vasodilation, fibrinolysis, or platelet aggregation in humans. Graded doses of bradykinin (47-377 pmol/min) and bradykinin 1-5 (47-18,850 pmol/min) were infused in the brachial artery in random order in 36 healthy subjects. Forearm blood flow (FBF) was measured, and simultaneously obtained venous and arterial plasma samples were analyzed for tissue plasminogen activator (t-PA) antigen. In seven subjects each, alpha- and gamma-thrombin-induced platelet aggregation was measured in platelet-rich plasma obtained from antecubital venous blood at baseline and during peptide infusions. Bradykinin caused dose-dependent increases in FBF and net t-PA release (P < 0.001 for both). Bradykinin 1-5 did not affect FBF (P = 0.13) or net t-PA release (P = 0.46) at concentrations >1500 times physiologic. In contrast, both bradykinin and bradykinin 1-5 inhibited alpha-and gamma-thrombin-induced platelet aggregation (P < 0.01 versus baseline). Bradykinin 1-5 inhibited gamma-thrombin-induced platelet aggregation 50% at a calculated dose of 183 +/- 3 pmol/min. Neither bradykinin nor bradykinin 1-5 affected thrombin receptor-activating peptide-induced platelet aggregation, consistent with the hypothesis that bradykinin and bradykinin 1-5 inhibit thrombin-induced platelet aggregation by preventing cleavage of the thrombin receptor and liberation of thrombin receptor-activating peptide. This study is the first to demonstrate biological activity of bradykinin 1-5 following in vivo administration to humans. By inhibiting thrombin-induced platelet aggregation without causing vasodilation, bradykinin 1-5 may provide a model for small molecule substrate-selective thrombin inhibitors.

Adult↗
PubMed2006

Metabolic syndrome and the risk of cardiovascular disease in older adults.

OBJECTIVES: The purpose of this study was to assess whether metabolic syndrome (MetSyn) predicts a higher risk for cardiovascular events in older adults. BACKGROUND: The importance of MetSyn as a risk factor has not previously focused on older adults and deserves further study. METHODS: We studied the impact of MetSyn (38% prevalence) on outcomes in 3,035 participants in the Health, Aging, and Body Composition (Health ABC) study (51% women, 42% black, ages 70 to 79 years). RESULTS: During a 6-year follow-up, there were 434 deaths overall, 472 coronary events (CE), 213 myocardial infarctions (MI), and 231 heart failure (HF) hospital stays; 59% of the subjects had at least one hospital stay. Coronary events, MI, HF, and overall hospital stays occurred significantly more in subjects with MetSyn (19.9% vs. 12.9% for CE, 9.1% vs. 5.7% for MI, 10.0% vs. 6.1% for HF, and 63.1% vs. 56.1% for overall hospital stay; all p < 0.001). No significant differences in overall mortality was seen; however, there was a trend toward higher cardiovascular mortality (5.1% vs. 3.8%, p = 0.067) and coronary mortality (4.5% vs. 3.2%, p = 0.051) in patients with MetSyn. After adjusting for baseline characteristics, patients with MetSyn were at a significantly higher risk for CE (hazard ratio [HR] 1.56, 95% confidence interval [CI] 1.28 to 1.91), MI (HR 1.51, 95% CI 1.12 to 2.05), and HF hospital stay (HR 1.49, 95% CI 1.10 to 2.00). Women and whites with MetSyn had a higher coronary mortality rate. The CE rate was higher among subjects with diabetes and with MetSyn; those with both had the highest risk. CONCLUSIONS: Overall, subjects over 70 years are at high risk for cardiovascular events; MetSyn in this group is associated with a significantly greater risk.

Aged↗
PubMed2006

Foxc2 is a common mediator of insulin and transforming growth factor beta signaling to regulate plasminogen activator inhibitor type I gene expression.

Elevated plasma levels of plasminogen activator inhibitor type I (PAI-1), a significant risk factor of ischemic heart disease, are associated with insulin resistance in which insulin and transforming growth factor (TGF)-beta play a pivotal role in regulating PAI-1 production. Forkhead transcription factor FOXC2 is an important regulator of insulin resistance. However, the underlying molecular mechanisms to link FOXC2 to PAI-1 levels in insulin resistance remain to be elucidated. Here, we demonstrate that Foxc2 is a common transcriptional activator of insulin and TGF-beta signaling to directly regulate PAI-1 expression via 2 distinct target sites, an insulin response element (IRE) and a novel forkhead-binding element (FBE), adjacent to a Smad-binding site. We found that in adipocytes and endothelial cells Foxc2 mediates insulin action competing with another Forkhead protein, FOXO1, via the insulin response element, and simultaneously cooperate with the TGF-beta/Smad pathway to transactivate PAI-1. Importantly, Foxc2 haploinsufficiency in mice significantly attenuates TGF-beta1-induced PAI-1 expression in the cardiovascular system and adipose tissue. Taken together, we propose that Foxc2 is a key molecule to regulate PAI-1 gene expression.

Animals↗
PubMed2006

Endogenous NO regulates plasminogen activator inhibitor-1 during angiotensin-converting enzyme inhibition.

To test the hypothesis that NO contributes to effects of angiotensin-converting enzyme inhibitors on fibrinolysis, fibrinolytic balance was assessed in 17 normal subjects during placebo and after randomized, double-blind 4-week treatment with the NO precursor L-arginine (3 g TID), ramipril (10 mg QD), or L-arginine+ramipril. Neither L-arginine nor ramipril alone affected basal plasminogen activator inhibitor-1 or tissue-type plasminogen activator (t-PA) antigen in these salt-replete subjects in whom plasma renin activity was suppressed (mean+/-SD 0.7+/-0.5 ng angiotensin I/mL per hour). In contrast, L-arginine+ramipril reduced morning plasminogen activator inhibitor-1 antigen (10.8+/-9.5 ng/mL) and the molar ratio of plasminogen activator inhibitor-1:t-PA (2.3+/-1.6) compared with placebo (13.5+/-10.8 ng/mL, P=0.006; ratio 2.9+/-2.1, P=0.015) or ramipril alone (15.2+/-13.2 ng/mL, P=0.009; ratio 3.7+/-3.3, P=0.005). L-arginine and ramipril synergistically increased d-dimers (23.1+/-31.5, 29.7+/-50.0, 35.1+/-50.0, and 57.1+/-144.8 ng/mL during placebo, L-arginine, ramipril, and L-arginine+ramipril, respectively; P<0.05 for L-arginine+ramipril versus any other group). During ramipril, the NO synthase inhibitor L-NG-nitro-arginine-methyl-ester (2 mg/kg) significantly increased plasminogen activator inhibitor-antigen after 2 hours (from 9.4+/-8.6 ng/mL during vehicle to 13.5+/-11.0 ng/mL during L-NG-nitro-arginine-methyl-ester; P=0.020), consistent with an effect on expression but rapidly increased t-PA activity (from 0.4+/-0.3 to 0.5+/-0.4 IU/mL; P=0.031), consistent with an effect on release. Both effects of L-NG-nitro-arginine-methyl-ester were reversed by L-arginine. During angiotensin-converting enzyme inhibition, endogenous NO decreases plasminogen activator inhibitor-1 antigen and improves fibrinolytic balance in normotensive salt-replete subjects.

Adult↗
PubMed2006

The gender-specific role of polymorphisms from the fibrinolytic, renin-angiotensin, and bradykinin systems in determining plasma t-PA and PAI-1 levels.

Tissue plasminogen activator (t-PA) and plasminogen activator inhibitor 1 (PAI-1) directly influence thrombus formation and degradation and thus risk for arterial thrombosis. We report here results from a genetic analysis of plasma t-PA and PAI-1 levels in a large population-based sample from the PREVEND study in Groningen, the Netherlands (n = 2,527). We measured polymorphisms from genes of the fibrinolytic system, the renin-angiotensin system (RAS), and the bradykinin system. We found that males had higher levels of natural-log transformed t-PA, and PAI-1 (P < 0.01) compared to females. When stratifying females by menopausal status, PAI-1 levels were only significantly different between pre-menopausal females and males (p < 0.001). Furthermore, we found that age, body mass index, and waist-to-hip ratio were significant predictors of t-PA and PAI-1 in both females and males, and that the regression relationships between these factors and plasma t-PA and PAI-1 were dependent on gender. In addition, we found that the PAI-1 4G/5G polymorphism was a significant predictor of PAI-1 levels in both females and males, that the angiotensin II type I receptor A1166C was a significant predictor of t-PA and PAI-1 levels in females, and that the bradykinin receptor B2 58CT polymorphism was a significant predictor of t-PA levels in females. In conclusion, this large population-based study showed that t-PA and PAI-1 levels are determined by several demographic and genetic factors involved in the fibrinolytic, RAS and bradykinin system. In addition, the results support the idea that the biology of t-PA and PAI-1 is different between females and males.

Adult↗
PubMed2005

Differential regulation of endothelial exocytosis of P-selectin and von Willebrand factor by protease-activated receptors and cAMP.

Thrombin-mediated endothelial-cell release of von Willebrand factor (VWF) and P-selectin functionally links protease-activated receptors (PARs) to thrombosis and inflammation. VWF release can be stimulated by both Ca2+ and cAMP, and, although both VWF and P-selectin are found in Weibel-Palade bodies (WPBs), we found that their release could be differentially regulated. In these studies, human umbilical vein endothelial cells stimulated with cAMP or PAR2-AP led to a delayed release of VWF and significantly less P-selectin release compared with histamine, thrombin, or PAR1-AP. Dose-response studies revealed that PAR2-AP was significantly less efficacious in promoting the release of P-selectin compared with VWF. PAR2-AP-induced robust stimulation of intracellular Ca2+ coupled with a significantly greater inhibitory effect of calcium chelation on release of VWF compared with cell-surface expression of P-selectin, suggests an additional Ca2+-independent pathway involved in release of P-selectin. PAR2-AP failed to increase global cAMP levels; however, inhibition of protein kinase A led to a significant attenuation of PAR2-AP-mediated release of VWF. Confocal microscopy studies revealed that PAR2 and forskolin caused preferential release of a population of Weibel-Palade bodies (WPBs) consisting of only VWF. Thus, WPBs are pharmacologically and morphologically heterogeneous, and distinct granule populations are susceptible to differential regulation.

Calcium Signaling↗
PubMed2005

Angiotensin-converting enzyme inhibition increases basal vascular tissue plasminogen activator release in women but not in men.

OBJECTIVE: Angiotensin-converting enzyme inhibition (ACEI) increases vascular tissue plasminogen activator (t-PA) release through endogenous bradykinin (BK). We tested the hypothesis that gender influences the effect of ACEI on t-PA release. METHODS AND RESULTS: We measured the effect of intra-arterial enalaprilat (0.33 microg/min per 100 mL forearm volume) on forearm blood flow (FBF) and net t-PA release before and during BK (25 to 400 ng/min) and methacholine (3.2 to 12.8 microg/min) in premenopausal women, postmenopausal women not using hormone replacement, young men, and older men. Baseline net t-PA release was similar among groups. Enalaprilat increased basal t-PA release in premenopausal (from 0.9+/-1.0 to 5.1+/-1.7 ng/min per 100 mL, P=0.023) and postmenopausal women (from -3.9+/-2.2 to 3.9+/-1.1 ng/min per 100 mL, P=0.010) but not in young or older men (P=0.028 men versus women). Enalaprilat potentiated the effect of exogenous BK on FBF similarly in all groups. However, during enalaprilat, BK-stimulated t-PA release was greatest in premenopausal women (339.9+/-86.4 ng/min per 100 mL at the 100 ng/min dose, P<0.05 versus any other group), intermediate in postmenopausal women (243.8+/-51.1 ng/min per 100 mL, P<0.05 versus either male group), and least in young (111.9+/-19.2 ng/min/100 mL) and older men (103.4+/-27.6 ng/min/100 mL). CONCLUSIONS: ACEI enhances basal t-PA release in women, independent of menopausal status, but not in men. During ACEI, both gender and menopausal status affect BK stimulated t-PA release.

Adult↗
PubMed2005

Pivotal role of PAI-1 in a murine model of hepatic vein thrombosis.

Hepatic veno-occlusive disease (VOD) is a common complication of high-dose chemotherapy associated with bone marrow transplantation. While the pathogenesis of VOD is uncertain, plasminogen activator inhibitor-1 (PAI-1) has emerged as a diagnostic marker and predictor of VOD in humans. In this study, we investigated the role of PAI-1 in a murine model of VOD produced by long-term nitric oxide synthase inhibition using L-NAME. After 6 weeks, wild-type (WT) mice developed extensive fibrinoid hepatic venous thrombi and biochemical evidence of hepatic injury and dysfunction. In contrast, PAI-1-deficient mice were largely protected from the development of hepatic vein thrombosis. Furthermore, WT mice that received tiplaxtinin, an antagonist of PAI-1, were effectively protected from L-NAME-induced thrombosis. Taken together, these data indicate that NO and PAI-1 play pivotal and antagonistic roles in hepatic vein thrombosis and that PAI-1 is a potential target in the prevention and treatment of VOD in humans.

Animals↗
PubMed2005

Differing effects of mineralocorticoid receptor-dependent and -independent potassium-sparing diuretics on fibrinolytic balance.

This study tests the hypothesis that spironolactone influences plasminogen activator inhibitor-1 (PAI-1) concentrations through mineralocorticoid receptor antagonism rather than through changes in potassium. Effects of spironolactone (50 mg per day) and triamterene (50 mg per day) on fibrinolytic balance were compared in 18 normotensive and 20 hypertensive subjects pretreated with hydrochlorothiazide (HCTZ; 12.5 mg per day). Blood pressure and serum potassium were similar in spironolactone and triamterene treatment groups. The effect of the 2 drugs on the renin-angiotensin-aldosterone system was also similar. In contrast, spironolactone and triamterene exerted opposing effects on PAI-1 antigen (P=0.006 for drug effect). In normotensive subjects, triamterene (from 10.1+/-7.8 to 16.9+/-9.9 ng/mL at 9 am, P=0.019; from 7.6+/-5.4 to 11.5+/-7.3 ng/mL at 11 am, P=0.027; from 9.3+/-7.7 to 13.7+/-8.5 ng/mL for average of all time points, P=0.054) but not spironolactone significantly increased PAI-1 antigen. In hypertensive subjects, spironolactone significantly decreased PAI-1 antigen (from 22.0+/-23.4 to 16.7+/-19.0 ng/mL at 10 am, P=0.041; from 17.5+/-21.7 to 12.7+/-16.8 ng/mL at 11 am, P=0.043; from 20.3+/-22.6 to 16.6+/-19.7 ng/mL for average of all time points, P=0.014), whereas there was no effect of triamterene. Only spironolactone significantly decreased the molar ratio of PAI-1 to tissue-type plasminogen activator (t-PA) in hypertensive subjects. By regression analysis, predictors of mean PAI-1 response were spironolactone versus triamterene (P=0.014), hypertension (P=0.002), and PAI-1 response to HCTZ (P=0.019), with a trend for aldosterone (P=0.061). Mineralocorticoid receptor antagonism prevents the effect of activation of the renin-angiotensin-aldosterone system on PAI-1 antigen in normotensive subjects and improves fibrinolytic balance in hypertensive subjects through a potassium-independent mechanism.

Adult↗
PubMed2005

Thrombin modulates the expression of a set of genes including thrombospondin-1 in human microvascular endothelial cells.

Thrombospondin-1 (THBS1) is a large extracellular matrix glycoprotein that affects vasculature systems such as platelet activation, angiogenesis, and wound healing. Increases in THBS1 expression have been liked to disease states including tumor progression, atherosclerosis, and arthritis. The present study focuses on the effects of thrombin activation of the G-protein-coupled, protease-activated receptor-1 (PAR-1) on THBS1 gene expression in the microvascular endothelium. Thrombin-induced changes in gene expression were characterized by microarray analysis of approximately 11,000 different human genes in human microvascular endothelial cells (HMEC-1). Thrombin induced the expression of a set of at least 65 genes including THBS1. Changes in THBS1 mRNA correlated with an increase in the extracellular THBS1 protein concentration. The PAR-1-specific agonist peptide (TFLLRNK-PDK) mimicked thrombin stimulation of THBS1 expression, suggesting that thrombin signaling is through PAR-1. Further studies showed THBS1 expression was sensitive to pertussis toxin and protein kinase C inhibition indicating G(i/o)- and G(q)-mediated pathways. THBS1 up-regulation was also confirmed in human umbilical vein endothelial cells stimulated with thrombin. Analysis of the promoter region of THBS1 and other genes of similar expression profile identified from the microarray predicted an EBOX/EGRF transcription model. Expression of members of each family, MYC and EGR1, respectively, correlated with THBS1 expression. These results suggest thrombin formed at sites of vascular injury increases THBS1 expression into the extracellular matrix via activation of a PAR-1, G(i/o), G(q), EBOX/EGRF-signaling cascade, elucidating regulatory points that may play a role in increased THBS1 expression in disease states.

Adenosine Diphosphate↗
PubMed2005

Kruppel-like factor 2 (KLF2) regulates endothelial thrombotic function.

The vascular endothelium maintains blood fluidity by inhibiting blood coagulation, inhibiting platelet aggregation, and promoting fibrinolysis. Endothelial cells lose these nonthrombogenic properties on exposure to proinflammatory stimuli. We recently identified the Kruppel-like factor KLF2 as a novel regulator of endothelial proinflammatory activation. Here it is found that KLF2 differentially regulates key factors involved in maintaining an antithrombotic endothelial surface. Overexpression of KLF2 strongly induced thrombomodulin (TM) and endothelial nitric oxide synthase (eNOS) expression and reduced plasminogen activator inhibitor-1 (PAI-1) expression. Furthermore, overexpression of KLF2 inhibited the cytokine-mediated induction of tissue factor (TF). In contrast, siRNA mediated knockdown of KLF2 reduced antithrombotic gene expression while inducing the expression of pro-coagulant factors. The functional importance of KLF2 was verified by in vitro clotting assays. By comparison to control infected cells, KLF2 overexpression increased blood clotting time as well as flow rates under basal and inflammatory conditions. In contrast, siRNA-mediated knockdown of KLF2 reduced blood clotting time and flow rates. These observations identify KLF2 as a novel transcriptional regulator of endothelial thrombotic function. The full text of this article is available online at http://circres.ahajournals.org.

Animals↗
PubMed2005

The sterol response element binding protein regulates cyclooxygenase-2 gene expression in endothelial cells.

We previously demonstrated that cholesterol deprivation increases endothelial cyclooxygenase-2 (COX-2)-dependent prostacyclin [prostaglandin I2 (PGI2)] production in vitro. Cholesterol directly regulates gene transcription through the sterol response element binding protein (SREBP). In this work, we demonstrate that SREBP directly regulates COX-2 expression. Cholesterol reduces human COX-2 promoter-luciferase reporter construct activity in transiently transfected endothelial cells. Conversely, cotransfection with a constitutively active mutant SREBP increases COX-2 promoter activity. SREBP-1a and -2 specifically bind a putative sterol response element (SRE) sequence in the COX-2 promoter. This sequence competes for SREBP binding to a low density lipoprotein receptor consensus sequence in an electromobility-shift assay. These data indicate that endothelial COX-2 is regulated by cholesterol via the SREBP pathway. The present study identifies COX-2 as the first vascular gene without a clear role in lipid metabolism transactivated by SREBP, and suggests that enhanced production of PGI2 through this pathway may be an additional benefit of cholesterol-lowering therapies.

Animals↗