Plasminogen Activator Inhibitor-1 Protein Research Articles

Pathogenetic and predictive value of biomarkers in patients with ALI and lower severity of illness: results from two clinical trials

Plasma and bronchoalveolar lavage (BAL) biomarkers related to the pathogenesis of acute lung injury (ALI) have previously been associated with poorer clinical outcomes and increased disease severity among patients with ALI. Whether these biomarkers have predictive value in a less severely ill population that excludes septic patients with high APACHE II scores is currently unknown. We tested the association of plasma and BAL biomarkers with physiological markers of ALI severity or clinically relevant outcomes in a secondary analysis of a clinical trial of activated protein C for the treatment of ALI. Plasma plasminogen activator inhibitor-1 (PAI-1) and mini-BAL protein were both significantly associated with increased oxygenation index (P = 0.02 and 0.01, respectively), whereas there was a trend toward an association between IL-6 and oxygenation index (P = 0.057). High plasma IL-6, thrombomodulin, and mini-BAL protein were all significantly associated with fewer ventilator-free days (VFDs) (P = 0.01, 0.01, and 0.05, respectively); no markers were associated with mortality, but we hypothesized that this was due to the small size of our cohort and the low death rate. To confirm these associations in a larger sample, we identified a restricted cohort of patients from the ARDS Network ALVEOLI study with similar baseline characteristics. We retested the associations of the significant biomarkers with markers of severity and clinical outcomes and studied IL-8 as an additional biomarker given its important predictive value in prior studies. In this restricted cohort, IL-6 was significantly associated with oxygenation index (P = 0.02). Both IL-6 and IL-8 were associated with decreased VFDs and increased 28-day mortality. Future studies should be focused on examining larger numbers of patients with less severe ALI to further test the relative predictive value of plasma and mini-BAL biomarkers for clinically relevant outcomes, including VFDs and mortality, and for their prospective utility in risk stratification for future clinical trials.

Challenging delivery of VLHL NS plasminogen activator inhibitor-1 by osmotic pumps in diabetic mouse: A case report

ALZET® osmotic pumps are implantable devices used in animals for the continuous infusion of drugs or proteins at controlled rates from 1 day to 4 weeks. Pumps have been used successfully in a number of studies on the effects of controlled delivery of a wide range of experimental agents, independent of their properties. In the present study, use of these pumps was made in mice with diabetic nephropathy. Plasminogen activator inhibitor-1 (PAI-1) mediates diabetic nephropathy, which is characterized by the excessive accumulation of extracellular matrix (ECM) in the kidney. Disproportionate PAI-1 inactivates tissue plasminogen activator, which is one of the proteolytic enzymes in a cascade responsible for ECM remodeling in the kidney. The decrease of PAI-1 in the kidney has been shown to arrest the progression of nephropathy in experimental animals. This was achieved using inactive PAI-1R which increased the clearance of wild-type PAI-1 in order to protect net proteolytic activity and ECM clearance. However, this protein has a brief half-life in vivo, therefore, high and frequent doses are required. Thus, VLHL NS PAI-1 protein with a long half-life of over 700 h (Gln197Cys, Gly355Cys) inactivated by single point mutation (Arg369Ala) was used. Following the sacrifice of animals the tips of the flow moderators of the osmotic pumps in the treated animals were found to be clogged. In addition, from each pump from the treatment group, but not controls, we collected 50–150 μl of clear liquid containing VLHL NS PAI-1, cellular and serum proteins suggesting early pump sealing by cellular material. In conclusion, despite encouraging results obtained for the PAI-1R protein, the method of VLHL PAI-1 delivery should be ameliorated.

Fasudil, a Rho-Kinase Inhibitor, Attenuates Bleomycin-Induced Pulmonary Fibrosis in Mice

The mechanisms underlying the pathogenesis of idiopathic pulmonary fibrosis (IPF) involve multiple pathways, such as inflammation, epithelial mesenchymal transition, coagulation, oxidative stress, and developmental processes. The small GTPase, RhoA, and its target protein, Rho-kinase (ROCK), may interact with other signaling pathways known to contribute to pulmonary fibrosis. This study aimed to determine the beneficial effects and mechanisms of fasudil, a selective ROCK inhibitor, on bleomycin-induced pulmonary fibrosis in mice. Our results showed that the Aschcroft score and hydroxyproline content of the bleomycin-treated mouse lung decreased in response to fasudil treatment. The number of infiltrated inflammatory cells in the bronchoalveolar lavage fluid (BALF) was attenuated by fasudil. In addition, fasudil reduced the production of transforming growth factor-β1 (TGF-β1), connective tissue growth factor (CTGF), alpha-smooth muscle actin (α-SMA), and plasminogen activator inhibitor-1 (PAI-1) mRNA and protein expression in bleomycin-induced pulmonary fibrosis. These findings suggest that fasudil may be a potential therapeutic candidate for the treatment of pulmonary fibrosis.

Ethyl pyruvate reduces ventilation-induced neutrophil infiltration and oxidative stress

Mechanical ventilation with high tidal volume causes intense inflammatory responses and oxidative stress, including the release of high-mobility group box-1 (HMGB1), plasminogen activator inhibitor-1 (PAI-1) and heme oxygenase-1 (HO-1). The mechanisms regulating ventilator-induced lung injury (VILI) are unclear. We hypothesized that ethyl pyruvate attenuated acute lung injury as adjunctive pharmacological strategy by down-regulating neutrophil infiltration, oxidative stress and HMGB1 mRNA expression. C57BL/6 mice, weighing 20-25 g, were exposed to either low-tidal-volume (6 mL/kg) or high-tidal-volume (30 mL/kg) mechanical ventilation with room air for 2-5 h and subjected to 100 mg/kg ethyl pyruvate administration intraperitoneally. Non-ventilated mice served as the control group. Evans blue dye, lung wet-to-dry weight ratio, lung neutrophil infiltration and myeloperoxidase, free radicals, gene expression of HMGB1, active PAI-1 and HMGB1 production and HO-1 expression were measured. The expression of HO-1 was studied by immunohistochemistry. High-tidal-volume ventilation induced microvascular leak, neutrophil recruitment, oxidative injury, HMGB1 and active PAI-1 protein production, and upregulation of HMGB1 mRNA and stress-inducible protein HO-1. In contrast, administration of ethyl pyruvate before high stretch mechanical ventilation prevented lung edema formation, inflammatory cytokine production, neutrophil accumulation, oxidative stress and HMGB1 and HO-1 expression. High-tidal-volume mechanical ventilation increased microvascular permeability, neutrophil influx and inflammatory cytokines. Ethyl pyruvate is capable of suppressing the VILI related to the reduction of HMGB1 and our findings support the potential use of ethyl pyruvate as a therapeutic agent for the prevention of VILI.

Alterations in the Common Pathway of Coagulation During Weight Loss Induced by Gastric Bypass in Severely Obese Patients

The objective of this study was to establish the relationship between the plasminogen activator inhibitor-1 (PAI-1), antithrombin-III (ATIII), fibrinogen, and white blood cell (WBC) levels in severely obese patients. We analyzed various plasma parameters implicated in the intrinsic and extrinsic coagulation pathway from 34 severely obese patients before and 1, 6, and 12 months after gastric bypass. In obese people, ATIII, fibrinogen, and WBC levels were in the upper limit of the normal range, and all were higher and significantly different from nonobese people. After bariatric surgery, the ATIII level continued to be high during the first month and increased until 12 months, while fibrinogen decreased only at that time. PAI-1 plasma protein and PAI-1 mRNA levels in liver and adipose tissue show similar profiles and had a strong positive correlation (r = 0.576, P = 0.0003 in liver; r = 0.433, P = 0.0004 in adipose tissue). They were higher in obese patients compared with nonobese control, but tended to recover normal values 1 month after surgery. Thus, the liver and adipose tissue could be an important source of PAI-1 protein in plasma. Gastric bypass surgery leads to a normalization of the hematological profile and a decrease in PAI-1 levels, which entails a decrease of risk for thromboembolism in severely obese.

Induction of insulin resistance by the adipokines resistin, leptin, plasminogen activator inhibitor-1 and retinol binding protein 4 in human megakaryocytes

In normal platelets, insulin inhibits agonist-induced Ca(2+) mobilization by raising cyclic AMP. Platelet from patients with type 2 diabetes are resistant to insulin and show increased Ca(2+) mobilization, aggregation and procoagulant activity. We searched for the cause of this insulin resistance. Platelets, the megakaryocytic cell line CHRF-288-11 and primary megakaryocytes were incubated with adipokines and with plasma from individuals with a disturbed adipokine profile. Thrombin-induced Ca(2+) mobilization and signaling through the insulin receptor and insulin receptor substrate 1 were measured. Abnormalities induced by adipokines were compared with abnormalities found in platelets from patients with type 2 diabetes. Resistin, leptin, plasminogen activator inhibitor-1 and retinol binding protein 4 left platelets unchanged but induced insulin resistance in CHRF-288-11 cells. Interleukin-6, tumor necrosis factor-α and visfatin had no effect. These results were confirmed in primary megakaryocytes. Contact with adipokines for 2 hours disturbed insulin receptor substrate 1 Ser(307)-phosphorylation, while contact for 72 hours caused insulin receptor substrate 1 degradation. Plasma with a disturbed adipokine profile also made CHRF-288-11 cells insulin-resistant. Platelets from patients with type 2 diabetes showed decreased insulin receptor substrate 1 expression. Adipokines resistin, leptin, plasminogen activator-1 and retinol binding protein 4 disturb insulin receptor substrate 1 activity and expression in megakaryocytes. This might be a cause of the insulin resistance observed in platelets from patients with type 2 diabetes.

Histone Deacetylase Inhibitor m-Carboxycinnamic Acid bis-Hydroxamide Attenuates Plasminogen Activator Inhibitor–1 Expression in Human Pleural Mesothelial Cells

Plasminogen activator inhibitor-1 (PAI-1), primarily up-regulated by transforming growth factor (TGF)-β, is essential in the development of fibrosis. Histone deacetylase (HDAC) was shown to modulate gene expression and fibrogenesis in various tissues. However, the implications of HDAC in terms of PAI-1 expression and pleural fibrosis remain unclear. In this study, we examined the effects of m-carboxycinnamic acid bis-hydroxamide (CBHA), a hybrid-polar HDAC inhibitor, on the TGF-β1-induced expression of PAI-1 in a human pleural mesothelial cell line (MeT-5A). MeT-5A cells were treated with TGF-β1 in the presence or absence of CBHA. We assayed the expression and stability of PAI-1 mRNA and protein, PAI-1 promoter activity, the activation of Smad signaling, the protein-protein interactions of Smads with transcriptional cofactors Sp1 and coactivator p300, and the expression of the mRNA-stabilizing protein nucleolin. The results indicate that CBHA significantly inhibited TGF-β1-induced PAI-1 mRNA and protein expression, and attenuated PAI-1 promoter activity in MeT-5A cells. CBHA abrogated TGF-β1-induced Smad4 nuclear translocation, but not Smad2/3 activation. Furthermore, the association of Smad4 with p300, but not with Sp1, was disrupted by CBHA. Alternatively, CBHA suppressed TGF-β1-induced nucleolin expression, and thereby destabilized PAI-1 mRNA and decreased PAI-1 protein concentrations. These findings suggest that the inhibition of HDAC activity by CBHA may attenuate PAI-1 expression through the modulation of cellular signaling at multiple levels. Given the down-regulating effect of CBHA on PAI-1 expression, HDAC inhibitors should be tested further in animal models as potential therapeutic agents for pleural fibrosis.

Mature breast adipocytes promote breast cancer cell motility

Adipocytes express substances involved in both normal physiology and pathological processes. One such adipocyte protein is the Serpin (serine protease inhibitor) plasminogen activator inhibitor-1 (PAI-1). PAI-1 functions to inhibit urokinase type plasminogen activator (uPA) though PAI-1 itself is also implicated in breast cancer progression. While the role of adipocytes in breast cancer development is not fully understood, obesity is a known risk factor associated with breast cancer. Thus, we characterized adipocytes from breast and omental tissues for PAI-1 and uPA, and the influence of adipocytes on breast cancer cell motility. Using preadipocyte cells from breast and omental adipose tissue, we differentiated each site into mature adipocytes. PAI-1 protein was found in breast adipocytes>omental preadipocytes>omental adipocytes>breast preadipocytes. Interestingly, uPA protein was not detected in any of these cell types. We then incubated breast adipocyte conditioned media (Adip-CM) and preadipocyte conditioned media (PreAdip-CM) on both normal (MCF-10A) and malignant (MCF-10CA1) breast epithelial cell lines. Adip-CM, but not PreAdip-CM, (a) increased cell motility in both MCF-10A and MCF-10CA1 cells; (b) increased cell-associated uPA activity in both cell lines; (c) increased phosphorylated-Akt levels in MCF-10CA1 cells; and (d) gene array profiles show altered expression of several genes associated with cancer adhesion, metastasis and signaling. Our results suggest that mature breast adipocytes are capable of altering the epithelial cell phenotype, producing a more motile cell type and further provide a potential link between obesity and risk of breast cancer.

A Novel mRNA Binding Protein Complex Promotes Localized Plasminogen Activator Inhibitor-1 Accumulation at the Myoendothelial Junction

Plasminogen activator inhibitor-1 (PAI-1) has previously been shown to be key to the formation of myoendothelial junctions (MEJs) in normal and pathological states (eg, obesity). We therefore sought to identify the mechanism whereby PAI-1 could be selectively accumulated at the MEJ. We identified PAI-1 protein enrichment at the MEJ in obese mice and in response to tumor necrosis factor (TNF-α) with a vascular cell coculture. However, PAI-1 mRNA was also found at the MEJ and transfection with a PAI-1-GFP with TNF-α did not demonstrate trafficking of the protein to the MEJ. We therefore hypothesized the PAI-1 mRNA was being locally translated and identified serpine binding protein-1, which stabilizes PAI-1 mRNA, as being enriched in obese mice and after treatment with TNF-α, whereas Staufen, which degrades PAI-1 mRNA, was absent in obese mice and after TNF-α application. We identified nicotinamide phosphoribosyl transferase as a serpine binding protein-1 binding partner with a functional τ-like microtubule binding domain. Application of peptides against the microtubule binding domain significantly decreased the number of MEJs and the amount of PAI-1 at the MEJ. We conclude that PAI-1 can be locally translated at the MEJ as a result of a unique mRNA binding protein complex.

Antineoplastic agent busulfan regulates a network of genes related to coagulation and fibrinolysis

Purpose Hepatic veno-occlusive disease (HVOD) is one of the major complications following hematopoietic stem cell transplantation (HSCT). Although high-dose busulfan is associated with the development of HVOD, the underlying molecular mechanisms are still unknown.Methods Transcriptional gene regulation by busulfan was profiled using Affymetrix GeneChip® Human Genome U133 Plus 2.0 arrays. Messenger RNA (mRNA) expression of regulated genes was assessed by TaqMan real-time polymerase chain reaction (PCR), and protein expression and secretion was determined by enzyme-linked immunosorbent assay (ELISA)in cell supernatants, lysates, and patient plasma.Results Plasma levels of plasminogen activator inhibitor(PAI)-1 significantly increased 48 h after starting busulfan treatment IV in children preconditioned for HSCT. In vitro,busulfan significantly induced plasminogen activator inhibitor-1 (PAI-1) expression in endothelium-like ECV304 cells in a concentration- and time-dependent manner. Comparative transcriptional profiling of busulfan-treated and control ECV304 cells identified differential expression of genes related to coagulation and fibrinolysis, including tissue factor, tissue factor pathway inhibitor-1, protein S, thrombospondin-1, urokinase receptor, and PAI-1, as well as activin A and transforming growth factor beta 1 (TGF-β1). Ingenuity pathway analysis (IPA) suggested TGF-β1 as a central modulator of gene regulation by busulfan. Consequently, expression of tissue factor, urokinase receptor, and PAI-1 mRNA and PAI-1 protein secretion induced by busulfan were significantly reduced by the activin A/TGF-β1 inhibitor SB 431542 in ECV304 and primary endothelial cells.Conclusions This is the first report that directly relates busulfan exposure to antifibrinolytic activity by PAI-1 and hypercoagulation possibly mediated by members of the TGF-β1 family. This suggests further research to evaluate activin A and TGF-β1 as potential targets for HVOD treatment.

Relationship between changes in selected thrombotic and inflammatory factors, echocardiographic parameters and the incidence of venous thrombosis after pacemaker implantation based on our own observations

IntroductionThrombosis (VTh) is a rare dangerous complication of pacemaker implantation (PM). The aim of the study was to determine the dynamics of change in selected thrombotic and inflammatory factors after PM.Material and methodsThe study involved 81 patients (30 female, mean age: 71.1 years) with PM, divided into two groups. Group A (71 patients) consisted of patients without VTh, whereas group B (10 patients) comprised the patients with VTh. A transthoracic echocardiogram (TTE) and a venous ultrasound (VU) examination were performed. The levels of D-dimers, fibrinogen, tissue factor (TF), factor VII, plasminogen activator inhibitor-1 (PAI-1), interleukin-6 (IL-6) and high-sensitivity C-reactive protein (hsCRP) were determined in the venous blood. After PM, the TTE and VU examinations were repeated at 6 and 12 months, and blood analyses were performed within 7 days after PM, and subsequently at 6 and 12 months.ResultsIn 10 patients of group B, symptomatic VTh occurred at a mean time of 13.06 months after PM. Initially, the levels of IL-6, hsCRP, D-dimers, fibrinogen, TF, VII factor and PAI-1 were considerably higher in group B than in group A. In all patients the levels of these factors kept on increasing for up to 7 days after the procedure. In group A they subsequently decreased, whereas in group B they continued to rise.ConclusionsIncreased levels of inflammatory and thrombotic factors were observed in patients with VTh before and after PM. The factors of highest risk of VTh occurrence were D-dimers, fibrinogen and TF.

Effects of Valsartan on Fibrinolysis in Hypertensive Patients With Metabolic Syndrome

The purpose of this study was to analyze the effect of valsartan on abnormal adipocyte metabolism and prothrombotic state in hypertensive patients with metabolic syndrome (MetS). We conducted a multicenter, prospective, randomized, parallel-group controlled trial in 150 hypertensive patients with MetS. They were randomly assigned to receive either 80-160 mg valsartan per day (valsartan group, n=79) or other conventional treatment without a renin-angiotensin system (RAS) inhibitor (non-RAS inhibitor group, n=71). After 1 year, there were no significant differences between the 2 groups in the changes in systolic and diastolic blood pressures (valsartan: 153±15/86±15 to 138±16/77±12 mmHg; non-RAS inhibitor: 150±14/82±15 to 137±15/76±10 mmHg). There was a significant difference in the change in the levels of plasminogen activator inhibitor-1 (PAI-1) between the 2 groups after 1 year (valsartan: 3.7±3.2 ng/ml; non-RAS inhibitor: 5.8±3.3 ng/ml, P=0.04). There was no significant difference between groups in the change in the concentration of adiponectin after 1 year (valsartan: 0.3±0.4 µg/ml; non-RAS inhibitor: 0.9±0.4 µg/ml, P=0.22). The animal study showed aortic PAI-1 protein expression was reduced in double knockout mice of angiotensin II type 1a receptor and apolipoprotein E (apoE) compared with the apoE knockout mice. Valsartan reduced plasma PAI-1 levels compared to non-RAS inhibitor in hypertensive patients with MetS, which suggests it may be useful for improving fibrinolytic function.

P38 Mitogen-Activated Protein Kinase Is Required for Glucosamine-Induced Endothelial Nitric Oxide Synthase Uncoupling and Plasminogen-Activator Inhibitor Expression

Hexosamine biosynthetic pathway (HBP) is implicated in increased plasminogen activator inhibitor-1 (PAI-1), and endothelial nitric oxide synthase (eNOS) dysfunction in diabetes. Glucosamine (GlcN) that directly activates HBP is a dietary supplement and is clinically used to treat osteoarthritis despite uncertain efficacy and adverse cardiovascular effects observed in animal models. p38 mitogen-activated protein kinase (p38mapk) has been shown to be involved in HBP-mediated biological processes. The aim of the present study was to investigate the role of p38mapk in GlcN-induced endothelial PAI-1 expression and eNOS dysfunction. In cultured human endothelial cells, GlcN time- and concentration-dependently increased PAI-1 protein level that was further enhanced by tumor necrosis factor (TNF)-α, which was accompanied by a transient synergistic activation of p38mapk. The stimulation of PAI-1 by GlcN alone or by GlcN and TNF-α in combination was inhibited by the specific inhibitor of p38mapk, but not that of JNK or ERK1/2. Moreover, in isolated mouse aortas, GlcN caused eNOS uncoupling resulting in enhanced superoxide and decreased NO production, as well as impaired endothelium-dependent relaxations, which were also fully prevented by the p38mapk inhibitor. HBP activated by GlcN increases PAI-1 expression and eNOS uncoupling depending on p38mapk, which not only explains hyperglycemic vascular complications, but also may bring into question the clinical use of GlcN. The present results, support currently ongoing clinical application of p38mapk inhibitor in patients with cardiovascular disease.

Poly(ADP-ribose) polymerase-1 in high glucose-induced epithelial-mesenchymal transition during peritoneal fibrosis

Peritoneal fibrosis is a major complication of continuous ambulatory peritoneal dialysis (CAPD). The present study tested the hypothesis that ADP-ribose polymerase-1 (PARP-1) may play a role in peritoneal epithelial-mesenchymal transition and fibrosis under high glucose conditions. High glucose (126 mmol/l)-induced peritoneal EMT and fibrosis via the PARP-1 mechanism was examined in the primary culture of rat peritoneal mesothelial cells (PMCs) and in the human peritoneal mesothelial cell line (HMrSv5) in the presence or absence of a PARP-1 inhibitor PJ34 (3x10-6 M) or by knocking down PARP-1 with the PARP-1 siRNA technique. High glucose significantly increased PARP-1 expression and EMT as demonstrated by de novo expression of a mesenchymal marker α-SMA and loss of epithelial phenotype E-cadherin by both rat and human PMC, resulting in peritoneal fibrosis including up-regulation of plasminogen activator inhibitor-1 (PAI-1), collagen I, and fibronectin mRNA and protein expression. All these fibrotic responses induced by high glucose were significantly inhibited by the PARP-1 inhibitor PJ34 (all P<0.05) or by knocking down PARP-1 with the siRNA technique. Results from this study suggested that high glucose stimulates peritoneal EMT and fibrosis via a PARP-1-dependent mechanism, and targeting the PARP-1 may represent an alternative therapeutic potential for CAPD-related peritoneal fibrosis.

Compound Astragalus and Salvia miltiorrhiza extract inhibits cell proliferation, invasion and collagen synthesis in keloid fibroblasts by mediating transforming growth factor-β / Smad pathway

The transforming growth factor (TGF)-β/Smad pathway plays a key role in keloid development. We have previously demonstrated that compound Astragalus and Salvia miltiorrhiza extract (CASE) inhibits liver fibrosis and reduces invasion capacity of HepG2 cells by mediating the TGF-β/Smad pathway. We therefore hypothesize that CASE may also exert antifibrotic effects in keloids by mediating the TGF-β/Smad pathway. To investigate the effects of CASE on cell proliferation, invasion and collagen synthesis in keloid fibroblasts, and to explore the effects of CASE on the TGF-β/Smad signal pathway in order to elucidate its mechanisms of action. The inhibitory effects of CASE on keloid fibroblasts were evaluated. Cell proliferation was studied by MTT assay; cell invasion was observed utilizing Transwell invasion chambers; and collagen synthesis in keloid fibroblasts was measured by (3) H-proline incorporation assay. Expression of proteins induced by TGF-β1 and their intracellular localization in keloid fibroblasts were investigated by Western blot and immunofluorescence, respectively. Plasminogen activator inhibitor-1 (PAI-1) transcriptional activity was measured by real-time reverse transcription-polymerase chain reaction. CASE significantly inhibited cell proliferation induced by newborn bovine serum as well as collagen synthesis and cell invasion induced by TGF-β1 in keloid fibroblasts, while it showed weak effects on normal fibroblasts. The phosphorylation of Smad2/3 was markedly reduced by CASE treatment, while CASE exhibited stronger inhibitory effects on linker region phosphorylation (pSmad2L and pSmad3L) compared with effects on C-terminal region phosphorylation (pSmad2C and pSmad3C). In addition, CASE blocked formation of Smad2/3/4 complexes and their nuclear translocation, but upregulated Smad7 expression in a dose-dependent manner. PAI-1 mRNA and protein levels were also suppressed by CASE treatment. These results suggest that CASE exhibits inhibitory effects on cell proliferation, invasion and collagen synthesis in keloid fibroblasts, and its mechanisms of action may involve the TGF-β/Smad pathway.

Pseudomonas aeruginosa toxin ExoU induces a PAF-dependent impairment of alveolar fibrin turnover secondary to enhanced activation of coagulation and increased expression of plasminogen activator inhibitor-1 in the course of mice pneumosepsis

BackgroundExoU, a Pseudomonas aeruginosa cytotoxin with phospholipase A2 activity, was shown to induce vascular hyperpermeability and thrombus formation in a murine model of pneumosepsis. In this study, we investigated the toxin ability to induce alterations in pulmonary fibrinolysis and the contribution of the platelet activating factor (PAF) in the ExoU-induced overexpression of plasminogen activator inhibitor-1 (PAI-1).MethodsMice were intratracheally instilled with the ExoU producing PA103 P. aeruginosa or its mutant with deletion of the exoU gene. After 24 h, animal bronchoalveolar lavage fluids (BALF) were analyzed and lung sections were submitted to fibrin and PAI-1 immunohistochemical localization. Supernatants from A549 airway epithelial cells and THP-1 macrophage cultures infected with both bacterial strains were also analyzed at 24 h post-infection.ResultsIn PA103-infected mice, but not in control animals or in mice infected with the bacterial mutant, extensive fibrin deposition was detected in lung parenchyma and microvasculature whereas mice BALF exhibited elevated tissue factor-dependent procoagulant activity and PAI-1 concentration. ExoU-triggered PAI-1 overexpression was confirmed by immunohistochemistry. In in vitro assays, PA103-infected A549 cells exhibited overexpression of PAI-1 mRNA. Increased concentration of PAI-1 protein was detected in both A549 and THP-1 culture supernatants. Mice treatment with a PAF antagonist prior to PA103 infection reduced significantly PAI-1 concentrations in mice BALF. Similarly, A549 cell treatment with an antibody against PAF receptor significantly reduced PAI-1 mRNA expression and PAI-1 concentrations in cell supernatants, respectively.ConclusionExoU was shown to induce disturbed fibrin turnover, secondary to enhanced procoagulant and antifibrinolytic activity during P. aeruginosa pneumosepsis, by a PAF-dependent mechanism. Besides its possible pathophysiological relevance, in vitro detection of exoU gene in bacterial clinical isolates warrants investigation as a predictor of outcome of patients with P. aeruginosa pneumonia/sepsis and as a marker to guide treatment strategies.

Plasminogen activator inhibitor-1 (PAI-1) 4 G/5 G polymorphism and endometrial cancer. Influence of PAI-1 polymorphism on tissue PAI-1 antigen and mRNA expression and tumor severity

Plasminogen activator inhibitor-1 (PAI-1) 4G/5G polymorphism may have significance for PAI-1 expression. High levels of PAI-1 in endometrial cancer patients are associated with a poor prognosis. The objective of this study was to evaluate the PAI-1 4G/5G polymorphism in women with and without endometrial cancer and to analyze the influence of this polymorphism on PAI-1 expression in endometrial tissue.In 423 women (212 patients with endometrial cancer and 211 controls) PAI-1 4G/5G polymorphism was determined by PCR amplification using allele-specific primers. Quantitative real-time RT-PCR assay was used to quantify PAI-1 mRNA and PAI-1 protein levels were quantified by ELISA in tissue extracts from 33 patients with endometrial cancer and from 70 endometrial tissues from control women. The frequency of PAI-1 4G/4G genotype (P=0.010) and the PAI-1 4G allele (P=0.009) was significantly higher in patients than in controls. The frequency of PAI-1 4G allele was significantly higher in patients with stage IB than in those with stage IA (P=0.03). Control women with the 4G/4G genotype had higher endometrial PAI-1 protein (P=0.018) and mRNA (P=0.004) levels than those with the 5G/5G genotype. A significant increase in PAI-1 protein and mRNA was observed in endometrial cancer tissue in comparison with the endometrial tissue from control women (P<0.01). In conclusion, frequencies of the PAI-1 4G allele and 4G/4G genotype were found significantly more often in women with endometrial cancer than in controls. PAI-1 levels in endometrial tissue seem to be associated with PAI-1 4G/5G polymorphism. These findings suggest that the PAI-1 4G/4G genotype may be associated with the risk of endometrial cancer in a Caucasian population. Further studies with a larger number of patients are needed to clarify the influence of this PAI-1 polymorphism in endometrial cancer.

Conventional and novel cardiovascular risk factors and markers of vascular damage in rural and urban Indian men

BackgroundIndia is undergoing rapid epidemiological and nutritional transition largely as a consequence of rapid urbanisation. We investigated conventional and novel cardiovascular risk factors in rural and urban Indian men and studied their association with markers of vascular damage. MethodsWe randomly selected and studied 149 rural, 142 urban slum residents and 150 urban middle class middle aged Indian men. We measured conventional (obesity, blood pressure, lipids, smoking habits) and novel (proinflammatory and prothrombotic factors) cardiovascular risk factors and markers of vascular damage (carotid intima media thickness (IMT), von Willebrand Factor (vWF), e-selectin). ResultsThere was a progressive increase in most of the conventional cardiovascular (CV) risk factors from rural to slum to urban middle class men. Plasminogen activator inhibitor-1 (PAI-1), platelet count, total homocysteine and C-reactive protein showed similar patterns. Carotid IMT was similar in the three groups; vWF was highest in rural and e-selectin in slum men. Adjusting for location, age explained 17%, obesity 3% and conventional risk factors 1% of the variance in carotid IMT, whilst novel cardiovascular risk factors were without any significant impact. ConclusionsUrbanisation increases obesity related as well as prothrombotic and proinflammatory CV risk factors in Indian men, but appears not to impact on IMT.

Effects of telmisartan and losartan on cardiovascular protection in Japanese hypertensive patients

The Telmisartan and Losartan Cardiac Evaluation Trial, a multicenter, prospective, randomized, open-labeled, blinded-endpoint trial, was designed to compare the effects of two angiotensin II receptor blockers (ARBs), telmisartan and losartan, on cardiovascular protection in Japanese patients with mild to moderate essential hypertension. We compared the effects of telmisartan and losartan on left ventricular (LV) hypertrophy, cardiac function, atherosclerosis of carotid arteries and surrogate markers related to the actions of peroxisome proliferator-activated receptor-γ. A total of 58 patients were enrolled in the present trial and the follow-up period was 1 year. There were no significant differences in blood pressure (BP) levels between the telmisartan group and the losartan group throughout the trial. The percentage of the patients treated with ARB monotherapy was significantly higher in the telmisartan group compared with the losartan group. In addition, the progression of intima-media thickness of common carotid artery was significantly inhibited in the telmisartan group compared with the losartan group. Neither group experienced significant changes in cardiac function and LV mass index. There were no differences between the groups with respect to changes in surrogate markers such as serum adiponectin, creatinine, homeostasis model assessment index, plasminogen activator inhibitor-1 and high sensitivity C-reactive protein. Although BP levels were equal and well controlled in both groups, telmisartan showed more protective vascular effects than losartan.

PAI-1 is an essential component of the pulmonary host response during Pseudomonas aeruginosa pneumonia in mice

RationaleElevated plasma and bronchoalveolar lavage fluid plasminogen activator inhibitor 1 (PAI-1) levels are associated with adverse clinical outcome in patients with pneumonia caused by Pseudomonas aeruginosa. However, whether PAI-1 plays...