Altered fibrin clot properties are associated with the progression of chronic kidney disease in atrial fibrillation.

INTRODUCTION: Formation of denser and resistant to lysis fibrin clot networks has been shown in chronic kidney disease (CKD) and atrial fibrillation (AF). We investigated whether such prothrombotic fibrin clot properties are associated with faster progression of CKD in AF patients. MATERIAL AND METHODS: We recruited 265 AF patients (men 49.1 %, median age of 64.0 years, median estimated glomerular filtration rate [eGFR] of 77.0 ml/min/1.73 m2), including 137 patients on non-vitamin K antagonist oral anticoagulants (NOACs) (51.7 %) and 109 patients (41.1 %) on vitamin K antagonists (VKAs). At baseline while off anticoagulation, we determined fibrin clot permeability (Ks), and clot lysis time (CLT), along with plasminogen activator inhibitor-1 (PAI-1), endogenous thrombin potential (ETP), and von Willebrand factor (vWF). The kidney function was assessed at baseline and after a median follow-up of 50.0 months. RESULTS: During follow-up, a median eGFR decreased by 8.0 (5.0-11.0) ml/min/1.73 m2, 1.8 ml/min/1.73 m2/year and this change correlated with age (R = 0.19, P = 0.002), Ks (R = 0.46, P < 0.0001), and CLT (R = -0.17, P = 0.005), but not ETP, fibrinogen, PAI-1 or vWF. A decrease in eGFR was lower in patients who used NOACs at baseline but not in those who started NOACs during follow-up (n = 101) as compared to the remaining patients. On multiple linear regression analysis, adjusted for age and fibrinogen, baseline Ks, eGFR, hypertension, and NOACs use independently predicted a decrease in eGFR. CONCLUSIONS: This study is the first to show that more compact fibrin clot networks may contribute to faster progression of CKD in AF, indicating novel kidney-related harmful effects of prothrombotic clot properties in humans.

Alantolactone Attenuates Renal Fibrosis via Inhibition of Transforming Growth Factor ß/Smad3 Signaling Pathway.

BACKGRUOUND: Renal fibrosis is characterized by the accumulation of extracellular matrix proteins and interstitial fibrosis. Alantolactone is known to exert anticancer, anti-inflammatory, antimicrobial and antifungal effects; however, its effects on renal fibrosis remains unknown. Here, we investigated whether alantolactone attenuates renal fibrosis in mice unilateral ureteral obstruction (UUO) and evaluated the effect of alantolactone on transforming growth factor (TGF) signaling pathway in renal cells. METHODS: To evaluate the therapeutic effect of alantolactone, cell counting kit-8 (CCK-8) assay, histological staining, Western blot analysis, and real-time quantitative polymerase chain reaction were performed in UUO kidneys in vivo and in TGF-ß-treated renal cells in vitro. RESULTS: Alantolactone (0.25 to 4 µM) did not affect the viability of renal cells. Mice orally administered 5 mg/kg of alantolactone daily for 15 days did not show mortality or liver toxicity. Alantolactone decreased UUO-induced blood urea nitrogen and serum creatinine levels. In addition, it significantly alleviated renal tubulointerstitial damage and fibrosis and decreased collagen type I, fibronectin, and α-smooth muscle actin (α-SMA) expression in UUO kidneys. In NRK-49F cells, alantolactone inhibited TGF-ßstimulated expression of fibronectin, collagen type I, plasminogen activator inhibitor-1 (PAI-1), and α-SMA. In HK-2 cells, alantolactone inhibited TGF-ß-stimulated expression of collagen type I and PAI-1. Alantolactone inhibited UUO-induced phosphorylation of Smad3 in UUO kidneys. In addition, it not only decreased TGF-ß secretion but also Smad3 phosphorylation and translocation to nucleus in both kidney cell lines. CONCLUSION: Alantolactone improves renal fibrosis by inhibiting the TGF-ß/Smad3 signaling pathway in obstructive nephropathy. Thus, alantolactone is a potential therapeutic agent for chronic kidney disease.

Endothelial extracellular vesicles induce acute lung injury via follistatin-like protein 1.

Cardiopulmonary bypass has been speculated to elicit systemic inflammation to initiate acute lung injury (ALI), including acute respiratory distress syndrome (ARDS), in patients after cardiac surgery. We previously found that post-operative patients showed an increase in endothelial cell-derived extracellular vesicles (eEVs) with components of coagulation and acute inflammatory responses. However, the mechanism underlying the onset of ALI owing to the release of eEVs after cardiopulmonary bypass, remains unclear. Plasma plasminogen-activated inhibitor-1 (PAI-1) and eEV levels were measured in patients with cardiopulmonary bypass. Endothelial cells and mice (C57BL/6, Toll-like receptor 4 knockout (TLR4-/-) and inducible nitric oxide synthase knockout (iNOS-/-)) were challenged with eEVs isolated from PAI-1-stimulated endothelial cells. Plasma PAI-1 and eEVs were remarkably enhanced after cardiopulmonary bypass. Plasma PAI-1 elevation was positively correlated with the increase in eEVs. The increase in plasma PAI-1 and eEV levels was associated with post-operative ARDS. The eEVs derived from PAI-1-stimulated endothelial cells could recognize TLR4 to stimulate a downstream signaling cascade identified as the Janus kinase 2/3 (JAK2/3)-signal transducer and activator of transcription 3 (STAT3)-interferon regulatory factor 1 (IRF-1) pathway, along with iNOS induction, and cytokine/chemokine production in vascular endothelial cells and C57BL/6 mice, ultimately contributing to ALI. ALI could be attenuated by JAK2/3 or STAT3 inhibitors (AG490 or S3I-201, respectively), and was relieved in TLR4-/- and iNOS-/- mice. eEVs activate the TLR4/JAK3/STAT3/IRF-1 signaling pathway to induce ALI/ARDS by delivering follistatin-like protein 1 (FSTL1), and FSTL1 knockdown in eEVs alleviates eEV-induced ALI/ARDS. Our data thus demonstrate that cardiopulmonary bypass may increase plasma PAI-1 levels to induce FSTL1-enriched eEVs, which target the TLR4-mediated JAK2/3/STAT3/IRF-1 signaling cascade and form a positive feedback loop, leading to ALI/ARDS after cardiac surgery. Our findings provide new insight into the molecular mechanisms and therapeutic targets for ALI/ARDS after cardiac surgery.

Pharmacological inhibition of plasminogen activator inhibitor-1 prevents memory deficits and reduces neuropathology in APP/PS1 mice.

RATIONALE: Extracellular proteolytic activity plays an important role in memory formation and the preservation of cognitive function. Previous studies have shown increased levels of plasminogen activator inhibitor-1 (PAI-1) in the brain of mouse models of Alzheimer's disease (AD) and plasma of AD patients, associated with memory and cognitive decline; however, the exact function of PAI-1 in AD onset and progression is largely unclear. OBJECTIVE: In this study, we evaluated a novel PAI-1 inhibitor, TM5A15, on its ability to prevent or reverse memory deficits and decrease Aß levels and plaque deposition in APP/PS1 mice. METHODS: We administered TM5A15 mixed in a chow diet to 3-month and 9-month-old APP/PS1 mice before and after neuropathological changes were distinguishable. We then evaluated the effects of TM5A15 on memory function and neuropathology at 9 months and 18 months of age. RESULTS: In the younger mice, 6 months of TM5A15 treatment protected against recognition and short-term working memory impairment. TM5A15 also decreased oligomer levels and amyloid plaques, and increased mBDNF expression in APP/PS1 mice at 9 months of age. In aged mice, 9 months of TM5A15 treatment did not significantly improve memory function nor decrease amyloid plaques. However, TM5A15 treatment showed a trend in decreasing oligomer levels in APP/PS1 mice at 18 months of age. CONCLUSION: Our results suggest that PAI-1 inhibition could improve memory function and reduce the accumulation of amyloid levels in APP/PS1 mice. Such effects are more prominent when TM5A15 is administered before advanced AD pathology and memory deficits occur.

Immunomodulation Effect of Regional Citrate Anticoagulation in Acute Kidney Injury Requiring Renal Replacement Therapy.

INTRODUCTION: Acute kidney injury (AKI) is a common syndrome in critically ill patients. Continuous renal replacement therapy (CRRT) is the standard treatment for patients with AKI. Research on the immunomodulating effects of regional citrate anticoagulation (RCA) remains limited in patients with AKI receiving CRRT. We aimed to evaluate the immunomodulating effects of RCA in patients with AKI receiving CRRT. METHODS: A randomized controlled trial study on critically ill adult patients with AKI undergoing CRRT was undertaken. Participants were randomized into either a regional citrate group or control group (either heparin anticoagulant or normal saline). Measurements were taken at baseline, 6 and 24 h after commencing CRRT for CD11b expression, C3a, C5a, and plasminogen activator inhibitor-1 (PAI-1) levels. Clinical outcomes assessed were 28-day survival rate, length of ICU stay, renal support duration, and renal function at discharge. RESULTS: Thirty patients were recruited and randomized into 2 groups of 15 subjects. Baseline demographic and clinical data were comparable between groups. In the citrate group, CD11b expression was significantly decreased at 24 h compared to the control group (1.84% [1.18-3.32] versus 4.92% [2.63-6.93], p < 0.01). The complement level, including c3b and c5a, was stable during CRRT. Additionally, the PAI-1 levels were significantly decreased at 24 h compared to the control group (114 ng/mL [19-193] versus 359 ng/mL [264-491], p < 0.01). No significant difference in survival rate was observed. CONCLUSIONS: RCA may have the potential to mitigate the inflammatory response by decreasing CD11b expression of neutrophil and improve fibrinolysis activity through a reduction of PAI-1 levels. Larger clinical trials are warranted to test this immunomodulation effect of RCA.

Oral contraceptive pill (OCP) treatment alters the gene expression of intercellular adhesion molecule-1 (ICAM-1), tumor necrosis factor-α (TNF-α), monocyte chemoattractant protein-1 (MCP-1) and plasminogen activator inhibitor-1 (PAI-1) in polycystic ovary syndrome (PCOS) women compared to drug-naive PCOS women.

BACKGROUND: Polycystic ovary syndrome (PCOS) presents clinical symptoms of menstrual abnormalities, excessive hair growth (hirsutism), scalp hair loss, acne and infertility. Metabolic abnormalities such as obesity, insulin resistance, glucose intolerance and cardiovascular problems constitute an essential part of PCOS, all of which can have significant long-term health consequences. Low-grade chronic inflammation demonstrated by persistent moderately elevated serum levels of inflammatory and coagulatory markers plays a critical role in the pathogenesis of PCOS. Oral contraceptive pills (OCPs) constitute the mainstay of pharmacologic therapy for women with PCOS to regularize cyclicity and ameliorate androgen excess. On the other hand, OCP use is associated with various venous thromboembolic and proinflammatory events in the general population. PCOS women always carriers the increased lifetime risk of these events. The studies on the effect of OCPs on inflammatory, coagulation and metabolic parameters in PCOS are less robust. Therefore in this study, we investigated and compared the messenger RNA (mRNA) expression profiles of genes implicated in inflammatory and coagulation pathways between drug-naive and OCP-treated PCOS women. The selected genes include intercellular adhesion molecule-1 (ICAM-1), tumor necrosis factor-α (TNF-α), monocyte chemoattractant protein-1 (MCP-1) and plasminogen activator inhibitor-1 (PAI-1). Furthermore, the correlation between the selected markers and various metabolic indices in the OCP group has also been explored. METHOD: The relative amounts of ICAM-1, TNF-α, MCP-1 and PAI-1 mRNA in peripheral blood mononuclear cells from 25 drug-naive PCOS subjects (controls) and 25 PCOS subjects who received OCPs containing 0.03 mg-ethinyl-estradiol and 0.15 mg-levonorgestrel for at least six months (cases) were estimated using real-time qPCR. The statistical interpretation was conducted using SPSS version 20.0 (SPSS, Inc, Chicago, IL), Epi Info version 2002 (Disease Control and Prevention Centres, Atlanta, GA) and GraphPad Prism 5 (GraphPad Software, La Jolla, CA) software. RESULT: Six months of OCP therapy enhanced the expression of inflammatory genes viz ICAM-1, TNF-α and MCP-1 mRNA in PCOS women by 2.54, 2.05 and 1.74 folds, respectively, in this study. However, PAI-1 mRNA in the OCP group showed no significant increase. Furthermore, in cases, ICAM-1 mRNA expression positively correlated with body mass index (BMI) (p = 0.01), fasting insulin (p = 0.01), insulin 2 h p = 0.02), glucose 2 h (p = 0.01) and triglycerides (p = 0.01). TNF-α mRNA expression positively correlated with fasting insulin (p = 0.0007). MCP-1 mRNA expression positively correlated with (BMI) (p = 0.002). CONCLUSION: OCPs helped reduce clinical hyperandrogenism and regularise menstrual cycles in women with PCOS. However, OCP use was associated with increased fold expression of inflammatory markers which positively correlated with metabolic abnormalities.

Oral 24-week probiotics supplementation did not decrease cardiovascular risk markers in patients with biopsy proven NASH: A double-blind placebo-controlled randomized study.

INTRODUCTION AND OBJECTIVES: Cardiovascular disease (CVD) is the major cause of death in non-alcoholic fatty liver disease (NAFLD), a clinical condition without any approved pharmacological therapy. Probiotics are often indicated for the disease, but their results are controversial in part due to the poor quality of studies. Thus, we investigated the impact of 24-week probiotics supplementation on cardiovascular risk (CVR) in biopsy-proven non-alcoholic steatohepatitis (NASH) patients. PATIENTS AND METHODS: Double-blind, placebo-controlled, single-center study (NCT03467282), adult NASH, randomized for 24 weeks daily sachets of probiotic mix (109CFU of Lactobacillus acidophilus, Lactobacillus rhamnosus, Lactobacillus paracasei and Bifidobacterium lactis) or placebo. Clinical scores (atherogenic indexes, atherosclerotic cardiovascular disease-ASCVD and systematic coronary risk evaluation-SCORE), biochemistry, miR-122, miR-33a, plasminogen activator inhibitor-1 (PAI-1), intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), were determined before and after the intervention. RESULTS: Forty-six patients were enrolled (23 received probiotics and 23 placebo), with a mean age of 51.7 years, most of them females and whites. Clinical and demographic features were similar between the groups at the baseline. The Median NAFLD activity score was 4.13 in both groups. Fibrosis was mild in most patients (15.2% and 65.2% F0 and F1, respectively). Treatment did not promote any clinically significant changes in body mass index or laboratory, including lipid and glucose profile. High CVR patients through atherogenic indexes decreased from baseline in both groups, as well as PAI-1 and miR-122 levels, although there was no difference between probiotics and placebo. CONCLUSIONS: A 24-week probiotic mix administration was not superior to placebo in reducing CVR markers in patients with NASH.

Application of Machine Learning to Assess Interindividual Variability in Rapid-Acting Insulin Responses After Subcutaneous Injection in People With Type 1 Diabetes.

OBJECTIVES: Circulating insulin concentrations mediate vascular-inflammatory and prothrombotic factors. However, it is unknown whether interindividual differences in circulating insulin levels are associated with different inflammatory and prothrombotic profiles in type 1 diabetes (T1D). We applied an unsupervised machine-learning approach to determine whether interindividual differences in rapid-acting insulin levels associate with parameters of vascular health in patients with T1D. METHODS: We re-analyzed baseline pretreatment meal-tolerance test data from 2 randomized controlled trials in which 32 patients consumed a mixed-macronutrient meal and self-administered a single dose of rapid-acting insulin individualized by carbohydrate counting. Postprandial serum insulin, tumour necrosis factor (TNF)-alpha, plasma fibrinogen, human tissue factor (HTF) activity and plasminogen activator inhibitor-1 (PAI-1) were measured. Two-step clustering categorized individuals based on shared clinical characteristics. For analyses, insulin pharmacokinetic summary statistics were normalized, allowing standardized intraindividual comparisons. RESULTS: Despite standardization of insulin dose, individuals exhibited marked interpersonal variability in peak insulin concentrations (48.63%), time to peak (64.95%) and insulin incremental area under the curve (60.34%). Two clusters were computed: cluster 1 (n=14), representing increased serum insulin concentrations; and cluster 2 (n=18), representing reduced serum insulin concentrations (cluster 1: 389.50±177.10 pmol/L/IU h-1; cluster 2: 164.29±41.91 pmol/L/IU h-1; p<0.001). Cluster 2 was characterized by increased levels of fibrinogen, PAI-1, TNF-alpha and HTF activity; higher glycated hemoglobin; increased body mass index; lower estimated glucose disposal rate (increased insulin resistance); older age; and longer diabetes duration (p<0.05 for all analyses). CONCLUSIONS: Reduced serum insulin concentrations are associated with insulin resistance and a prothrombotic milieu in individuals with T1D, and therefore may be a marker of adverse vascular outcome.

Plasminogen Activator Inhibitor-1 as a Marker of Esophageal Functional Changes in Pediatric Eosinophilic Esophagitis.

BACKGROUND & AIMS: Esophageal remodeling in eosinophilic esophagitis (EoE) can lead to esophageal rigidity with eventual luminal compromise and stenoses. Gauging esophageal functional alterations in EoE is challenging. An epithelial marker of functional remodeling would impact EoE management. METHODS: Esophageal biopsy specimens from children with and without EoE and primary human esophageal epithelial cells were used for PAI-1 immunohistochemistry, and cell proliferation experiments. PAI-1 immunostaining and basal cell hyperplasia were assessed in the context of concurrently obtained esophageal compliance measures on endoscopic functional lumen imaging probe (EndoFLIP). RESULTS: EndoFLIPs were performed in 45 children (32 with and 13 without EoE). Epithelial PAI-1 was increased in patients with active EoE versus inactive or control patients (P < .01). Esophageal compliance was lower in EoE patients versus controls, particularly in the proximal esophagus (P < .001). Proximal compliance was the strongest predictor of EoE (AUROC 0.88, 95% CI 0.77, 0.98) with esophageal compliance of less than 2.6%mL/mmHg demonstrating 82% sensitivity and 84% specificity for EoE. PAI-1 inhibition significantly diminished esophageal epithelial cell proliferation, suggesting PAI-1 could trigger basal cell hyperplasia. A composite mid-esophageal BZH + PAI-1 score was the strongest predictor of altered compliance (P = .02, AUROC 0.89 (95% CI 0.80, 0.99). CONCLUSIONS: PAI-1 is significantly elevated in pediatric EoE and distinguishes altered compliance in children. PAI-1 may be a novel disease marker and therapeutic target.

[uPA/PAI-1 and EPClin®: Comparison of their impact on the management of intermediate-prognosis breast cancers]. / uPA/PAI-1 et EPClin® : comparaison de leur impact sur la prise en charge des cancers du sein de pronostic intermédiaire.

OBJECTIVE: The uPA/PAI-1 assay and the EPClin® test are useful tools that add to clinico-anatomical characteristics to determine the indication of adjuvant chemotherapy in case of intermediate-prognosis invasive breast cancer. The principal purpose of our study was to analyze the concordance of uPA/PAI-1 and EPClin® in classification of patients into two groups: low and high risk of relapse. METHODS: We prospectively included 63 patients treated for intermediate-prognosis invasive breast cancer. All of these patients received a uPA/PAI-1 assay and an EPClin® test. RESULTS: The uPA/PAI-1 assay and EPClin® test were consistent for 56.2% and inconsistent for 43.8%. In the event of a discrepancy, the treatment decision was based in 95.2% of patients on the EPClin® test result. In total, 38 patients were selected for adjuvant chemotherapy after achievement of the two tests. The mean time to report results after surgery was 9 days for the uPA/PAI-1 assay and 35 days for the EPClin® test. No cases of recurrence or death were found, with an average follow-up of 32 months. CONCLUSION: The EPClin® test resulted in more chemotherapy prescriptions than indicated by uPA/PAI-1. However, we can't conclude to the superiority of one of these two tests, survival data and the effectiveness of our study being insufficient. In general, studies comparing different signatures useful to the therapeutic decision of intermediate prognosis breast cancers should be encouraged.

Inhibition of PAI-1 Promotes Lipolysis and Enhances Weight Loss in Obese Mice.

OBJECTIVE: This study investigates the therapeutic potential of a small molecule inhibitor of plasminogen activator inhibitor-1 (PAI-1), TM5441, in reversing diet-induced obesity in mice. METHODS: Wild-type C57BL/6J mice were fed a high-fat high-sugar (HFHS) diet for 8 weeks to induce obesity. After the first 8 weeks, TM5441 was added to the diet for an additional 8 weeks. In order to determine the efficacy of PAI-1 inhibition in conjunction with dietary modification, mice were fed an HFHS diet for 8 weeks to induce obesity and were then switched to a low-fat diet with or without TM5441 for an additional 2 to 8 weeks. RESULTS: Obese mice showed weight reduction and significant improvement in hepatic steatosis when TM5441 was added to the HFHS diet. Obese mice that were treated with TM5441 in conjunction with dietary modification showed enhanced weight loss and a more rapid reversal of hepatic steatosis compared with obese mice treated with dietary modification alone. The enhanced weight loss among mice treated with TM5441 was associated with increased adipose tissue expression of adipose triglyceride lipase, phosphorylated hormone-sensitive lipase, and phosphorylated perilipin-1 as well as induction of adipose tissue lipolysis. CONCLUSIONS: Pharmacologic PAI-1 inhibition stimulates adipose tissue lipolysis and enhances weight loss in obese mice.

Glycated Haemogloin, Fasting Plasma Glucose, Plasminogen Activator Inhibitor Type-1, and Soluble Thrombomodulin Levels in Patients with Type 2 Diabetes Mellitus.

Diabetes mellitus has become increasingly prevalent over the years. The chronic hyperglycaemia of diabetes is associated with long-term damage, dysfunctions, and failure of different organs suggesting that the most effective tool to prevent complications is the effective control of hyperglycaemia itself. The study is set to determine the effect of glycemic control on plasminogen activator inhibitor type 1 (PAI-1), soluble thrombomodulin (STM) alongside fasting plasma glucose (FPG) and glycated hemoglobin (HbA1c) among type 2 diabetic subjects. One hundred diabetic subjects accessing care at the University of Calabar Teaching Hospital Calabar and 100 non -diabetics that served as controls were enrolled. Blood samples from participants were analyzed for FPG, HbA1c, PAI-1 and STM by standard methods. The result shows 74% of the diabetic to be females. Half of the diabetics were managed on only oral anti-diabetic drugs while the remaining half were either on insulin injection or a combination of oral and insulin injection.  Poor glycemic control was observed in 56% of the studied subjects. The mean age of 54.69 ± 9.94 years for the diabetics was comparable to the age-matched controls (p=.097). Diabetics showed significantly higher FPG, HbA1c, PAI-1and STM (P=0.001) compared to control values. Correlations between STM, PAI 1 and glycated hemoglobin (figures 2 p=0.001, p =0.001) and STM, PAI-1 and FPG revealed significantly robust association (p=0.001, p=0.001).  The study concludes that there is poor glycemic control among the treated diabetic subjects with PAI-1 and STM showing a very strong positive correlation with HbA1c than FPG.

Tissue-type plasminogen activator and plasminogen activator inhibitor type 1 in patients with symptomatic lower extremity artery disease.

BACKGROUND: Lower extremity artery disease (LEAD) involves progressive arterial narrowing manifested by intermittent claudication (IC). LEAD entails endothelial dysfunction and fibrinolytic disorders. In the current study, we analyze the selected parameters of the fibrinolytic system in the blood of patients with symptomatic LEAD depending on clinical parameters. METHODS: The test group was comprised of 80 patients with diagnosis of LEAD based on Ankle-Brachial Index (ABI) test (27 female/53 male) with an average age of 63.5±9 years. The control group included 30 healthy, non-smoking volunteers (10 female/20 male), with the median age of 56±6 years. The research material - venous blood - was sampled to determine the concentrations of tissue-type plasminogen activator (t-PA Ag), plasminogen activator inhibitor type 1 (PAI-1 Ag), D-dimer, fibrinogen, and platelet count (PLT). RESULTS: We found elevated concentrations of t-PA Ag, PAI-1 Ag, D-dimer, and fibrinogen in the plasma of subjects with symptomatic LEAD. Various stages of the Fontaine classification demonstrated a gradual, statistically significant increase in the concentrations of fibrinogen and PLT count as the disease progressed. More so, in the subgroup of LEAD patients aged ≥65 years, we observed significantly higher levels of D-dimer than in the group of younger subjects. In addition to that, the LEAD group demonstrated negative correlations of IC distance, fibrinogen concentrations, and PLT count, negative correlations of ABI at rest and concentrations of D-dimer and PLT count, as well as positive correlations between age and D-dimer levels. CONCLUSIONS: High t-PA Ag concentrations in LEAD patients suggest damage to the endothelium which comprises the main source of this factor. With high PAI-1 Ag levels, inactive fibrinolytic t-PA-PAI-1 complexes are formed. Increasing fibrinogen concentrations at the subsequent stages in accordance with the Fontaine classification, indicate increasing inflammation. Moreover, heightened values of D-dimer reflect an increased secondary fibrinolysis activation as patients get older and impaired extremity vascularization, manifested by the decreasing ABI, progresses.

A novel PAI-1 inhibitor prevents ageing-related muscle fiber atrophy.

Sarcopenia is among the most common medical problems of the aging population worldwide and a major social concern. Here, we explored the therapeutic potential of TM5484, a novel orally available PAI-1 inhibitor, to prevent sarcopenia. The sarcopenic phenotypes of the calf muscle of 12- and 6-month-old middle-aged mice were compared. Although significant decline of isometric gastrocnemius muscle force was detected in the older untreated mice, those administered TM5484 had significantly greater calf muscle force, as determined using isometric measurements by electrical stimulation. Histological analysis indicated that cross-sectional gastrocnemius muscle fibers in untreated older mice were thinner than those in younger mice; however, TM5484-treated group showed thicker fibers than younger mice. Treatment with TM5484 for 6 months enhanced Igf1, Atrogin-1, Mt-Co1, and Chrna1 mRNA expression in the mice gastrocnemius muscle, with increased serum IGF-1 concentration. TM5484 induced dose-dependent Igf1, Atrogin-1, and Chrna1 expression in C2C12 myoblastic cells, confirming cell autonomous effect. Further, the presence of plasmin for 72 h caused significantly increased Igf1 expression in C2C12 cells. These findings suggest that oral PAI-1 inhibitors represent a promising therapeutic candidate for preventing sarcopenia progression in humans.

Embelin ameliorated sepsis-induced disseminated intravascular coagulation intensities by simultaneously suppressing inflammation and thrombosis.

Disseminated intravascular coagulation (DIC), an acute syndrome of systemic thrombus formation in microvasculatures throughout the body, can be induced by severe infections, e.g. sepsis. Anticoagulants are clinically used to alleviate the intensities of DIC. However, anticoagulants only reduce the thrombus formation but have negligible effects on the inflammatory conditions. We previously reported embelin, a natural product, as an inhibitor of plasminogen activator inhibitor-1 (PAI-1), suggesting the potent antithrombotic property. In this study, we used three thrombotic mice models to confirm the antithrombotic property of embelin. By combining the anti-inflammatory and the antithrombotic properties, we proposed embelin as a potent therapeutic agent for sepsis-induced DIC, which involves both inflammation and thrombosis. In a lipopolysaccharides-induced septic mice model, embelin not only significantly ameliorated the inflammation levels, but also effectively reduced the pulmonary hemorrhages and the micro-thrombi formations in lung. In contrast, low-molecular-weight-heparin, an anticoagulant, only moderately ameliorated the pulmonary hemorrhages and thrombotic obstructions, but had non-measurable effect on the inflammatory conditions. In addition, embelin alleviated the dysregulation of the global coagulation in septic mice, but did not affect the global coagulation in normal mice. Our current study demonstrates the antithrombotic property of embelin and the potency of the treatment or prevention of syndromes combining inflammation and thrombosis, e.g. sepsis-induced DIC.

Role of plasminogen activator inhibitor-1 in coronary pathophysiology.

The standard of care for obstructive atherosclerotic coronary disease is revascularization, predominantly achieved via percutaneous placement of a stent with concurrent medical therapy. Advancements in percutaneous coronary intervention (PCI) have dramatically improved outcomes. However, major complications from PCI due to target lesion failure continue to occur at rates between 5 and 10% in the first twelve months following intervention limiting its therapeutic efficacy. Plasminogen activator inhibitor-1 (PAI-1) is a protein of interest for both arterial remodeling and thrombotic risk as it regulates cell migration and vascular thrombosis. Elevated PAI-1 antigen levels have been identified as a potential biomarker for coronary artery disease and metabolic syndrome while being modulated by a number of atherosclerotic risk factors. Although linked by some studies as a marker of disease severity and prognosis, it remains to be understood whether it is also a mediator and/or therapeutic target of vascular disease. In this review, we discuss the current understanding of PAI-1 in vascular disease and its potential role in in-stent restenosis and stent thrombosis.

Is there a role for urokinase-type plasminogen activator inhibitors as maintenance therapy in patients with ovarian cancer?

There is abundant evidence that the urokinase-type plasminogen activator (uPA), its inhibitors PAI-1 and PAI-2 (plasminogen activator inhibitor type-1 and type-2) and its cells surface receptor (uPA-R, CD87) play a fundamental role in tumor invasion and metastasis and are of significant prognostic significance for many tumor types. We performed a systematic Med-line search on uPA, PAI, uPA-R and (epithelial) ovarian cancer (EOC). The majority of malignant EOC specimens show moderate to strong immunostating of tumor and stromal cells. Overexpression of u-PA and PAI-1 can be found in more the 75% of primary ovarian carcinomas, in most metastatic EOC samples and all examined epithelial ovarian cancer cell lines. uPA overexpression in primary specimens was significantly associated with tumor stage, grade, residual disease status after cytoreductive surgery, and poor clinical outcome. This may be explained by increased chemoresistance, a lower resectability and more aggressive tumor biology and tumor dissemination in patients with high uPA and PAI-1. Several therapeutical approaches aimed at inhibiting the uPA/uPAR functions have shown to possess anti-tumor effects in vitro and in animal models. When treating a patient with advanced ovarian cancer it may to be assumed that inhibiting the progression of established (micro) metastases may be more therapeutically relevant than trying to destroy all tumor cells which is not possible in most cases with current systemic treatment modalities. Taking into account the role of uPA and PAI in cell detachment, formation of new stroma, tumor cell reimplantation and metastasis uPA inhibition should be further investigated as maintenance treatment in patients with advanced EOC.

Truncated Plasminogen Activator Inhibitor-1 Protein Protects From Pulmonary Fibrosis Mediated by Irradiation in a Murine Model.

PURPOSE: To determine whether the delivery of recombinant truncated plasminogen activator inhibitor-1 (PAI-1) protein (rPAI-1(23)) would protect from the development of radiation-induced lung injury. METHODS AND MATERIALS: C57Bl/6 mice received intraperitoneal injections of rPAI-1(23) (5.4 µg/kg/d) or vehicle for 18 weeks, beginning 2 days before irradiation (IR) (5 daily fractions of 6 Gy). Cohorts of mice were followed for survival (n=8 per treatment) and tissue collection (n=3 per treatment and time point). Fibrosis in lung was assessed with Masson-Trichrome staining and measurement of hydroxyproline content. Senescence was assessed with staining for ß-galactosidase activity in lung and primary pneumocytes. RESULTS: Hydroxyproline content in irradiated lung was significantly reduced in mice that received rPAI-1(23) compared with mice that received vehicle (IR+vehicle: 84.97 µg/lung; IR+rPAI-1(23): 56.2 µg/lung, P=.001). C57Bl/6 mice exposed to IR+vehicle had dense foci of subpleural fibrosis at 19 weeks, whereas the lungs of mice exposed to IR+rPAI-1(23) were largely devoid of fibrotic foci. Cellular senescence was significantly decreased by rPAI-1(23) treatment in primary pneumocyte cultures and in lung at multiple time points after IR. CONCLUSIONS: These studies identify that rPAI-1(23) is capable of preventing radiation-induced fibrosis in murine lungs. These antifibrotic effects are associated with increased fibrin metabolism, enhanced matrix metalloproteinase-3 expression, and reduced senescence in type 2 pneumocytes. Thus, rPAI-1(23) is a novel therapeutic option for radiation-induced fibrosis.

Functional stability of plasminogen activator inhibitor-1.

Plasminogen activator inhibitor-1 (PAI-1) is the main inhibitor of plasminogen activators, such as tissue-type plasminogen activator (t-PA) and urokinase-type plasminogen activator (u-PA), and a major regulator of the fibrinolytic system. PAI-1 plays a pivotal role in acute thrombotic events such as deep vein thrombosis (DVT) and myocardial infarction (MI). The biological effects of PAI-1 extend far beyond thrombosis including its critical role in fibrotic disorders, atherosclerosis, renal and pulmonary fibrosis, type-2 diabetes, and cancer. The conversion of PAI-1 from the active to the latent conformation appears to be unique among serpins in that it occurs spontaneously at a relatively rapid rate. Latency transition is believed to represent a regulatory mechanism, reducing the risk of thrombosis from a prolonged antifibrinolytic action of PAI-1. Thus, relying solely on plasma concentrations of PAI-1 without assessing its function may be misleading in interpreting the role of PAI-1 in many complex diseases. Environmental conditions, interaction with other proteins, mutations, and glycosylation are the main factors that have a significant impact on the stability of the PAI-1 structure. This review provides an overview on the current knowledge on PAI-1 especially importance of PAI-1 level and stability and highlights the potential use of PAI-1 inhibitors for treating cardiovascular disease.

Right ventricle free wall mechanics in metabolic syndrome without type-2 diabetes: effects of a 3-month lifestyle intervention program.

BACKGROUND: Growing evidence demonstrates subtle left ventricular myocardial dysfunction in patients with metabolic syndrome (MetS), with central obesity, glucose intolerance and inflammation emerging as important contributors. Whether these results can be translated to the right ventricle (RV) is not yet fully elucidated. Furthermore, although lifestyle intervention favorably impacts MetS components and inflammatory biomarkers, its effect on RV myocardial function remains unknown today. METHODS: Thirty-nine MetS adults free of diabetes were enrolled in a three month lifestyle intervention program including diet and physical exercise, and compared with forty healthy controls. Blood biochemistry, echocardiography including tissue Doppler imaging (TDI), and vector velocity imaging of the RV free wall to assess global longitudinal strain (GLS) and strain rates (SR) were obtained at baseline and after the intervention. RESULTS: Compared with controls, MetS patients presented similar right atrial and RV morphology but reduced systolic (P = 0.04) and early diastolic (P = 0.02) velocities of the tricuspid annulus. They showed attenuated RV GLS (-21.4 ± 4.5 vs -25.7 ± 4.9%, P < 0.001) as well as early diastolic (P = 0.003) and systolic (P < 0.001) SR. Multiple regression analyses revealed log PAI-1 active, (P < 0.001), log adiponectin, (P = 0.01), LV mass indexed (P = 0.004) and central fat (P = 0.03) as independent predictors of RV GLS (R2 = 0.46, P < 0.001). Biological markers of MetS and inflammation as well as RV GLS (-21.8 ± 3.8 vs -24.3 ± 3.0%, P = 0.009) and systolic (P = 0.003) and early diastolic (P = 0.01) SR, but not TDI indexes, significantly improved after diet and exercise training, and vector velocity imaging data in MetS following the lifestyle intervention no longer differed from controls. CONCLUSIONS: MetS is associated with subtle impairments in both RV free wall diastolic and systolic myocardial function which could be partly related to central-obesity induced changes in pro- and anti-inflammatory cytokines and left ventricular remodeling. The favorable impact of healthy dieting and physical activity on RV free wall mechanics indicates that cellular and sub-cellular alterations responsible for the RV myocardial abnormalities are probably not permanent and modifiable throughout adequate interventional strategies. TRIAL REGISTRATION: American National Institutes of Health database NCT00917917.