Liraglutide and not lifestyle intervention reduces soluble CD163 after comparable weight loss in obese participants with prediabetes or type 2 diabetes mellitus.

BACKGROUND: The GLP-1 receptor agonist liraglutide is used to treat hyperglycemia in type 2 diabetes but is also known to induce weight loss, preserve the beta cell and reduce cardiovascular risk. The mechanisms underlying these effects are however still not completely known. Herein we explore the effect of liraglutide on markers of immune cell activity in a population of obese individuals with prediabetes or newly diagnosed type 2 diabetes mellitus. METHOD: Plasma levels of the monocyte/macrophage markers, soluble (s)CD163 and sCD14, the neutrophil markers myeloperoxidase (MPO) and neutrophil gelatinase-associated lipocalin (NGAL),the T-cell markers sCD25 and T-cell immunoglobulin mucin domain-3 (sTIM-3) and the inflammatory marker TNF superfamily (TNFSF) member 14 (LIGHT/TNFSF14) were measured by enzyme-linked immunosorbent assays in obese individuals with prediabetes or diabetes diagnosed within the last 12 months, prior to and after comparable weight loss achieved with lifestyle changes (n = 20) or liraglutide treatment (n = 20), and in healthy subjects (n = 13). RESULTS: At baseline, plasma levels of the macrophage marker sCD163, and the inflammatory marker LIGHT were higher in cases as compared to controls. Plasma levels of sCD14, NGAL, sTIM-3 and sCD25 did not differ at baseline between patients and controls. After weight reduction following lifestyle intervention or liraglutide treatment, sCD163 decreased significantly in the liraglutide group vs. lifestyle (between-group difference p = 0.023, adjusted for visceral adipose tissue and triglycerides basal values). MPO and LIGHT decreased significantly only in the liraglutide group (between group difference not significant). Plasma levels of MPO and in particular sCD163 correlated with markers of metabolic dysfunction and inflammation. After weight loss, only sCD163 showed a trend for decreased levels during OGTT, both in the whole cohort as in those of liraglutide vs lifestyle group. CONCLUSION: Weight loss following treatment with liraglutide was associated with reduced circulating levels of sCD163 when compared to the same extent of weight loss after lifestyle changes. This might contribute to reduced cardiometabolic risk in individuals receiving treatment with liraglutide.

Kinetics of Circulating Extracellular Vesicles Over the 24-Hour Dosing Interval After Low-Dose Aspirin Administration in Patients at Cardiovascular Risk.

Extracellular vesicles (EVs) are small vesicles deriving from all cell types during cell activation, involved in transcellular communication, and regarded as predictors of vascular damage and of cardiovascular events. We tested the hypothesis that, in patients on chronic low-dose aspirin treatment for cardiovascular prevention, aspirin may affect the release of EVs within the 24-hour interval. We enrolled 84 patients, mostly at high or very high cardiovascular risk, on chronic low-dose aspirin treatment. The numbers of circulating EVs (cEVs) and annexinV+ cEVs (total, platelet-derived, endothelial-derived, and leucocyte-derived) were assessed immediately before, and after 10 and 24 hours of a witnessed aspirin administration. Platelet cyclooxygenase 1 (COX-1) recovery was characterized by measuring serum thromboxane B2 (sTXB2 ) at the same timepoints. Nine healthy participants were also enrolled. In patients, daily aspirin administration acutely inhibited after 10 hours following aspirin administrations the release of cEVs (total and leukocyte-derived) and annexinV+ cEVs (total, platelet-derived, endothelial-derived, and leukocyte-derived), with a rapid recovery at 24 hours. The inhibition after 10 hours suggests a COX-1-dependent mechanism. Interestingly, the slope of platelet-derived and of annexinV+ platelet-derived cEVs were both directly related to sTXB2 slope and COX-1 messenger RNA, raising the hypothesis that vice versa, cEVs may affect the rate of COX-1 recovery and the subsequent duration of aspirin effect. In healthy participants, no circadian difference was observed, except for leukocyte-derived cEVs. Our findings suggest a previously unappreciated effect of aspirin on the kinetics of a subset of cEVs possibly contributing to the cardioprotective effects of this drug.

Reduced platelet glycoprotein Ibα shedding accelerates thrombopoiesis and COX-1 recovery: implications for aspirin dosing regimen.

Cardiovascular (CV) disease prevention with low-dose aspirin can be less effective in patients with a faster recovery of platelet (PLT) cyclooxygenase (COX)-1 activity during the 24-hour dosing interval. We previously showed that incomplete suppression of TXA2 over 24 hours can be rescued by a twice daily aspirin regimen. Here we show that reduced PLT glycoprotein (GP)Ibα shedding characterizes patients with accelerated COX-1 recovery and may contribute to higher thrombopoietin (TPO) production and higher rates of newly formed PLT, escaping aspirin inhibition over 24 hours. Two hundred aspirin-treated patients with high CV risk (100 with type 2 diabetes mellitus) were stratified according to the kinetics of PLT COX-1 activity recovery during the 10- to 24-hour dosing interval. Whole proteome analysis showed that PLT from patients with accelerated COX-1 recovery were enriched in proteins involved in cell survival, inhibition of apoptosis and cellular protrusion formation. In agreement, we documented increased plasma TPO, megakaryocyte maturation and proplatelet formation, and conversely increased PLT galactose and reduced caspase 3, phosphatidylserine exposure and ADAM17 activation, translating into diminished GPIbα cleavage and glycocalicin (GC) release. Treatment of HepG2 cells with recombinant GC led to a dose-dependent reduction of TPO mRNA in the liver, suggesting that reduced GPIbα ectodomain shedding may unleash thrombopoiesis. A cluster of clinical markers, including younger age, non-alcoholic fatty liver disease, visceral obesity and higher TPO/GC ratio, predicted with significant accuracy the likelihood of faster COX-1 recovery and suboptimal aspirin response. Circulating TPO/GC ratio, reflecting a dysregulation of PLT lifespan and production, may provide a simple tool to identify patients amenable to more frequent aspirin daily dosing.

Study of pulsed electrostatic field (PESF) in the perfusion of peripheral tissues: Microangiopathy, nutrition and quality of perceiver life.

Microangiopathy compromises the structural and functional integrity of organs and tissues in patients with type II diabetes mellitus (T2DM) negatively affecting the perceived quality of life. Nitric oxide (NO) is a multifunctional signalling molecule, acting as a vasodilator, neurotransmitter, and modulator of inflammatory processes. Patients with type II diabetes mellitus and chronic kidney disease, controlled from glycaemic status, were treated or not with pulsed electrostatic field (PESF) cycles to evaluate effect on the perfusion of peripheral tissues. Everyone was monitored for the metabolic profile, and we tested circulating NO with a commercial enzyme immunoassay kit. In addition, we tested the perceived quality of life of patients before/after a PESF cycle using a questionnaire. Patients treated with PESF were improved circulating NO levels, significant changes in systolic and diastolic blood pressure, heart rate and were more homogeneous for their metabolic profile. The questionnaire showed also a marked improvement in the perceived quality of life. The use of pulsed electrostatic fields has allowed us to observe an improvement in the metabolic, psychological, and clinical profile in patients with T2DM and chronic kidney disease whose pathological profile is strongly compromised.

Effects of liraglutide vs. lifestyle changes on soluble suppression of tumorigenesis-2 (sST2) and galectin-3 in obese subjects with prediabetes or type 2 diabetes after comparable weight loss.

BACKGROUND: Soluble suppression of tumorigenesis-2 (sST2) and galectin (Gal)-3 are two biomarkers related to inflammation, metabolic disturbances and to myocardial fibrosis that characterize several cardiac pathological conditions. Increased circulating levels of these molecules have been associated with risk of cardiovascular death. Treatment with liraglutide, a glucagon-like peptide 1 analog, is associated with weight loss, improved glycemic control, and reduced cardiovascular risk. We wanted to assess (I) potential differences between subjects with prediabetes or type 2 diabetes mellitus (T2DM) and healthy controls in sST2 and Gal-3 circulating levels, and their relationship with glycemic control and markers of beta cell function and myocardial injury; (II) whether liraglutide treatment modulates these markers in subjects with prediabetes or early T2DM independently of weight loss; (III) whether baseline levels of any of these two molecules may predict the response to liraglutide treatment. METHODS: Forty metformin-treated obese subjects (BMI ≥ 30) with prediabetes [impaired fasting glucose (IFG) or impaired glucose tolerance (IGT) or both (n = 23)] or newly diagnosed T2DM (n = 17), were randomized to liraglutide or lifestyle counseling until achieving a comparable weight loss (7% of initial body weight). Thirteen subjects were enrolled as healthy controls for baseline sST2 and Gal-3 levels. RESULTS: Baseline sST2 levels were comparable between controls and obese patients (p = 0.79) whereas Gal-3 levels were significantly higher in patients as compared to controls (p < 0.001). Liraglutide treatment, but not weight loss achieved by lifestyle counseling, decreased plasma sST2 levels (- 9%, beta = - 14.9, standard deviation 6.9, p = 0.037) while Gal-3 levels did not change. A reduction in serum hs-Troponin I was observed after intervention, due to a 19% (p = 0.29) increase in the lifestyle arm, and a 25% decrease (p = 0.033) in the liraglutide arm (between-group difference p = 0.083). Lower baseline Gal-3 levels predicted a better improvement in beta cell function after liraglutide treatment. CONCLUSIONS: Liraglutide-induced reduction in sST2 and possibly hs-TnI suggests that in obese patients with prediabetes or early T2DM this drug may have a positive effect on (cardiac) fibrosis, whereas plasma level of Gal-3 before liraglutide initiation may predict response to the drug in terms of beta cell function improvement. Trial registration Eudract: 2013-001356-36.

Insulin resistance and NAFLD may influence memory performance in obese patients with prediabetes or newly-diagnosed type 2 diabetes.

BACKGROUND AND AIMS: Diabetes has consistently been shown to increase risk for cognitive decline. Cognitive deficits may occur at the very earliest stages of diabetes. We sought to estimate the determinants of memory function in a group of middle-aged obese subjects with prediabetes or newly-diagnosed type 2 diabetes mellitus. METHODS AND RESULTS: Sixty-two obese patients in treatment with metformin-with prediabetes (n = 41) or newly diagnosed T2DM (n = 21), were studied. Short- and long-term memory function was assessed through a neuropsychological assessment consisting of two tests and a composite domain z score was calculated. Cardiometabolic variables, such as abdominal MRI quantification of subcutaneous (SAT) and visceral (VAT) adipose tissue content, and of intra-hepatocellular lipid content, as well as insulin sensitivity (Matsuda Index, HOMA-IR) and beta cell performance (Beta Index), by multiple sampling, 8-point oral glucose tolerance test, were also evaluated. Age, non-alcoholic fatty liver disease (NAFLD), and lnHOMA-IR together explained 18% (R square) of the variance in memory domain. Including NAFLD increased the explained variance by 8% and including lnHOMA-IR by 9.1%, whereas the contribution of age and other factors was negligible. CONCLUSION: Preventing and managing insulin resistance in precocious and possibly earlier stages of diabetes might provide benefit in slowering down future cognitive decline.

Endogenous PCSK9 may influence circulating CD45neg/CD34bright and CD45neg/CD34bright/CD146neg cells in patients with type 2 diabetes mellitus.

Protease proprotein convertase subtilisin/kexin type 9 (PCSK9) is a regulator of LDL cholesterol clearance and has been associated with cardiovascular risk. PCSK9 inhibitors increase in vivo circulating endothelial progenitor cells (EPCs), a subtype of immature cells involved in ongoing endothelial repair. We hypothesized that the effect of PCSK9 on vascular homeostasis may be mediated by EPCs in patients with or without type 2 diabetes mellitus (T2DM). Eighty-two patients (45 with, 37 without T2DM) at high cardiovascular risk were enrolled in this observational study. Statin treatment was associated with higher circulating levels of PCSK9 in patients with and without T2DM (p < 0.001 and p = 0.036) and with reduced CD45neg/CD34bright (total EPC compartment) (p = 0.016) and CD45neg/CD34bright/CD146neg (early EPC) (p = 0.040) only among patients with T2DM. In the whole group of patients, statin treatment was the only independent predictor of low number of CD45neg/CD34bright (ß = - 0.230; p = 0.038, adjusted R2 = 0.041). Among T2DM patients, PCSK9 circulating levels were inversely related and predicted both the number of CD45neg/CD34bright (ß = - 0.438; p = 0.003, adjusted R2 = 0.173), and CD45neg/CD34bright/CD146neg (ß = - 0.458; p = 0.002, adjusted R2 = 0.191) independently of age, gender, BMI and statin treatment. In high-risk T2DM patients, high endogenous levels of PCSK9 may have a detrimental effect on EPCs by reducing the endothelial repair and worsening the progression of atherothrombosis.

Sex-Specific Association of Endogenous PCSK9 With Memory Function in Elderly Subjects at High Cardiovascular Risk.

Background: Growing evidence indicates that cognitive decline and cardiovascular diseases (CVDs) share common vascular risk factors. Protease proprotein convertase subtilisin/kexin type 9 (PCSK9) is associated with CV disease risk and has been also involved in neuronal differentiation. Aim: Evaluate whether in patients at high CV risk cognitive function is related to PCSK9 levels. Methods. One hundred sixty-six patients (67 female) were enrolled. A detailed neuropsychological (NP) assessment was performed. PCSK9 levels were measured with ELISA. Results: Men had significantly higher short-term memory, executive function, and praxic and mental representation skills, as reflected by Forward Digit Span (FDS) (p = 0.005), Trail Making Test-A (TMT-A) (p = 0.047), Clock Drawing Test (CDT) (0.016). Endogenous PCSK9 levels were higher in female (p = 0.005). On linear regression analysis PCSK9 predicts short term memory only in females (Beta = 0.408, p = 0.001), with an interaction between PCSK9 and gender (p = 0.004 for interaction PCSK9 by sex). The association of PCSK9 with FDS in female was partially mediated by waist circumference (mediation effect 8.5%). Conclusions: In patients at high CV risk short term memory was directly related to PCSK9 levels only in women, revealing the relevance of sex in this relationship. The association of PCSK9 with memory function may be mediated, at least in part, by waist circumference.

Liraglutide improves memory in obese patients with prediabetes or early type 2 diabetes: a randomized, controlled study.

BACKGROUND/OBJECTIVES: Diabetic subjects are at increased risk of subtle cognitive impairment since the disease early stages and of dementia later in life. In animal models, glucagon-like peptide-1 receptor agonizts (GLP1-RAs) have been shown to exert neuroprotective effects, expecially in the memory domain. We assessed whether treatment with a GLP1-RA might affect cognitive functions in type 2 diabetic subjects independently on the weight loss it might induce. SUBJECTS/METHODS: Forty metformin-treated obese subjects with prediabetes or newly diagnosed type 2 diabetes mellitus, received liraglutide (1.8 mg/d) (n = 20) or lifestyle counseling (dietary intervention and exercise training) (n = 20) until achieving a modest and comparable weight loss (-7% of initial body weight). INTERVENTIONS/METHODS: A detailed neuropsychological assessment before and after weight loss was completed in 16 patients per arm, who were administered a total of seven psychological tests, thus assessing three composite domain z-scores for attention, memory, and executive control. RESULTS: After comparable weight loss and superimposable glycemic control and insulin sensitivity, a significant increase in short term memory (mean Digit Span Z score from -0.06 to 0.80, p = 0.024) and memory composite z-score (mean memory z-score from -0.67 to 0.032, p = 0.0065) was observed in the liraglutide exposed subjects (between group p = 0.041 and p = 0.033, respectively). CONCLUSIONS: Liraglutide might slow down memory function decline in diabetic patients in early, and possibly preclinical stages of the disease.

In vivo thromboxane-dependent platelet activation is persistently enhanced in subjects with impaired glucose tolerance.

BACKGROUND: Impaired glucose tolerance (IGT) is associated with increased cardiovascular morbidity and mortality. Enhanced thromboxane (TX)-dependent platelet activation plays a pivotal role in atherothrombosis and characterizes type 2 diabetes mellitus (DM). Whether this also pertains to IGT is currently unknown. We investigated whether TXA2 -dependent platelet activation, as reflected by 11-dehydro-TXB2 (TXM) urinary excretion, is comparably abnormal in IGT as in DM, is persistent over long-term follow-up, changes as a function of metabolic disease progression, and is influenced by food intake. METHODS: We prospectively investigated subjects with IGT (n = 48) and two control groups with DM diagnosed either less than 12 months (n = 60) or 12 months or more (n = 58). RESULTS: Baseline TXM excretion was comparable between subjects with IGT and DM, with no evidence of a circadian variation. During a 36-month follow-up, urinary TXM excretion was stable over time in the DM groups, while tended to increase in subjects with IGT. Increasing urinary TXM excretion over time was observed in the subjects who progressed to diabetes vs nonprogressors. CONCLUSIONS: We conclude that TXA2 -dependent platelet activation was at least as high in IGT as in patients with DM and further increased over time, especially in those who progressed to overt diabetes.

Thromboxane-Dependent Platelet Activation in Obese Subjects with Prediabetes or Early Type 2 Diabetes: Effects of Liraglutide- or Lifestyle Changes-Induced Weight Loss.

Thromboxane (TX)-dependent platelet activation and lipid peroxidation, as reflected in vivo by the urinary excretion of 11-dehydro-TXB2 and 8-iso-prostaglandin (PG)F2α, play a key role in atherothrombosis in obesity and type 2 diabetes mellitus (T2DM) since the earlier stages. Thirty-five metformin-treated obese subjects with prediabetes or newly-diagnosed T2DM were randomized to the glucagon-like peptide receptor agonist (GLP-RA) liraglutide (1.8 mg/day) or lifestyle counseling until achieving a comparable weight loss (-7% of initial body weight), to assess whether changes in subcutaneous (SAT) and visceral (VAT) adipose tissue distribution (MRI), insulin sensitivity (Matsuda Index) and beta-cell performance (multiple sampling OGTT beta-index), with either intervention, might affect TX-dependent platelet activation, lipid peroxidation and inflammation. At baseline, Ln-8-iso-PGF2α (Beta = 0.31, p = 0.0088), glycosylated hemoglobin (HbA1c) (Beta = 2.64, p = 0.0011) Ln-TNF-α (Beta = 0.58, p = 0.0075) and SAT (Beta = 0.14, p = 0.044) were significant independent predictors of 11-dehydro-TXB2. After achievement of the weight loss target, a comparable reduction in U-11-dehydro-TXB2 (between-group p = 0.679) and 8-iso-PGF-2α (p = 0.985) was observed in both arms in parallel with a comparable improvement in glycemic control, insulin sensitivity, SAT, high-sensitivity C-reactive protein (hs-CRP). In obese patients with initial impairment of glucose metabolism, the extent of platelet activation is related to systemic inflammation, isoprostane formation and degree of glycemic control and abdominal SAT. Successful weight loss, achieved with either lifestyle changes or an incretin-based therapy, is associated with a significant reduction in lipid peroxidation and platelet activation.

Significance of urinary 11-dehydro-thromboxane B2 in age-related diseases: Focus on atherothrombosis.

Platelet activation plays a key role in atherogenesis and atherothrombosis. Biochemical evidence of increased platelet activation in vivo can be reliably obtained through non-invasive measurement of thromboxane metabolite (TXM) excretion. Persistent biosynthesis of TXA2 has been associated with several ageing-related diseases, including acute and chronic cardio-cerebrovascular diseases and cardiovascular risk factors, such as cigarette smoking, type 1 and type 2 diabetes mellitus, obesity, hypercholesterolemia, hyperhomocysteinemia, hypertension, chronic kidney disease, chronic inflammatory diseases. Given the systemic nature of TX excretion, involving predominantly platelet but also extraplatelet sources, urinary TXM may reflect either platelet cyclooxygenase-1 (COX-1)-dependent TX generation or COX-2-dependent biosynthesis by inflammatory cells and/or platelets, or a combination of the two, especially in clinical settings characterized by low-grade inflammation or enhanced platelet turnover. Although urinary 11-dehydro-TXB2 levels are largely suppressed with low-dose aspirin, incomplete TXM suppression by aspirin predicts the future risk of vascular events and death in high-risk patients and may identify individuals who might benefit from treatments that more effectively block in vivo TX production or activity. Several disease-modifying agents, including lifestyle intervention, antidiabetic drugs and antiplatelet agents besides aspirin have been shown to reduce TX biosynthesis. Taken together, these aspects may contribute to the development of promising mechanism-based therapeutic strategies to reduce the progression of atherothrombosis. We intended to critically review current knowledge on both the pathophysiological significance of urinary TXM excretion in clinical settings related to ageing and atherothrombosis, as well as its prognostic value as a biomarker of vascular events.

Oxidant stress as a major determinant of platelet activation in invasive breast cancer.

The hypothesis that increased oxidative stress in breast cancer (BC) patients could induce enhanced lipid peroxidation, which, in turn, would contribute to platelet activation and poor clinical outcome is attractive. To address this issue, we investigated pre-surgical urinary 8-iso-prostaglandin (PG)F2α (marker of in vivo oxidative stress) and 11-dehydro-thromboxane (TX)B2 (marker of in vivo platelet activation) levels in patients with primary BC (n = 115) compared with control women paired for comorbidities and their association with patients' metabolic profile and clinical prognostic factors. The results obtained showed that presurgical urinary excretion of both biomarkers was enhanced in BC patients compared to controls and was associated with patients' estrogen receptor (ER) expression, but not HER2 status or Ki67 proliferation index. Accordingly, both urinary biomarkers were increased in patients with luminal-like BC molecular subtypes compared with triple negative or HER2-enriched tumors. ER status was an independent predictor of 8-iso-PGF2α urinary levels, which, in turn, significantly predicted 11-dehydro-TXB2 urinary levels together with disease stage and ER status. The prognostic value of 11-dehydro-TXB2 was then evaluated showing a significant correlation with BC pathological response to neoadjuvant treatment. Furthermore, among relapsing patients, those with elevated urinary biomarker levels were more likely to develop distant metastasis rather than local recurrence. In conclusion, we may speculate that enhanced oxidative stress due to estrogen-related mechanisms might cause a condition of persistent platelet activation capable of sustaining BC growth and progression through the release of bioactive stored molecules, ultimately contributing to tumor invasiveness. Further studies specifically addressing this hypothesis are presently needed.

Increased circulating resistin is associated with insulin resistance, oxidative stress and platelet activation in type 2 diabetes mellitus.

Resistin is an adipokine that promotes inflammation and insulin resistance by targeting several cells including platelets. We hypothesised that in type 2 diabetes (T2DM), resistin may foster in vivo oxidative stress, thromboxane-dependent platelet activation and platelet-derived inflammatory proteins release, key determinants of atherothrombosis. A cross-sectional comparison of circulating resistin, sCD40L, as a marker of platelet-mediated inflammation, asymmetric dimethylarginine (ADMA), endothelial dysfunction marker, Dickkopf (DKK)-1, reflecting the inflammatory interaction between platelets and endothelial cells, and urinary 8-iso-PGF2α and 11-dehydro-TxB2, reflecting in vivo lipid peroxidation and platelet activation, respectively, was performed between 79 T2DM patients and 30 healthy subjects. Furthermore, we investigated the effects of the α-glucosidase inhibitor acarbose and the PPARγ agonist rosiglitazone, targeting hyperglycaemia or insulin resistance, versus placebo, in 28 and 18 T2DM subjects, respectively. Age- and gender-adjusted serum resistin levels were significantly higher in patients than in controls. HOMA (ß=0.266, p=0.017) and 11-dehydro-TXB2 (ß=0.354, p=0.002) independently predicted resistin levels. A 20-week treatment with acarbose was associated with significant reductions (p=0.001) in serum resistin, DKK-1, urinary 11-dehydro-TXB2 and 8-iso-PGF2α with direct correlations between the change in serum resistin and in other variables. A 24-week rosiglitazone treatment on top of metformin was associated with significant decreases in resistin, DKK-1, 11-dehydro-TXB2 and 8-iso-PGF2α, in parallel with HOMA decrease. In conclusion, resistin, antagonising insulin action in part through PPARγ activation, may favour insulin resistance and enhance oxidative stress, endothelial dysfunction and platelet activation. The adipokine-platelet interactions may be involved in platelet insulin resistance and their consequent pro-aggregatory phenotype in this setting.

LIGHT/TNFSF14 is increased in patients with type 2 diabetes mellitus and promotes islet cell dysfunction and endothelial cell inflammation in vitro.

AIMS/HYPOTHESIS: Activation of inflammatory pathways is involved in the pathogenesis of type 2 diabetes mellitus. On the basis of its role in vascular inflammation and in metabolic disorders, we hypothesised that the TNF superfamily (TNFSF) member 14 (LIGHT/TNFSF14) could be involved in the pathogenesis of type 2 diabetes mellitus. METHODS: Plasma levels of LIGHT were measured in two cohorts of type 2 diabetes mellitus patients (191 Italian and 40 Norwegian). Human pancreatic islet cells and arterial endothelial cells were used to explore regulation and relevant effects of LIGHT in vitro. RESULTS: Our major findings were: (1) in both diabetic cohorts, plasma levels of LIGHT were significantly raised compared with sex- and age-matched healthy controls (n = 32); (2) enhanced release from activated platelets seems to be an important contributor to the raised LIGHT levels in type 2 diabetes mellitus; (3) in human pancreatic islet cells, inflammatory cytokines increased the release of LIGHT and upregulated mRNA and protein levels of the LIGHT receptors lymphotoxin ß receptor (LTßR) and TNF receptor superfamily member 14 (HVEM/TNFRSF14); (4) in these cells, LIGHT attenuated the insulin release in response to high glucose at least partly via pro-apoptotic effects; and (5) in human arterial endothelial cells, glucose boosted inflammatory response to LIGHT, accompanied by an upregulation of mRNA levels of HVEM (also known as TNFRSF14) and LTßR (also known as LTBR). CONCLUSIONS/INTERPRETATION: Our findings show that patients with type 2 diabetes mellitus are characterised by increased plasma LIGHT levels. Our in vitro findings suggest that LIGHT may contribute to the progression of type 2 diabetes mellitus by attenuating insulin secretion in pancreatic islet cells and by contributing to vascular inflammation.

Oxidative stress and thromboxane-dependent platelet activation in inflammatory bowel disease: effects of anti-TNF-α treatment.

Patients with inflammatory bowel disease (IBD) are at higher risk of venous thromboembolism and coronary artery disease despite having a lower burden of traditional risk factors. Platelets from IBD patients release more soluble CD40 ligand (CD40L), and this has been implicated in IBD platelet hyper-activation. We here measured the urinary F2-isoprostane 8-iso-prostaglandin (PG)2α (8-iso-PGF2α), urinary 11-dehydro-thromboxane (TX) B2 (11-dehydro-TXB2) and plasma CD40L in IBD patients, and explored the in vitro action of anti-tumour necrosis factor (TNF)-α antibody infliximab on IBD differentiating megakaryocytes. Urinary and blood samples were collected from 124 IBD patients and 37 healthy subjects. Thirteen IBD patients were also evaluated before and after 6-week infliximab treatment. The in vitro effect of infliximab on patient-derived megakaryocytes was evaluated by immunoflorescence microscopy and by flow cytometry. IBD patients had significantly (p<0.0001) higher urinary 8-iso-PGF2α and 11-dehydro-TXB2 as well as plasma CD40L levels than controls, with active IBD patients displaying higher urinary and plasma values when compared to inactive patients in remission. A 6-week treatment with infliximab was associated with a significant reduction of the urinary excretion of 8-iso-PGF2α and 11-dehydro-TXB2 (p=0.008) and plasma CD40L (p=0.001). Infliximab induced significantly rescued pro-platelet formation by megakaryocytes derived from IBD patients but not from healthy controls. Our findings provide evidence for enhanced in vivo TX-dependent platelet activation and lipid peroxidation in IBD patients. Anti-TNF-α therapy with infliximab down-regulates in vivo isoprostane generation and TX biosynthesis in responder IBD patients. Further studies are needed to clarify the implication of infliximab induced-proplatelet formation from IBD megakaryocytes.

Pentraxin 3 and Platelet Activation in Obese Patients After Gastric Banding.

BACKGROUND: Circulating pentraxin 3 (PTX3), the main regulator of the inflammatory response, rapidly increases following cardiovascular events, and low PTX3 is associated with high body mass index. METHODS AND RESULTS: We conducted a 12-month longitudinal study, to test the hypothesis that laparoscopic adjustable gastric banding (LAGB)-induced weight loss was associated with changes in platelet activation markers and PTX3. Twelve obese patients, scheduled to undergo LAGB, were enrolled at the University Obesity Center. Urinary 11-dehydro-thromboxane (Tx)B2excretion rate was measured on radioimmunoassay, and PTX3 and CD40L were determined on immunoassay. Plasma PTX3 increased by 178.8 and 214.9% (P<0.0001), respectively, 6 and 12 months after LAGB. High-sensitivity CRP decreased by 24 and 29.7% (P<0.0001), whereas CD40L decreased by 64.3 and 58.6% (P=0.002), respectively. Urinary 11-dehydro-TxB2decreased from 1,443 to 715 and 564 pg/mg creatinine, respectively 6 months and 12 months after LAGB (P<0.0001). PTX3 was inversely related to platelet activation markers, 11-dehydro-TxB2and CD40L. Moreover, multiple regression analysis on pooled data showed that plasma PTX3 was an independent predictor of urinary 11-dehydro-TxB2. CONCLUSIONS: There is an association between inflammation, platelet activation and metabolic dysfunction in obesity, and PTX3 is a key player within these circuits.

Arterial stiffness and sedentary lifestyle: Role of oxidative stress.

Sedentary lifestyle is a risk factor for the development of cardiovascular disease, and leads to a quantifiable impairment in vascular function and arterial wall stiffening. We tested the hypothesis of oxidative stress as a determinant of arterial stiffness (AS) in physically inactive subjects, and challenged the reversibility of these processes after the completion of an eight-week, high-intensity exercise training (ET). AS was assessed before and after ET, measuring carotid to femoral pulse wave velocity (PWV) with a Vicorder device. At baseline and after ET, participants performed urine collection and underwent fasting blood sampling. Urinary 8-iso-PGF2α, an in vivo marker of lipid peroxidation, total, HDL and LDL cholesterol, and triglyceride concentrations were measured. ET was associated with significantly reduced urinary 8-iso-PGF2α(p<0.0001) levels. PWV was significantly reduced after ET completion (p<0.0001), and was directly related to urinary 8-iso-PGF2α(Rho=0.383, p=0.021). After ET, cardiovascular fitness improved [peak oxygen consumption (p<0.0001), peak heart rate (p<0.0001)]. However, no improvement in lipid profile was observed, apart from a significant reduction of triglycerides (p=0.022). PWV and triglycerides were significantly related (Rho=0.466, p=0.005) throughout the study period. PWV levels were also related to urinary 8-iso-PGF2α in our previously sedentary subjects. We conclude that regular physical exercise may be a natural antioxidant strategy, lowering oxidant stress and thereby the AS degree.

Platelets and diabetes mellitus.

Platelet activation plays a key role in atherothrombosis in type 2 diabetes mellitus (T2DM) and increased in vivo platelet activation with enhanced thromboxane (TX) biosynthesis has been reported in patients with impairment of glucose metabolism even in the earlier stages of disease and in the preclinical phases. In this regards, platelets appear as addresses and players carrying and transducing metabolic derangement into vascular injury. The present review critically addresses key pathophysiological aspects including (i) hyperglycemia, glycemic variability and insulin resistance as determinants and predictors of platelet activation, (ii) inflammatory mediators derived from platelets, such as soluble CD40 ligand, soluble CD36, Dickkopf-1 and probably soluble receptor for advanced glycation-end-products (sRAGE), which expand the functional repertoire of platelets from players of hemostasis and thrombosis to powerful amplifiers of inflammation by promoting the release of cytokines and chemokines, cell activation, and cell-cell interactions; (iii) molecular mechanisms underpinning the less-than-expected antithrombotic protection by aspirin (ASA), despite regular antiplatelet prophylaxis at the standard dosing regimen, and (iv) stratification of patients deserving different antiplatelet strategies, based on the metabolic phenotype. Taken together, these pathophysiological aspects may contribute to the development of promising mechanism-based therapeutic strategies to reduce the progression of atherothrombosis in diabetic subjects.

Circulating myeloid-related protein-8/14 is related to thromboxane-dependent platelet activation in patients with acute coronary syndrome, with and without ongoing low-dose aspirin treatment.

BACKGROUND: Platelet activation is involved in acute coronary syndromes (ACS). Incomplete suppression by low-dose aspirin treatment of thromboxane (TX) metabolite excretion (urinary 11-dehydro-TXB2) is predictive of vascular events in high-risk patients. Myeloid-related protein (MRP)-8/14 is a heterodimer secreted on activation of platelets, monocytes, and neutrophils, regulating inflammation and predicting cardiovascular events. Among platelet transcripts, MRP-14 has emerged as a powerful predictor of ACS. METHODS AND RESULTS: We enrolled 68 stable ischemic heart disease (IHD) and 63 ACS patients, undergoing coronary angiography, to evaluate whether MRP-8/14 release in the circulation is related to TX-dependent platelet activation in ACS and IHD patients and to residual TX biosynthesis in low-dose aspirin-treated ACS patients. In ACS patients, plasma MRP-8/14 and urinary 11-dehydro-TXB2 levels were linearly correlated (r=0.651, P<0.001) but significantly higher than those in IHD patients (P=0.012, P=0.044) only among subjects not receiving aspirin. In aspirin-treated ACS patients, MRP-8/14 and 11-dehydro-TXB2 were lower versus those not receiving aspirin (P<0.001) and still significantly correlated (r=0.528, P<0.001). Higher 11-dehydro-TXB2 significantly predicted higher MRP-8/14 in both all ACS patients and ACS receiving aspirin (P<0.001, adj R(2)=0.463 and adj R(2)=0.497) after multivariable adjustment. Conversely, plasma MRP-8/14 (P<0.001) and higher urinary 8-iso-prostaglandin F2α (P=0.050) levels were significant predictors of residual, on-aspirin, TX biosynthesis in ACS (adjusted R(2)=0.384). CONCLUSIONS: Circulating MRP-8/14 is associated with TX-dependent platelet activation in ACS, even during low-dose aspirin treatment, suggesting a contribution of residual TX to MRP-8/14 shedding, which may further amplify platelet activation. Circulating MRP-8/14 may be a target to test different antiplatelet strategies in ACS.