Association of vitronectin and plasminogen activator inhibitor-1 levels with the risk of metabolic syndrome and type 2 diabetes mellitus. Results from the D.E.S.I.R. prospective cohort.

It was the objective of this study to investigate the relation between vitronectin and plasminogen activator inhibitor (PAI)-1 plasma levels with nine-year incidences of the metabolic syndrome (MetS) and of type 2 diabetes mellitus (T2DM). Baseline plasma concentrations of vitronectin and PAI-1 were measured in 627 healthy participants from the prospective D.E.S.I.R. cohort who subsequently developed MetS (n = 487) and T2DM (n = 182) over a nine-year follow-up (42 presented both) and who were matched with two healthy control subjects each by use of a nested case-control design. Parameters composing the MetS explained about 20% of plasma vitronectin levels. An increase of one standard deviation of vitronectin was associated with increased risks of both the MetS (odds ratio [OR] = 1.21 [1.07 - 1.37], p = 0.003) and T2DM (OR = 1.24 [1.01 - 1.53], p = 0.045). Corresponding ORs for PAI-1 levels were 1.46 [1.27 - 1.68] (p<10(-4)) and 1.40 [1.14 - 1.72] (p = 0.0012). However, the effects of vitronectin and PAI-1 levels on outcomes were not independent. The vitronectin-MetS association was restricted to individuals with low to modest PAI-1 levels (OR = 1.33 [1.14 - 1.54], p = 0.0003) while no association was observed in individuals with high PAI-1 levels (OR = 0.87 [0.68 - 1.10], p = 0.24), the test for interaction being highly significant (p = 0.0009). In conclusion, baseline plasma vitronectin is a marker of incident MetS at nine years. Its predictive ability for MetS and T2DM should not be assessed independently of PAI-1 levels.

Effect of diet-induced weight loss on endothelial dysfunction: early improvement after the first week of dieting.

Obesity is associated with impaired endothelial function, and this may lead to increased cardiovascular risk. To gain insight into the beneficial effects of diet-induced weight loss on endothelial function, endothelium-dependent, flow-mediated dilation (FMD) of the brachial artery and several metabolic and inflammatory markers were assessed in 40 obese women (BMI 34.9 ± 4.88 kg/m(2)) at baseline, after the 1st week and after 5 months on a low-calorie diet of 5.0 MJ/day. Twenty lean women served as controls. At entry, the obese women had a lower FMD than the lean women (7.7 ± 1.8 vs. 11.5 ± 4.2%, p < 0.001). After 1 week of the intervention and 4% reduction of BMI, FMD improved by 22% (p = 0.005), and a decrease in circulating triglycerides, insulin, leptin, tissue type plasminogen activator and its inhibitor, von Willebrand factor, C-reactive protein and tumor necrosis factor receptor 1 was observed. Improvement of FMD was associated only with a decrease in BMI (r = 0.39, p = 0.03). Twenty-two women completed the weight reduction program and reduced their BMI by 16%. FMD was further improved by 64% (to 12.4 ± 5.3%, p = 0.001) and became comparable to that of lean women. None of the significant changes in the observed parameters was associated with improvement of FMD at the end of the program. Improvements in obesity-related endothelial dysfunction began in the 1st week of dieting and continued during the following months of this simple non-pharmacological lifestyle modification to reach normalisation of endothelial function. The favourable effect of dieting on endothelial function is independent of the accompanying improvement of classical risk factors.

p38 mitogen activated protein kinase controls two successive-steps during the early mesodermal commitment of embryonic stem cells.

Embryonic stem (ES) cells differentiate in vitro into all cell lineages. We previously found that the p38 mitogen activated kinase (p38MAPK) pathway controls the commitment of ES cells toward either cardiomyogenesis (p38 on) or neurogenesis (p38 off ). In this study, we show that p38α knock-out ES cells do not differentiate into cardiac, endothelial, smooth muscle, and skeletal muscle lineages. Reexpression of p38MAPK in these cells partially rescues their mesodermal differentiation defects and corrects the high level of spontaneous neurogenesis of knock-out cells. Wild-type ES cells were treated with a p38MAPK-specific inhibitor during the differentiation process. These experiments allowed us to identify 2 early independent successive p38MAPK functions in the formation of mesodermal lineages. Further, the first one correlates with the regulation of the expression of Brachyury, an essential mesodermal-specific transcription factor, by p38MAPK. In conclusion, by genetic and biochemical approaches, we demonstrate that p38MAPK activity is essential for the commitment of ES cell into cardiac, endothelial, smooth muscle, and skeletal muscle mesodermal lineages.

Polymorphisms of the lamina maturation pathway and their association with the metabolic syndrome: the DESIR prospective study.

Laminopathies are rare monogenic diseases, some of them exhibiting features of the metabolic syndrome. These diseases are mainly due to mutations in LMNA, encoding A-type lamins. One LMNA polymorphism, rs4641, has been associated with the metabolic syndrome, but results have been controversial. We therefore investigated the effect of single nucleotide polymorphisms (SNPs) in the LMNA gene in combination with four other genes encoding enzymes influencing lamin post-translational maturation on risk of metabolic syndrome (MS). Twenty-three tagging SNPs characterising the haplotypic variability of five genes (LMNA, ICMT, ZMPSTE24, FNTA and FNTB) were genotyped in 3,916 French men and women who took part in the prospective DESIR study. Single locus and haplotype analyses were performed but did not detect any significant association with the risk of MS. No robust interaction between SNPs located in different genes on the risk of MS was identified. In conclusion, we did not observe any convincing evidence that common polymorphisms of the lamina pathway could modulate the risk of MS.

Molecular characterization of a novel mutation in the factor XIII a subunit gene associated with a severe defect: importance of prophylactic substitution.

A congenital FXIII A subunit deficiency was diagnosed in a male child because of umbilical bleeding at birth. Venous infusion was difficult and prophylactic FXIII infusion was delayed. At age 1, he suffered a spontaneous intracranial haemorrhage. Substitutive FXIII was initiated, and at age 12, no other significant bleeding event had occurred. His 5 years younger brother also bears the same FXIII deficiency. The younger brother's treatment was initiated at birth and never discontinued, and no bleeding occurred. Mutation gene analysis found a homozygous four bases insertion predicting a stop codon seven residues after PRO675. Antigen assay indicated that the mutant molecule is secreted. This case highlights the importance of prophylactic FXIII infusion.

Respective contribution of conventional risk factors and antihypertensive treatment to stable angina pectoris and acute coronary syndrome as the first presentation of coronary heart disease: the PRIME Study.

OBJECTIVE: To test whether conventional risk factors and antihypertensive treatment were more predictive of stable angina (SA) than acute coronary syndrome (ACS) as the first presentation of coronary heart disease (CHD). DESIGN: We used data from the PRIME Study (Prospective Epidemiological Study of Myocardial Infarction), a prospective cohort of 9758 asymptomatic middle-aged men recruited from WHO MONICA centers in Northern Ireland and France between 1991 and 1993. SA and ACS events were registered during 5 years of follow-up. METHODS: Hazard ratios (HRs) of each risk factor measured at baseline for SA and ACS events were assessed using separate Cox proportional hazard models. Difference between HRs was estimated by the bootstrap method. RESULTS: After 5 years of follow-up, there were 114 SA and 178 ACS as the first presentation of CHD. Diastolic blood pressure [adjusted HRs for 1 standard deviation increase = 1.34; 95% confidence interval (CI): 1.17-1.54 vs. 1.04; 95% CI: 0.87-1.25; P for comparison between HRs = 0.012], and possibly cigarette smoking over or equal to 20 pack-years (adjusted HR = 2.07; 95% CI: 1.43-2.99 vs. 1.29; 95% CI: 0.83-2.01; P for comparison between HRs = 0.062) were more predictive of ACS than SA, whereas this was the opposite for antihypertensive treatment (adjusted HR = 2.18; 95% CI: 1.39-3.41 for SA vs. 1.28; 95% CI: 0.85-1.93 for ACS, P for comparison between HRs = 0.049). CONCLUSION: The present data support that SA and ACS, as the first presentation of CHD, may not share exactly the same determinants.

Platelet microparticles: a new player in malaria parasite cytoadherence to human brain endothelium.

Cerebral malaria (CM) is characterized by accumulation of circulating cells within brain microvessels, among which platelets play an important role. In vitro, platelets modulate the cytoadherence of Plasmodium falciparum-parasitized red blood cells (PRBCs) to brain endothelial cells. Here we show for the first time that platelet microparticles (PMPs) are able to bind to PRBCs, thereby transferring platelet antigens to the PRBC surface. This binding is largely specific to PRBCs, because PMPs show little adherence to normal red blood cells. PMP adherence is also dependent on the P. falciparum erythrocyte membrane protein 1 variant expressed by PRBCs. PMP binding to PRBCs decreases after neutralization of PRBC surface proteins by trypsin or after treatment of PMPs with a mAb to platelet-endothelial cell adhesion molecule-1 (CD31) and glycoprotein IV (CD36). Furthermore, PMP uptake is a dynamic process that can be achieved by human brain endothelial cells (HBECs), inducing changes in the endothelial phenotype. Lastly, PMPs dramatically increase PRBC cytoadherence to HBECs. In conclusion, our study identifies several mechanisms by which PMPs may participate in CM pathogenesis while interacting with both PRBCs and HBECs. PMPs thereby provide a novel target for antagonizing interactions between vascular cells that promote microvascular sludging and blood brain barrier alteration during CM.

Common susceptibility alleles are unlikely to contribute as strongly as the FV and ABO loci to VTE risk: results from a GWAS approach.

Venous thromboembolism (VTE) is a complex disease that has a major genetic component of risk. To identify genetic factors that may modify the risk of VTE, we conducted a genome-wide association study by analyzing approximately 317 000 single nucleotide polymorphisms (SNPs) in 453 VTE cases and 1327 controls. Only 3 SNPs located in the FV and ABO blood group genes were found associated with VTE at a genome-wide significant level of 1.7 x 10(-7). Detailed analysis of these SNPs in additional cohorts of more than 1700 cases and 1400 controls revealed that the association observed at the FV locus was the result of the increased risk mediated by the FV Leiden mutation, whereas O and A2 blood groups were found to be at lower risk for VTE. Apart from the FV and ABO loci, no other locus was found strongly associated with VTE. However, using this large cohort of subjects, we were able to replicate the mild effects of 2 nonsynonymous SNPs, rs1613662 in GP6 and rs13146272 in CYP4V2, recently suspected to be associated with VTE.

Relationship between aspirin and clopidogrel responses in acute coronary syndrome and clinical predictors of non response.

OBJECTIVES: We have prospectively investigated the association between aspirin and clopidogrel responses and the clinical predictors of non response. METHODS: 635 Non ST Elevation Acute Coronary Syndrome (NSTE ACS) patients were included and received loading doses of 250 mg aspirin and 600 mg clopidogrel. We analyzed post-treatment maximal intensity of arachidonic acid and ADP-induced platelet aggregation (AA-Ag and ADP-Ag) and Platelet Reactivity Index of VAsodilator-Stimulated Phosphoprotein (PRI VASP). Aspirin and clopidogrel non responses were defined respectively by AA-Ag>30% and ADP-Ag>70%. RESULTS: Aspirin non responders patients had significantly higher ADP-Ag and PRI VASP than aspirin responders: 63.7+/-15.9% vs 55+/-19% (p=0.0001) and 73.6+/-13.3% vs 53+/-23% (p=0.0001) respectively and the rate of clopidogrel non responders was higher among aspirin non responders than aspirin responders: 36.7% vs 22.7% (p=0.003). In addition, clopidogrel non responders had significantly higher AA-Ag and rate of aspirin non responders than clopidogrel responders: 21.6+/-24% vs 10.3+/-19% (p=0.0001) and 22.8% vs 12.9% (p=0.003) respectively. Age and Body Mass Index (BMI) were significantly associated with non response to Clopidogrel (p=0.035 and 0.02 respectively) and diabetes mellitus by trend (p=0.07). CONCLUSION: We observed a relationship between aspirin and clopidogrel non responses and an association between age, BMI and diabetes mellitus and clopidogrel response.

Prospective analysis of factor XI deficiencies in the Marseilles area identified four novel mutations among 12 consecutive unrelated families.

Hereditary factor XI (FXI) deficiency is an autosomal bleeding disorder of variable severity but without a clear relationship between bleeding and FXI levels or mutation location or both. In the present study, the molecular basis of FXI deficiency in 16 patients from 12 families originating from the Marseilles area in the south of France was studied. FXI defect was evidenced by routine laboratory tests showing prolonged activated partial thromboplastin times and decreased factor XI activities. The promotor region, exons 1-15, and the exon-intron boundaries of the FXI gene were sequenced. Four novel mutations were found; three were missense mutations (Cys212Ser, Gly350Arg and Thr381Leu resulting from heterozygote mutation in exon 7, 10 and 11, respectively), and one was a one base deletion in exon 4 that induces a frameshift creating a stop codon four residues later (Thr57Ile fsX4). Eight previously reported mutations were also found. Contrarily to the Jewish, Basques or Briton populations, no recurrent mutation was identified. This cohort also illustrates that bleeding events occur not exclusively and not systematically in severe FXI deficiency but also in patients characterized by a mild FXI deficiency.

Contribution of novel biomarkers to incident stable angina and acute coronary syndrome: the PRIME Study.

AIMS: To compare whether novel inflammatory and haemostatic biomarkers are more predictive of well-characterized incident acute coronary syndrome (ACS) than stable angina (SA). METHODS AND RESULTS: We used data from the PRIME Study, a prospective cohort of 9758 asymptomatic middle-aged men recruited in Northern Ireland and France between 1991 and 1993. A nested case-control study was established with the baseline plasma sample of 269 incident cases and 538 matched controls. Odds ratios (ORs) for SA and ACS were estimated by conditional logistic regression analysis. After 5 years of follow-up, 107 incident SA and 162 ACS cases were validated. After adjustment for traditional risk factors, higher circulating levels of hs-CRP, ICAM1, interleukin 6 and interleukin 18 were equally predictive of SA and ACS (all P-values of OR comparison >0.05). In contrast, elevated levels of fibrinogen, von Willebrand factor, and possibly higher level of D-dimers and lower level of tissue factor pathway inhibitor were associated with ACS only. The comparison of the ORs showed a statistically significant difference for von Willebrand factor only [OR(4th vs. 1st quartile) = 2.99 (1.49-6.02) for ACS vs. 0.80 (0.33-1.94) for SA; P(z test) = 0.02]. CONCLUSION: This is the first population-based study suggesting that higher levels of circulating haemostatic markers and of von Willebrand factor, in particular, are significantly more predictive of incident ACS than SA.

Metabolic syndrome, haemostasis and thrombosis.

The metabolic syndrome (metS), a concurrence of abdominal fat, disturbed glucose and insulin metabolism, dyslipidemia, and hypertension has been strongly associated not only with subsequent development of type 2 diabetes but also with atherothrombosis. The physiopathology of this association is complex. The metS affects the thrombogenicity of circulating blood. Apart from its effect on platelets, a procoagulant and hypofibrinolytic state has been identified; mainly the result of the inflammatory state, dyslipidemia, and liver fat accumulation that accompany the MetS. Among haemostasis disturbances, the strong rise in the inhibitor of plasminogen activator type 1 plasma level is the most documented abnormality implicating the participation of the oxidative stress and inflammatory state developed during the metS. Endothelial dysfunction is also a central feature. Moreover, secretion products of fat tissues (adipokines) are now thought to have direct modulating effects on the vascular and the circulating cells. In support of these data, the metS, may predispose not only to atherosclerosis but also to venous thrombosis.

Effect of cytochrome p450 polymorphisms on platelet reactivity after treatment with clopidogrel in acute coronary syndrome.

Genetic polymorphisms of cytochrome P450 (CYP) isoforms may promote variability in platelet response to clopidogrel. This study was conducted to analyze, in 603 patients with non-ST elevation acute coronary syndromes, the effect of CYP3A4, CYP3A5, and CYP2C19 gene polymorphisms on clopidogrel response and post-treatment platelet reactivity assessed by adenosine diphosphate (ADP)-induced platelet aggregation, vasodilator-stimulated phosphoprotein phosphorylation index, and ADP-induced P-selectin expression. The CYP2C19*2 polymorphism was significantly associated with ADP-induced platelet aggregation, vasodilator-stimulated phosphoprotein phosphorylation index, and ADP-induced P-selectin expression in recessive (p <0.01, p <0.007, and p <0.06, respectively) and codominant (p <0.08, p <0.0001, and p <0.009, respectively) models, but the CYP3A4*1B and CYP3A5*3 polymorphisms were not. The CYP2C19*2 allele carriers exhibited the highest platelet index levels in multivariate analysis (p = 0.03). After covariate adjustment, the CYP2C19*2 allele was more frequent in clopidogrel nonresponders, defined by persistent high post-treatment platelet reactivity (ADP-induced platelet aggregation >70%; p = 0.03). In conclusion, the present data suggest that the CYPC19*2 allele influences post-treatment platelet reactivity and clopidogrel response in patients with non-ST elevation acute coronary syndromes.

Comparison of skin microvascular reactivity with hemostatic markers of endothelial dysfunction and damage in type 2 diabetes.

AIM: Patients with non-insulin-dependent diabetes mellitus (NIDDM) are at increased cardiovascular risk due to an accelerated atherosclerotic process. The present study aimed to compare skin microvascular function, pulse wave velocity (PWV), and a variety of hemostatic markers of endothelium injury [von Willebrand factor (vWF), plasminogen activator inhibitor-1 (PAI-1), tissue plasminogen activator (t-PA), tissue factor pathway inhibitor (TFPI), and the soluble form of thrombomodulin (s-TM)] in patients with NIDDM. METHODS: 54 patients with NIDDM and 38 sex- and age-matched controls were studied. 27 diabetics had no overt micro- and/or macrovascular complications, while the remainder had either or both. The forearm skin blood flow was assessed by laser-Doppler imaging, which allowed the measurement of the response to iontophoretically applied acetylcholine (endothelium-dependent vasodilation) and sodium nitroprusside (endothelium-independent vasodilation), as well as the reactive hyperemia triggered by the transient occlusion of the circulation. RESULTS: Both endothelial and non-endothelial reactivity were significantly blunted in diabetics, regardless of the presence or the absence of vascular complications. Plasma vWF, TFPI and s-TM levels were significantly increased compared with controls only in patients exhibiting vascular complications. Concentrations of t-PA and PAI-1 were significantly increased in the two groups of diabetics versus controls. CONCLUSION: In NIDDM, both endothelium-dependent and -independent microvascular skin reactivity are impaired, whether or not underlying vascular complications exist. It also appears that microvascular endothelial dysfunction is not necessarily associated in NIDDM with increased circulating levels of hemostatic markers of endothelial damage known to reflect a hypercoagulable state.

ADP-induced platelet aggregation and platelet reactivity index VASP are good predictive markers for clinical outcomes in non-ST elevation acute coronary syndrome.

Clopidogrel responsiveness has been proposed to be involved in recurrent ischemic events after stenting for non-ST elevation acute coronary syndromes (NSTE ACS). However, its biological definition is not consensual. We assess the value of ADP-induced platelet aggregation (ADP-Ag) and platelet reactivity index VASP (PRI VASP) in predicting recurrent ischemic events in patients with NSTE ACS undergoing percutaneous coronary intervention (PCI). We studied 195 consecutive NSTE ACS patients undergoing PCI after a 600 mg loading dose of clopidogrel. ADP-Ag and PRI VASP were analysed. The primary end-point was recurrent ischemic events within 30 days of PCI. It occurred in 14 patients (7%). Construction of ROC curves to examine the value of predictive models showed that sensitivity and specificity for primary endpoint were 79% and 76%, respectively, for a maximal intensity of ADP-Ag >or=70%, 93% and 50% for PRIVASP > 53%. The positive and negative predictive values were 21% and 98%, respectively, for ADP-Ag >or=70%, 12% and 99% for PRIVASP > 53%. In patients with NSTE ACS undergoing PCI, ADP-Ag and PRI VASP identify low responders to clopidogrel with an increased risk of recurrent ischemic events with respective cut-off values of 70% and 53%.

Plasminogen activator inhibitor-1, adipose tissue and insulin resistance.

PURPOSE OF REVIEW: Plasminogen activator inhibitor (PAI)-1 is a physiological inhibitor of plasminogen activators (urokinase and tissue types) and vitronectin. It is synthesized by adipose tissue, and its levels in plasma are increased in obesity and reduced with weight loss. Circulating PAI-1 level predicts development of type 2 diabetes, suggesting that it may be causally related to development of obesity. A role for PAI-1 in development of obesity has only partially been established, however. This review summarizes current knowledge, gives context to developments thus far and discusses controversies. RECENT FINDINGS: In addition to its role in atherothrombosis, PAI-1 might be involved in adipose tissue development. PAI-1 is produced by ectopic fat depots under the influence of inducers. Among the most recently described inducers are inflammation, oxidative stress and circadian clock protein. PAI-1 may play several roles in contributing to obesity: through indirect effects on insulin signalling, by influencing adipocyte differentiation and by regulating recruitment of inflammatory cells within adipose tissue. SUMMARY: These recent findings emphasize the involvement of PAI-1 in controlling the biology of adipose tissue; PAI-1 is an attractive new therapeutic target to retard the metabolic complications that accompany obesity.

Lack of association between the 807 C/T polymorphism of glycoprotein Ia gene and post-treatment platelet reactivity after aspirin and clopidogrel in patients with acute coronary syndrome.

Variability in platelet response to antiplatelet therapy and its clinical relevance have been well described. However, the underlying mechanisms remain unclear. It was the aim of the present study to assess whether the response to aspirin and clopidogrel may be influenced by the 807 C/T polymorphism of the glycoprotein Ia (GpIa) gene in patients with non-ST elevation acute coronary syndrome (NSTE ACS). Six hundred one NSTE ACS patients were included in our study and were divided into three groups: CC homozygotes, CT heterozygotes ad TT homozygotes. All patients received loading doses of 600 mg clopidogrel and 250 mg aspirin at least 12 hours before blood samples were drawn. Post-treatment platelet reactivity was assessed by post treatment ADP 10 microM-induced platelet aggregation (ADP-Ag), VASP phosphorylation (PRI VASP) and P-selectin expression. Non-response to dual antiplatelet therapy was defined by high post-treatment platelet reactivity (HPPR=ADP-Ag > 70%). Significant variability in the distribution of platelet parameters was observed in the overall study population. No significant difference in platelet parameters profiles was observed within patients having the same genotype, for ADP-Ag (p=0.33), PRIVASP (p=0.72) and P-selectin expression (p=0.37). The genotype frequencies of the 807 C/T polymorphism of the GpIa gene were similar in responders and non-responders defined by persistent HPPR (p=0.104). In conclusion, our study did not show any influence of 807 C/T polymorphism of GpIa gene on post-treatment platelet reactivity assessed by ADP-Ag, PRI VASP or P-selectin expression in 601 NSTE ACS patients.

Polymorphism A36G of the tumor necrosis factor receptor 1 gene is associated with PAI-1 levels in obese women.

The tumor necrosis factor (TNF) pathway may be implicated in etiopathogenesis of PAI-1 overexpression during obesity. The aim of this study was to investigate the influence of polymorphism A36G of the TNF receptor 1 (TNFRSF1A +36A/G) on plasma concentrations of PAI-1 in 163 obese (31 with the metabolic syndrome, MetS) and 150 lean, healthy women. Genotypic and allele frequencies did not significantly differ between obese and lean subjects. TNFRSF1A genotypes were significantly associated with sTNFR1 plasma levels in obese women only (p < 0.01); TNFRSF1A +36G/G obese carriers exhibited higher sTNFR1 and PAI-1 levels than A carriers (p < 0.01 and p < 0.05, respectively). In obese women, the presence of the MetS significantly potentiated the elevation of sTNFR1 and PAI-1 levels observed in the TNFRSF1A + 36G/G carriers. Our results suggest that association between TNFRSF1A +36G/G genotype and the MetS renders obese women more prone to activation of the TNF pathway reflected by high circulating sTNFR1 and PAI-1 levels.

Tiplaxtinin impairs nutritionally induced obesity in mice.

To investigate the effect of tiplaxtinin, designed as a synthetic inhibitor of plasminogen activator inhibitor-1 (PAI-1), on obesity, male C57Bl/6 mice (13-14 weeks old) were kept on a high-fat diet (20.1 kJ/g) for four weeks without or with addition of tiplaxtinin (PAI-039) at a dose of 2 mg/g food. At the time of sacrifice, body weights were significantly lower in the inhibitor-treated mice (p < 0.0005). The weights of the isolated subcutaneous and gonadal fat deposits were also significantly lower (both p < 0.0005), associated with adipocyte hypotrophy. Inhibitor-treated adipose tissues displayed similar blood vessel size, but a higher blood vessel density. Fasting glucose and insulin levels, as well as glucose-tolerance tests were not significantly affected by the inhibitor treatment, whereas plasma triglyceride levels were significantly reduced (p = 0.02) and LDL-cholesterol levels significantly enhanced (p = 0.0002). Insulin-tolerance tests revealed significantly lower glucose levels at the end of the test in the inhibitor treated mice (p = 0.03). Thus, in this model of diet-induced obesity in mice administration of tiplaxtinin resulted in impaired adipose tissue development.

The shedding activity of ADAM17 is sequestered in lipid rafts.

The tumor necrosis factor-alpha (TNF) converting enzyme (ADAM17) is a metalloprotease-disintegrin responsible for the cleavage of several biologically active transmembrane proteins. However, the substrate specificity of ADAM17 and the regulation of its shedding activity are still poorly understood. Here, we report that during its transport through the Golgi apparatus, ADAM17 is included in cholesterol-rich membrane microdomains (lipid rafts) where its prodomain is cleaved by furin. Consequently, ADAM17 shedding activity is sequestered in lipid rafts, which is confirmed by the fact that metalloproteinase inhibition increases the proportion of ADAM17 substrates (TNF and its receptors TNFR1 and TNFR2) in lipid rafts. Membrane cholesterol depletion increases the ADAM17-dependent shedding of these substrates demonstrating the importance of lipid rafts in the control of this process. Furthermore, ADAM17 substrates are present in different proportions in lipid rafts, suggesting that the entry of each of these substrates in these particular membrane microdomains is specifically regulated. Our data support the idea that one of the mechanisms regulating ADAM17 substrate cleavage involves protein partitioning in lipid rafts.