Dominique de Prost

Association of the Toll-Like Receptor 4 Gene Asp299Gly Polymorphism With Acute Coronary Events

Objective—Atherosclerosis is a chronic inflammatory disease of the blood vessels. Toll-like receptor 4 (TLR4) is a transmembrane receptor that is involved in mediating inflammatory responses to bacterial endotoxin and other ligands. The aim of this study was to search for an association between a common functional polymorphism of TLR4 —Asp299Gly—and acute coronary syndrome. Methods and Results—We conducted a case-control study of 183 patients with acute coronary syndromes and 216 controls. We screened the TLR4 gene for the Asp299Gly polymorphism using a 5′ fluorogenic assay. The 299Gly allele was associated with a decreased risk of acute coronary events independently of standard coronary risk factors. The adjusted odds ratio associated with this allele was 0.41 (95% CI, 0.18 to 0.95; P =0.037). In controls, TLR4 heterozygosity was also associated with a significant decrease in plasma fibrinogen and soluble vascular cellular adhesion molecule-1 levels (P <0.01). Conclusions—These results, which must be confirmed by a prospective longitudinal study, provide evidence of an association between the Asp299Gly polymorphism of the human TLR4 receptor and acute coronary syndromes. They confirm the previously reported involvement of TLR4 in carotid and femoral artery atherosclerosis.

Cardiovascular and Thromboembolic Risk Factors in Idiopathic Sudden Sensorineural Hearing Loss: A Case-Control Study

Objective: The pathogenesis of idiopathic sudden sensorineural hearing loss (ISSHL) remains unknown, but vascular involvement is one of the main hypotheses. The main objective of this study was to investigate the association between ISSHL and cardiovascular and thromboembolic risk factors. Study Design: Multicentric case-control study. Methods: Ninety-six Caucasian patients with ISSHL and 179 sex- and age-matched controls were included. Patients were evaluated on the day of the inclusion and 1 week, 3 weeks and 3 months later. Clinical information concerning personal and familial cardiovascular and thromboembolic risk factors and concerning the ISSHL was collected. Blood samples were collected for genetic analysis of factor V Leiden and G20210A polymorphism in the factor II gene. The severity of the hearing loss was classified as mild (21–40 dB), moderate (41–70 dB), severe (71–90 dB) and profound or total (>90 dB). Hearing improvement was calculated as a relative improvement of hearing thresholds using the contralateral ear as baseline. Results: Systolic blood pressure was higher in patients (130 ± 1.7 mm Hg) than in controls (124 ± 1.1 mm Hg, p = 0.003). The personal/familial history of cardiovascular events was also more prevalent in patients (p = 0.023 and p = 0.014, respectively), whereas no difference was found in the prevalence of personal cardiovascular risk factors (hypertension, diabetes mellitus, hyperlipidemia, smoking habits). There was no correlation between the audiogram type, the hearing outcome and the presence of cardiovascular risk factors. No significant difference was observed in the personal/familial history or in the presence of thromboembolic risk factors. The prothrombin and factor V mutations were uncommon in both patients and controls. The final hearing threshold was only correlated with the severity of the initial hearing loss (p < 0.001), but not influenced by the presence of vertigo, audiogram type, time elapsed from onset of ISSHL to hospitalization or failure of a previous oral therapy. Hearing stabilization was obtained at 21 days in 92% of patients. Conclusion: These results support the theory of vascular involvement as the etiology of some cases of ISSHL. The sole predictive factor of poor final hearing is the severity of the initial hearing loss.

Von Willebrand Factor and ADAMTS13 A Candidate Couple for Preeclampsia Pathophysiology

Objective—The goal of this study was to search for an association between a desintegrin-like and metalloprotease thrombospondin type 1 motif, member 13 (ADAMTS13) levels and the occurrence of preeclampsia, its characteristics (time-onset and severity), and its consequences (occurrence of fetal growth restriction or preterm delivery). Methods and Results—We studied 140 pairs of women in a case-control study with 3 matching criteria: maternal age, gestational age, and ethnic origin. We measured ADAMTS13 activity using a fluorescence resonance energy transfer assay with the fluorescence resonance energy transfer-VWF73 peptide. ELISA was used to assess protein antigen levels: ADAMTS13, von Willebrand Factor (VWF), interleukin-6, C-reactive protein, P-selectin, and thrombospondin-1. The lowest levels of ADAMTS13 (activity ⩽70% or antigen ⩽592 ng/mL) were significantly associated with preeclampsia (odds ratios [OR] [95% confidence interval] of 4.2 [1.1 to 15] and 14.3 [1.7 to 123], respectively). This association was independent of VWF levels and preeclampsia risk factors but dependent on interleukin-6 and C-reactive protein levels for ADAMTS13 activity. Levels of ADAMTS13 activity (⩽57%) were significantly associated with early-onset preeclampsia (OR=2.5 [1.1 to 5.8]). Severe preeclampsia was associated with the highest levels of P-selectin (>57 ng/mL) (OR=3.4 [1.2 to 9.7]). Conclusion—Preeclampsia is associated with decreased levels of ADAMTS13, independently of VWF. This decrease is quantitative, occurs early, and seems to be dependent on inflammation. Our results suggest that ADAMTS13 could participate in the pathophysiology of preeclampsia.

Recombinant Activated Protein C Attenuates Endothelial Injury and Inhibits Procoagulant Microparticles Release in Baboon Heatstroke

Objectives—We tested the hypothesis that the antithrombotic and cytoprotective effects of recombinant human activated protein C (rhAPC) protect baboons against the lethal effects of heatstroke. Methods and Results—Fourteen anesthetized baboons assigned randomly to rhAPC (n=7) or control group (n=7) were heat-stressed in a prewarmed incubator at 44 to 47°C until systolic blood pressure fell below 90 mm Hg, which signaled severe heatstroke. rhAPC was administered intravenously (24 &mgr;g/kg/h) for 12 hours at onset of heatstroke. Heat stress induced coagulation and fibrinolysis activation as evidenced by a significant increase from baseline levels in plasma levels of thrombin-antithrombin (TAT) complexes, tissue plasminogen activator, and D-dimer. Heat stress elicited cell activation/injury as assessed by the release of interleukin (IL)-6, soluble thrombomodulin, and procoagulant microparticles (MPs). rhAPC did not significantly reduce heatstroke-induced thrombin generation, and D-dimer and had no effect on fibrinolytic activity. In contrast, rhAPC infusion attenuated significantly the plasma rise of IL-6 and inhibited the release of soluble thrombomodulin and MPs as compared with control group. No difference in survival was observed between rhAPC-treated and control group. Conclusions—rhAPC given to heatstroke baboons provided cytoprotection, but had no effect on heatstroke-induced coagulation activation and fibrin formation. Inhibition of MPs by rhAPC suggested a novel mechanism of action for this protein.

The V249I polymorphism of the CX3CR1 gene is associated with fibrostenotic disease behavior in patients with Crohn's disease.

Objectives CX3CR1, the receptor of CX3CL1/fractalkine, is involved in regulation of inflammatory response and the CX3CR1-I249-M280 naturally occurring mutants are associated with altered binding to the ligand. Our aim was to evaluate the frequency of CX3CR1 V249I and T280M polymorphisms and NOD2/CARD15 mutations in Crohns disease patients and to search for a relationship with phenotype. Methods Clinical data were retrospectively collected. V249I and T280M polymorphisms of CX3CR1 gene and NOD2/CARD15 mutations (R702W, G908R, 3020InsC) were identified. Results Two hundred and thirty-nine patients (140 females, 39.7±14.1 years) were included. About 37.4% were heterozygous and 8.8% were homozygous for the V249I CX3CR1 polymorphism, 18.1% were heterozygous and 1.3% homozygous for the T280M CX3CR1 polymorphism and 35.9% had at least one of the three mutations of NOD2/CARD15. The T280M CX3CR1 polymorphism was not associated with any phenotype. In univariate analysis, stenosis was significantly associated with both V249I CX3CR1 polymorphism and 3020InsC NOD2/CARD15 mutations. In smoker patients carrying the CX3CR1 allele I249, there was a significant increase in the frequency of fibrostenosing disease [P=0.005, odds ratio (OR): 3.25] whereas this relationship disappeared in the group of nonsmokers (P=0.72). In multivariate analysis, 3020InsC NOD2/CARD15 mutations and the V249I CX3CR1 polymorphism were independent risk factors for intestinal stenosis (P=0.046, OR: 1.8 and P=0.044, OR: 2.4, respectively). Conclusion In Crohns disease, V249I CX3CR1 polymorphism is associated with intestinal strictures, particularly in smokers. This association is independent of CARD15 mutations.

Reference values for rotation thromboelastometry (ROTEM®) parameters following non-haemorrhagic deliveries. Correlations with standard haemostasis parameters

Reference values for rotation thromboelastometry (ROTEM®) parameters following non-haemorrhagic deliveries. Correlations with standard haemostasis parameters -

Metabolic activity of human polymorphonuclear leucocytes: relation to ingestion rate.

Abstract. The ingestion rate and oxygen‐dependent metabolic activities of normal human polymorphonuclear leucocytes were measured with heat‐killed Klebsiella as the particle. Since the experimental conditions were similar for each measurement, it was possible to make direct correlations between each oxygen‐dependent reaction and (1) ingestion rate and (2) the other oxygen‐dependent reactions.

Modulation of tissue factor expression by rapamycin and FK-506 in lipopolysaccharide-stimulated human mononuclear cells and serum-stimulated aortic smooth muscle cells.

Inflammation is a key pathogenic component of atherosclerosis; it also promotes thrombosis, a process underlying acute coronary events and stroke. Cells present in atherosclerotic plaque show abnormal tissue factor (TF) expression. Macrolides, in addition to their antimicrobial properties, have antiinflammatory effects that might help prevent atherothrombosis. The aim of this study was to determine the effect of an immunosuppressant macrolide, rapamycin (Sirolimus), on the expression of TF and its inhibitor (TFPI) by monocytic cells (human blood mononuclear and THP-1 cells) and human aortic smooth muscle cells, in comparison with FK-506 and azithromycin. In monocytic cells, rapamycin and FK-506 inhibited LPS-induced TF activity, antigen and mRNA expression through a transcriptional mechanism involving NF-kappaB. In smooth muscle cells, rapamycin and azithromycin had no effect on serum-induced TF expression, while FK-506 increased serum-induced TF protein and mRNA expression. TFPI levels in the culture supernatants of serum-stimulated smooth muscle cells were not modified by any of the three macrolides. Rapamycin slightly inhibits TFPI induction by LPS in monocytic cells. In addition to its recently established efficacy in the prevention of stent restenosis, the inhibitory effect of rapamycin on the TF pathway might have interesting therapeutic implications.

Combined factor V leiden (G1691A) and prothrombin (G20210A) genotyping by multiplex real-time polymerase chain reaction using fluorescent resonance energy transfer hybridization probes on the Rotor-Gene 2000.

Several methods have been developed to detect common single point mutations in the factor V and prothrobin genes that are risk factors for thrombophilia. Most are basd on PCR followed by restriction enzyme digestion and electrophoresis (RFLP), but gel analysis has certain limitations, and alternative detection methods, including real-time PCR, have therefore been developed. In this study we developed and evaluated a combined factor V Leiden and prothrombin (G20210A) genotyping method based on multiplex real-time PCR with fluorescent resonance energy transfer (FRET) hybridization probes on the Rotor-Gene 2000. Two hundred subjects were screened for the two mutations. The FRET assay clearly discriminated among wild-type, homozygous and heterozygous status for the two mutations, and the results were in full agreement with those of the RFLP assay. This robust FRET probe-based assay also has a higher throughput capacity than conventional methods, handling up to 72 samples in 90 min.

Search for an association between V249I and T280M CX3CR1 genetic polymorphisms, endothelial injury and preeclampsia: the ECLAXIR study.

Background Preeclampsia and coronary-artery disease share risk factors, suggesting common pathophysiological mechanisms. CX3CR1/CX3CL1 mediates leukocyte migration and adhesion and has been implicated in the pathophysiology of several inflammatory diseases. M280/I249 variants of CX3CR1 are associated with an atheroprotective effect and reduced endothelial dysfunction. The aim of this study was to search for an association between V249I and T280M polymorphisms of CX3CR1, preeclampsia and endothelial dysfunction. Methodology/Principal Findings We explored these polymorphisms with real-time polymerase chain reaction in a case-control study (184 white women with preeclampsia and 184 matched normotensive pregnant women). Endothelial dysfunction biomarkers including von Willebrand factor, VCAM-1 and thrombomodulin, as well as the soluble form of CX3CL1 were measured by enzyme-linked immunosorbent assays (ELISA). The I249 and M280 alleles were associated neither with preeclampsia, nor with its more severe form or with endothelial injury. In contrast, we found a trend toward increased CX3CL1 levels in preeclampsia patients, especially in early-onset- preeclampsia as compared to its level in later-onset- preeclampsia. Conclusions/Significance This is the first study to characterize the CX3CR1 gene polymorphisms in patients with preeclampsia. We found no differences in genotype or haplotype frequencies between patients with PE and normal pregnancies, suggesting that maternal CX3CR1 V249I and T280M polymorphisms do not increase susceptibility to preeclampsia. Further studies should be performed to directly evaluate the pathophysiological role of CX3CL1, a molecule abundantly expressed in endometrium, which has been shown to stimulate human trophoblast migration.