Topic 07 – Basic science
241
Genome-wide association analysis identifies 3 common variants pre-disposing to Brugada syndrome
Julien Barc (1), Connie Bezzina (1), Yuka Mizusawa (1), Carol Remme (1), Jean-Baptiste Gourraud (2), Floriane Simonet (2), Arie Verkerk (3), Peter Schwartz (4), Lia Crotti (4), Federica Dagradi (4), Pascale Guicheney (5), Véronique Fressart (5), Antoine Leenhardt (6), Charles Antzelevitch (7), Susan Bartkowiak (7), Eric Schulze-Bahr (8), Sven Zumhagen (8), Elijah Behr (9), Rachel Bastiaenen (9), Jacob Tfelt-Hanson (10), Morten Salling Ole-sen (10), Stefan Kaab (11), Britt Beckmann (11), Peter Weeke (12), Hiroshi Watanabe (13), Naoto Endo (14), Tohru Minamino (13), Minoru Horie (15), Seiko Ohno (15), Kanae Hasegawa (15), Naomasa Makita (16), Akihiko Nogami (17), Wataru Shimizu (18), Takeshi Aiba (18), Philippe Froguel (19), Beverley Balkau (20), Olivier Lantieri (21), Cornelia Wiese (22), David Weber (22), Rianne Wolswinkel (1), Ruben Coronel (1), Bas Boukens (1), Eric Charpentier (2), Stéphanie Chatel (2), Aurore Despres (2), Françoise Gros (2), Florence Kyndt (2), Simon Lecointe (2), Pierre Lindenbaum (2), Vincent Portero (2), Jade Violleau (2), Manfred Gessler (22), Hanno Tan (1), Dan Roden (12), Vincent Christoffels (3), Hervé Le Marec (2), Arthur Wilde (1), Vincent Probst (2), Jean-Jacques Schott (2), Christian Dina (2), Richard Redon (2)
(1) Heart Failure Research Center, Department of Experimental Cardiology, Amsterdam, Pays-Bas – (2) INSERM, UMR1087, l’institut du thorax, Nantes, France – (3) Heart Failure Research Center, Department of Anatomy, Embry-ology and PhysiEmbry-ology, Amsterdam, Pays-Bas – (4) University of Pavia, Department of Molecular Medicine, Pavia, Italie – (5) Faculté de Médecine Pierre et Marie Curie, site Pitié-Salpêtrière, INSERM, U956, Paris, France – (6) Hôpital Bichat-Claude Bernard, Service de Cardiologie et Centre de Référence des Maladies Cardiaques Héréditair, Paris, France – (7) Masonic Medical Research Laboratory, Department of Experimental Cardiology, Utica, Etats-Unis – (8) Institute for Genetics of Heart Diseases (IfGH), Department of Cardiovascular Medicine University Hospital, Munster, Alle-magne – (9) St George's University of London, Cardiovascular Sciences Research Centre, Londres, Royaume-Uni – (10) University of Copenhagen, Laboratory of Molecular Cardiology, Copenhague, Danemark – (11) Univer-sity Hospital Munich, Department of Medicine I, Munich, Allemagne – (12) Vanderbilt University School of Medicine, Department of Medicine and Pharmacology, Nashville, Etats-Unis – (13) Niigata University Graduate School of Medical and Dental Sciences, Department of Cardiovascular Biology and Medicine, Niigata, Japon – (14) Niigata University Graduate School of Medical and Dental Sciences, Department of Regenerative and Transplant Medicine, Niigata, Japon – (15) Shiga University of Medical Science, Depart-ment of Cardiovascular and Respiratory Medicine, Otsu, Japon – (16) Naga-saki University, Department of Molecular Physiology, NagaNaga-saki, Japon – (17) Yokohama Rosai Hospital, Division of Heart Rhythm Management, Yoko-hama, Japon – (18) National Cerebral and Cardiovascular Center, Division of Arrhythmia and Electrophysiology, Osaka, Japon – (19) Lille Pasteur Institute, UMR 8199, Lille, France – (20) Centre for research in Epidemiology and Pop-ulation Health, Inserm U1018, Villejuif, France – (21) Institut inter Régional pour la Santé (IRSA), La Riche, France – (22) University of Wuerzburg, Devel-opmental Biochemistry, Theodor-Boveri-Institute, Wuerzburg, Allemagne
The Brugada Syndrome (BrS) is considered as a rare Mendelian disorder with autosomal dominant transmission. BrS is associated with an increased risk of sudden cardiac death and specific electrocardiographic features consisting of ST-segment elevation in the right precordial leads. Loss-of-function mutations in SCN5A, encoding the pore-forming subunit of the cardiac sodium channel (NaV1.5), are identified in ~20% of patients. However, studies in families
har-boring mutations in SCN5A have demonstrated low disease penetrance and in some instances absence of the familial SCN5A mutation in some affected mem-bers. These observations suggest a more complex inheritance model.
To identify common genetic factors modulating disease risk, we conducted a genome-wide association study on 312 individuals with BrS and 1,115 ancestry-matched controls.
Two genomic loci displayed significant association. Replication testing on two independent case/control sets from Europe (598/855) and Japan (208/ 1016), confirmed both associations and revealed a third locus. The cumulative effect of the 3 loci on disease susceptibility was large with an odds ratio of 21.5 in the presence of more than 4 risk alleles versus less than 2.
Two out of three loci had previously been shown to influence ECG conduction parameters in the general population. The third locus encompasses a transcription factor which has not previously been implicated in cardiac electrical function and arrhythmia. Functional studies in mice heterozygous knock-out for this transcription factor identified differences in expression of NaV1.5, in addition to differences in conduction of the cardiac electrical
impulse in the right ventricular outflow tract.
Altogether, our findings (1) indicate that common genetic variation may have a strong impact on predisposition to BrS, and (2) identify a new gene involved in the pathogenesis of the disease.
242
Scn5a+/ΔQKP mice develop exhibit long QT syndrome and dilated cardiomyopathy
Jérôme Montnach, Agnès Carcouet, Isabelle Baró, Flavien Charpentier
Inserm UMR1087, CNRS UMR6291, l’institut du thorax, Université de Nantes, Nantes, France
Deletion of QKP1507-1509 amino-acids in SCN5A gene product, the Nav1.5 voltage-gated Na+channel, was first discovered in a family with type
3 long QT syndrome (LQT3) and is now associated with a larger phenotypic spectrum of LQT3, conduction disorder, dilated cardiomyopathy (DCM) and high incidence of youth sudden death. This mutation does not affect the peak Na+
current but rather increases the late Na+
current. In order to identify the mechanism of DCM in these patients, a knock-in mouse model presenting the delQKP1510-1512 mutation, has been generated.
Four groups of mice were studied: homozygous wild-type (Scn5a+/+
), heterozygous flp deleter (Scn5a+/flp), heterozygous neomycin-ΔQKP
(Scn5a+/neo) and Scn5a+/ΔQKPmice. Scn5a+/ΔQKPmice exhibited high early
mortality with a percentage of death of 50% at the age of 5-6 weeks. Only 35% of Scn5a+/ΔQKPmice survived until week 10 whereas 91% of control mice. Six-lead ECG recorded on 4 week-old mice anesthetized with isoflu-rane showed that 6/15 Scn5a+/ΔQKPexhibited numerous ventricular extrasystoles
and/or runs of non-sustained ventricular tachycardia. Six Scn5a+/ΔQKPmice were in sinus rhythm and showed a prolonged QT interval (QTc=95 ± 12 ms, versus 46 ± 2 ms in controls, n=22). Finally, 3 Scn5a+/ΔQKPmice were in 2:1
atrio-ventricular block because of highly prolonged QT interval. All control mice were in sinus rhythm. Sudden cardiac death secondary to ventricular fibrillation could be recorded in one Scn5a+/ΔQKPmouse, suggesting that
part of early mortality could be attributed to arrhythmias. Alternatively, mice could also die from heart failure. Indeed, Scn5a+/ǻQKPmice surviving
until the age of 10 weeks showed a heart weight / tibia length ratio of 12.1 ± 3.4 (n=4) versus 7.5 ± 0.5 in control mice (n=14, p< 0.05).
This study shows that Scn5a+/ǻQKPmice reproduce the phenotype of the
human mutation carriers, long QT syndrome, heart failure and increased risk of sudden death at a young age.
243
LMNA: a new gene involved in idiopathic atrioventricular blocks?
Xavier Daumy (1), Daniel Trujillano (2), Florence Kyndt (1), Stephan Ossowski (2), Hervé Le Marec (1), Xavier Estivill (2), Jean-Baptiste Gour-raud (1), Vincent Probst (1), Richard Redon (1), Jean-Jacques Schott (1)
(1) L'institut du thorax (INSERM), Unité Inserm UMR 1087 / CNRS UMR 6291, Nantes, France – (2) Center for Genomic Regulation, Barcelona, Espagne
Progressive cardiac conduction defect (PCCD) is one of the most common cardiac conduction disturbances characterized by progressive alteration of car-diac conduction through the His-Purkinje system with right or left bundle branch block and widening of QRS complexes.
January 18
thsimple fibrosis process affecting the conduction system. Familial forms of car-diac conduction disturbance have been reported with an autosomal dominant inheritance since then.
Several genes coding for ionic channels have already been described in this condition: SCN5A, SCN1B and TRPM4. Other genes have been identified in PCCD forms associated with other cardiac disease such as congenital heart disease or cardiomyopathy: NKX2.5, TBX5, PRKAG2, LMNA...
A study based on an epidemiological approach has enabled us to identify four families in the western part of France. In one of them, the exomes of four patients have been sequenced using the next generation sequencing techno-logies (Illumina HiSeq). This strategy has allowed us to identify one missense variant that seems to segregate with the pathology according to first validation tests.
This family was composed of 49 members (11 affected; 5 PM). The most frequent abnormal conduction was PB with LAHB (n=5) and LAHB (n=3). In this family, one patient had a complete AV block. No ECG was recorded before implantation for one patient. Nine patients presented a first-degree AV block. On average, age and heart rate were not different between the affected and non-affected members. There was a significant difference between PR duration of the affected group and non-affected group (224±28 ms vs 173±19 ms; p<0.0001) and QRS duration (120±16 ms vs 87±10 ms, respectively; p<0.0001).
The variant is localized in the LMNA gene (c.G1039A; p.E347K). Muta-tions in that gene are known to cause a wide range of human diseases (more than 10 different clinical syndromes have been attributed to LMNA mutations) including conduction system disease associated with dilated cardiomyopathy. Our study is the first showing LMNA as a potential responsible of isolated PCCD.
244
Long QT syndrome transmission distortion
Hideki Itoh (1), Myriam Berthet (1), Véronique Fressart (2), Isabelle Denjoy (3), Svetlana Maugenre (1), Didier Klug (4), Yuka Mizusawa (5), Takeru Makiyama (5), Nynke Hofman (6), Anja Husemann (7), Wataru Shimizu (8), Arthur Am Wilde (6), Eric Schulze-Bahr (7), Minoru Horie (9), Sophie Tezenas Du Montcel (10), Pascale Guicheney (1)
(1) INSERM UMR S956, Faculté de Médecine Pierre et Marie Curie, Paris, France – (2) Hôpital La Pitié-Salpétrière, Service de Biochimie Métabolique, Paris, France – (3) Hôpital Bichat-Claude Bernard, Paris, France – (4) Hôpital Cardiologique, Lille, France – (5) Kyoto University Graduate School of Medicine, Kyoto, Japon – (6) Academic Medical Cen-ter, University of Amsterdam, Amsterdam, Pays-Bas – (7) Institute for Genetics of Heart Diseases, Münster, Allemagne – (8) Nippon Medical School, Department of Cardiovascular Medicine, Tokyo, Japon – (9) Shiga University of Medical Science, Otsu, Japon – (10) Hôpital La Pitié-Salpêtrière, Unité de Biostatistiques et Information Médicale, Paris, France
Long QT syndrome (LQTS) is an inherited syndrome with QT interval pro-longation, syncope, and risk of sudden death from ventricular fibrillation. Fourteen candidate genes are implicated, of which KCNQ1 (LQT1), KCNH2 (LQT2) and SCN5A (LQT3) are the major genetic subtypes, accounting for approximately 80% of all genotyped LQTS patients. We previously reported transmission ratio distortion (TRD) in European LQTS families, with an excess of mutation carriers and female predominance suggesting a survival advantage for affected gametes. To determine whether this phenomenon depends on LQTS mutations or gene loci, we studied parental and grandpa-rental origins of alleles transmitted in LQTS and control families and the role of channel dysfunction in the transmission distortion.
We studied genotyped members from 679 European and Japanese LQTS families (2748 carriers). We determined grandparental and parental origins of mutant alleles in 1892 children and 619 grandchildren, and grandparental origin of normal alleles in healthy children from 44 three-generation control CEPH families. In families from Europe and Japan, mutant alleles were significantly more frequently maternal than paternal in origin (61%, 1155 vs. 737 alleles, p<0.001). The ratio of maternal alleles in LQT1 (66%) was signif-icantly higher than in LQT2 (56%, p<0.001) and LQT3 (57%, p=0.01). Unlike control families' Mendelian distribution of grandparental alleles, mutant grandparental LQT1 and LQT2 alleles in grandchildren showed excess maternal grandmother alleles.
For LQT1, maternal origin mutant alleles bore a significant relationship to the level of dysfunction (dominant negative 67%, non-dominant 58%, p<0.03), however for LQT2 or LQT3 this association was not significant. In conclusion, there is an excess of LQTS mutant alleles of maternal origin, most pronounced in LQT1, which is homogeneous across ethnic groups. This is not linked to a locus-specific grandparental origin allele transmission distortion, but most probably to potassium channel dysfunction.
245
A long QT mutation substitutes cholesterol for phosphatidylinositol-4,5-bisphosphate in KCNQ1 channel regulation
Fabien Coyan (1), Mohamed Yassine Amarouch (1), Julien Piron (1), Jérôme Mordel (1), Céline Nicolas (1), Jean Mérot (1), Annick Thomas (2), Robert Brasseur (2), Isabelle Baró (1), Gildas Loussouarn (1)
(1) INSERM U1087, Nantes, France – (2) Centre de Biophysique Molécu-laire Numérique, Gembloux, Belgique
KCNQ1 is the α-subunit of a voltage-dependent potassium channel. In the heart, KCNQ1 forms a channel complex with KCNE1 to generate the slow component of the delayed rectifier current IKs. Phosphatidylinositol-4,5-bisphosphate (PIP2) is a cofactor necessary for the activity of KCNQ1
channels. Some Long QT mutations of KCNQ1, including R243H, R539W and R555C have been shown to decrease KCNQ1 interaction with PIP2. PIP2
sensitivity of these mutants was determined by using the short acyl chains analog diC8-PIP2. Since then, hERG channels, known to be regulated by
PIP2, have been shown to be insensitive to this analog, suggesting that a
channel affinity for PIP2 and diC8-PIP2 might be quite different. Hence, we
set out to use other approaches to further study the PIP2 affinity of KCNQ1
mutant channels.
We measured the kinetics of Mg2+– and wortmannin-induced current
rundown in COS-7 cells expressing the WT or mutant channels and used either currents amplitudes at the end of the depolarizing step or tail-cur-rents amplitudes as readout. We show that R539W is very peculiar: as opposed to WT and R555C channels, the R539W channel current is barely running down when available PIP2 is decreased, either by wortmannin
application in whole-cell, or by magnesium application in inside-out con-figuration. Consistent with that, the R539W channel is also insensitive to extracellular osmolarity, known to modulate the channel activity via PIP2.
These results suggest that KCNQ1-R539W mutation shortcuts PIP2 in the
channel open pore stabilization. Structural model prediction suggests that the introduced tryptophan in R539W interacts with cholesterol. Both cyclodextrin application (to deplete membrane cholesterol) on R539W and substitution of R539 by residues other than tryptophan restore channel run-down, consistent with the supposed tryptophan-cholesterol interaction. We conclude that the R539W/cholesterol interaction replaces R539/PIP2
inter-action in the channel open pore stabilization.
246
Complex Brugada syndrome inheritance in a family harbouring com-pound SCN5A and CACNA1C mutations
Julien Barc (1), Delphine Béziau (2), Thomas O'Hara (3), Lauriane Le Gloan (2), Mohamed Yassine Amarouch (2), Aude Solnon (4), Domi-nique Pavin (4), Simon Lecointe (2), Patricia Bouillet (4), Pascale Gui-cheney (5), Isabelle Denjoy (6), Richard Redon (2), Philippe Mabo (4), Hervé Le Marec (2), Gildas Lousouarn (2), Florence Kyndt (2), Jean-Jacques Schott (2), Vincent Probst (2), Isabelle Baro (2)
(1) Heart Failure Research Center, Department of Experimental Cardio-logy, Amsterdam, Pays-Bas – (2) INSERM, UMR1087, l’institut du thorax, Nantes, France – (3) Institute for Computational Medicine,Johns Hopkins University, Baltimore, Etats-Unis – (4) CHU Rennes, Département de car-diologie et des maladies vasculaires, INSERM U642, Rennes, France – (5) Faculté de Médecine Pierre et Marie Curie, site Pitié-Salpêtrière, INSERM, U956, Paris, France – (6) Hôpital Bichat-Claude Bernard, Ser-vice de Cardiologie et Centre de Référence des Maladies Cardiaques Héréditaires, Paris, France
the right precordial leads and is associated with an increased risk of SCD from polymorphic ventricular tachyarrhythmia and ventricular fibrillation. We have recently reported families with BrS and SCN5A mutations where some affected members do not carry the familial mutation. These findings suggest the involvement of additional genetic determinants for BrS in affected families.
We report the identification of 2 distinct gene mutations within a family presenting a BrS (5 affected individuals), cardiac conduction defects (CCD, 3 affected individuals) and a shortened QT interval (SQT, 4 affected individ-uals). The first mutation is nonsense and lies within the SCN5A gene, which codes for the α-subunit of the cardiac Na+ channel NaV1.5. The second
muta-tion is missense and alters the CACNA1C gene, which encodes the α-subunit CaV1.2 of the L-type cardiac Ca2±channel. The SCN5A mutation strictly
seg-regates with CCD while 4 out of the 5 BrS patients carrying the CACNA1C variant present with SQT and/or BrS. Patch-clamp studies identified a net loss-of-function of the CaV1.2 channel in presence of the missense mutation.
In addition, western-blot experiments showed a global expression defect while increased mobility of CaV1.2 channels on cell surface was revealed by FRAP
experiments. Finally, computer simulations of the 2 mutations recapitulated patient phenotypes.
This study reports a rare mutation in the CACNA1C gene as causing BrS and/or SQT interval in a family carrying a stop mutation in the SCN5A gene, which does not segregate with the BrS. Furthermore one affected family member does not harbor any mutation among the 12 BrS known genes under-lying the complexity of the BrS inheritance and the difficulties of the preven-tion by a pre-symptomatic genetic screening.
247
Prevalence and impact of prosthesis-patient mismatch in patients with paradoxical low flow, severe aortic stenosis
Cyrille Boulogne (1), Dania Mohty (1), Victor Aboyans (1), Philippe Pibarot (2), Najmeddine Echahidi (1), Jean Gaston Dumesnil (2), Elisa-beth Cornu (3), Patrice Virot (1), Marc Laskar (3)
(1) CHU Limoges, Cardiologie, Limoges, France – (2) Institut de Cardio-logie de Quebec, Echocardiographie, Quebec, Canada – (3) CHU Limoges, Chirurgie thoracique, cardiovasculaire et angiologie, Limoges, France
Purpose: Patients with severe aortic stenosis (AS) and paradoxical low
flow (PLF) (indexed stroke volume< 35ml/m2) despite preserved left ventric-ular ejection fraction >50%, have worse outcome compared to those with AS but normal flow. Moreover, Prosthesis-Patient Mismatch (PPM) (indexed prosthetic valve effective orifice area< 0.85cm2/m2) after aortic valve
replace-ment (AVR) is a predictor of higher mortality. However, the impact of PPM in patients with PLFAS on long-term survival is unknown. Our aim was to analyze the prevalence and the impact on long-term survival of PPM in patients with PLFAS.
Methods: 667 consecutive patients (age 74±8 years, 42% female, AVA
0.69±0.16 cm²) with preserved LVEF who underwent AVR for severe AS at our institution between 2000 and 2010 were included in this study. Patients were divided into 4 groups according to the presence/absence of PLF at car-diac catheterization and presence/absence of PPM following AVR and we compared short and long-term survival between these groups.
Results: Among the 667 patients, 26% had PLFAS and PPM occurred
in 54% of patients after AVR. Compared to patients with no PLF & no PPM (36% of the total cohort), those with PLF & PPM (15%) were signif-icantly older, with more comorbidities. The 30-day mortality did not differ between the PLF- PPM and no-PLF-no PPM group. However, the 10-yr survival rate was significantly reduced in the PLF-PPM (37±9%) group compared to no PLF-no PPM (70±5%; p=0.003). In multivariate analysis adjusting for all predictors of survival, concomitant presence of PLF & PPM was an independent predictor of survival (HR= 2.68 95%CI: 1.5-4.4; p=0.0003)
Conclusion: In this catheterization-based study, patients with PLF and
PPM have worse outcome when compared to those without these 2 condi-tions.
248
Tramadol an opiate synthesis, accused of increased bleeding risk of AVK
Ahmed Hamzaoui (1), Hamdi Sonia (2), Chakroun Mahdi (3), Ben Hamda Kholdoun (4), Mohssen Hssine (1), Matouk Faouzi (2)
(1) Hôpital Fattouma Bourguiba, Hématologie biologique et banque du sang, Monastir, Tunisie – (2) Hôpital Fattouma Bourguiba, Cardiologie, Monastir, Tunisie – (3) Faculté de médecine Sfax Hôpital Fattouma Bour-guiba, Cardiologie, Monastir, Tunisie – (4) Faculté de Monastir Hôpital Fattouma Bourguiba, Cardiologie, Monastir, Tunisie –
Tramadol, a centrally acting analgesic derived from chemical synthesis, is introduced into the armamentarium since its 1977. Due to different pharma-ceutical presentations and analgesic action fast and efficient (step II analgesics WHO), its use clinic is widespread both oral and parenteral and by locore-gional. The scale that takes use of tramadol after removal of dextropr-poxyphéne market is not quite trivial. In our cardiology department, we observed over a period of 5 months INR disturbances in a population who received tramadol IVL. This population is a group of 6 patients of average age 46 – 19.5 years including 4 females and 2 males treated with acenocoumarol. Stopping tramadol subsequently restores its target INR between 2 to 3 or 3 to 4.5 depending on the clinical context of anticoagulation. This restoration is not immediate but the INR is restored after an average of 5 days. In our study, we have refuted the hypothesis suggesting that tramadol interfere with the biolog-ical analysis of PT and INR for the absence of signs of bleeding. This is by witnesses containing the blood of patients not anticoagulated and receiving tramadol. The most likely hypothesis is that pharmacokinetic interaction involving the step of biotransformation and more specifically one that takes place in the liver microsomes via cytochrome P450
249
Hyperhomocysteinemia and severity of coronary artery disease in the elderly
Nadia Koubaa (1), Sonia Hammami (2), Raja Chaaba (1), Sounira Mehri (1), Mohamed Hammami (1)
(1) Faculté de Medecine, Biochimie, Monastir, Tunisie – (2) Hôpital Fattouma Bourguiba, Service de médecine interne, Monastir, Tunisie
Hyperhomocysteinemia is an emerging factor in arterial disease, which is now taken more seriously by specialists. In the elderly, it makes the post-myocardial recovery precarious and impairs mental functions.
mean age 68.3 years. None of these patients was taking hormone supple-ments or multivitamins.A group of people age and sex matched served as controls.
The study showed that the plasma concentrations of total homocysteine (Hcy) were significantly higher in coronary patients compared to controls 15.9 (15 – 19.25) vs. 12.4ȝmol / L (10.6 – 12.8). Homocysteine was more positively correlated with levels of high-sensitivity CRP: Hs CRP (r=–0705, p=0.00) and negatively correlated with the activity of HDL thiolactonase, a minor compound with a homocysteine thiolactone hydrolase activity limiting the atherogenic effects of homocysteine. A multivariate ana-lysis (Hcy; HTase, CT, TG, Apo A/B ; Hs Crp) showed that only trigly-cerides influenced the severity of coronary artery disease in these patients (p=0.04).
Hyperhomocysteinemia as well as dyslipidaemia found in elderly CVD patients can be mastered by a proper diet and a close monitoring of therapeutic in elderly patients.
250
Hyperhomocysteinemia and severity of cardiovascular disease in type 2 elderly diabetics
Nadia Koubaa (1), Sonia Hammami (2), Raja Chaaba (1), Sounira Mehri (1), Mohamed Hammami (1)
(1) Faculté de Médecine, Biochimie, Monastir, Tunisie – (2) HôpitalFat-touma Bourguiba, Service de médecine interne, Monastir, Tunisie
Diabetics are prone to cardiovascular disease (CVD), in fact, the rate of death due to heart problems is two to four times higher in people with type 2 diabetes. The common risk factors for the accumulation of cholesterol plaques on the walls of the arteries (arteriosclerosis), such as hypertension, obesity, smoking and high blood lipid levels (dyslipidemia), are known to increase the cardiovascular risk in people with diabetes. However, other factors may also be involved namely the excess of an amino acid intermediate in the metabo-lism of methionine: the Homocysteine.
This study involved a total of 53 people grouped in diabetics (n=27, age=68.3 years) and non-diabetics (n=26, age=68.7ans).
A significant higher moderate hyperhomocysteinemia was found in the dia-betic group (15.8 (14.3-17.2) vs 18.5 mmol / L (15.4 – 24.8). P=0.057) asso-ciated with a significant decreased thiolactonase activity ( HTase: 327.4 (240.3 -406.8) vs. 266.8U / L (162-365.2). p=0.058). Moreover, in the diabetic group, the plasma homocysteine levels were negatively correlated with HTase activity (r=–0.637 p <0.00). Also in the group of non-diabetic patients on plasma homocysteine levels were negatively correlated with the HTase activity(r=0759, p<0.00.) and positively with apolipoprotein B (apo B: r=0.862 p <0.05).
A binary regression showed that homocysteine is an important cardio-vascular risk factor in elderly diabetic patients p=0.058 (diet score; WHR; TG, HDL cholesterol Hcy)
The hyperhomocysteinemia found in diabetics is responsible for macro-and micro-vascular diseases leading to death macro-and seems to be an important risk factor in the elderly. As hyperhomocysteinemia can be improved easily and safely with folic acid, a particular attention should be given to the screening and the treatment of hyperhomocysteinemia in elderly patients
251
Polymorphisms of tumor necrosis factor-alpha (TNF-α) gene and myocardial infarction in a Tunisian population
Riadh Jemaa (1), Amani Kallel (1), Mohamed-Hedi Sebai (1), Moncef Feki (1), Mhammed Sami Mourali (3), Rachid Mechmeche (2), Naziha Kaabachi (1)
(1) CHU la Rabta, Biochimie, Tunis, Tunisie – (2) CHU la Rabta, Cardio-logie, Tunis, Tunisie
Background: Tumor necrosis factor alpha (TNF-α) is a primary
pro-inflammatory cytokine and has been implicated in cardiovascular disease pathogenesis. The TNF-α gene may play an important role in coronary heart disease (CHD) and myocardial infarction (MI) risk. Controversial results
regarding the association of polymorphisms of the TNF-α gene with CHD/MI have been reported. The aim of this study was to investigate the possible asso-ciation between the G-308 A (rs1800629), the C-863 A (rs1800630) and the T–1031 C (rs1799964) polymorphisms in the promoter region of the TNF-α gene and MI in a sample of the Tunisian male population. In addition, we also examined the association of TNF-α gene haplotypes with MI in Tunisian male subjects.
Methods: A total of 276 unrelated patients with MI and 287 controls were
included in this study. The SNPs of the TNF-α gene were determined by poly-merase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis. The genotype distribution was in Hardy –Weinberg equilibrium for all variants.
Results: The genotype distribution and the relative allelic frequencies for
the 3 variants investigated were not significantly different between MI and control subjects (P=NS). Moreover, the odds ratio for MI associated with the –308A (OR=1.06, 95% CI 0.78 – 1.44), –863A (OR=1.22, 95% CI 0.83 – 1.81) and –1031C (OR=1.06, 95% CI 0.81 – 1.39) variants failed to reach statistical significance. Haplotype-based analysis also failed to reveal an association between MI and investigated variants.
Conclusion: Our data suggested that these polymorphisms of the TNF-α
gene are unlikely to be major factor in the susceptibility to MI in the Tunisian population studied.
252
The effect of apolipoprotein E and CYP3A4 polymorphisms on the response to lipid-lowering treatment with atorvastatin.
Sihem Mbarki (1), Salme Abdesselem (1), Ameni Kallel (2), Jemaa Riadh (2), Bassem Rekik (1), Mohsen Jemaa (1), Naziha Kaabechi (2), Rachid Mech-meche (1)
(1) CHU la Rabta, Cardiologie, Tunis, Tunisie – (2) CHU la Rabta, Bio-chimie, Tunis, Tunisie
Background: Statins are one of the most commonly prescribed classes of
drug worldwide and therapy is highly effective in reducing low-density lipo-protein cholesterol (LDLc) levels and cardiovascular events. Inspite of these recognized benefits, there is large variability in clinical response to statin treatment. This may be due to the interaction of environmental and genetic factors that affect drug bioavailability, receptor function or ligand structure. Our objective was to assess the effect of 2 polymorphysms in apolipoprotein E (ApoE) and CYP3A4 genes on response to atorvastatin.
Methods: Our study included 60 patients, hospitalized for ACS. DNA was
extracted from leucocytes and analyzed by the PCR-RFLP protocol. Lipid pro-files and calculation of LDLc were performed before and 8 weeks after initi-ation of atorvastatin 40mg/day. We proposed a definition of resistance to atorvastatin (reduction in LDLc<20%) and then looked for correlations between clinical factors, polymorphisms of CYP3A4 and Apo E and the lipid response to atorvastatin 40mg/day.
Results: The average baseline LDLc was 1.13g/l ±0.4; baseline LDL
cholesterol was lower in E4 allele carriers (p=0.06) and G allele carriers (p<0.01). At the end of the follow up period, the absolute reduction of LDLc was –0.35g/l ±0.43 and the relative reduction of –23.90%±31.68. Resistance to atorvastatin was noted in 37.9% of patients. In univariate analysis, this resistance was inversely correlated with baseline LDLc(p<0.01). The G allele (CYP3A4*1G) was associated with poor response to atorvastatin (p<0.01) and the E4 allele (ApoE) was associated with a higher risk of resistance (p=0.01). In multivariate analysis the studied polymorphisms were not independently associated with response to atorvastatin and only the baseline LDLc was predictive of the response and of the resistance to atorvastatin.
Conclusion: This study objectified, in univariate analysis, a correlation
between low baseline LDLc, poor response to atorvastatin and E4 and G alleles. Multivariate analysis, however, calls into question the interest of genetic markers compared to the only baseline rate of LDLc to predict the resistance to atorvastatin.
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Corticotropin-releasing hormone (CRH) is involved in cardiac energy homeostasis
Theodora Tzanavari, Emilia Varela, Katia P. Karalis, Dennis V. Cokkinos
BRFAA, Athens, Grèce
Background: The response of various systems to stressful stimuli is
influ-enced by the HPA axis. We have previously demonstrated that the stress hor-mone corticotropin (CRH), the hypothalamic component of the axis with various peripheral actions, contributes to cardiac function in basal and stressful (LPS-induced) conditions; with Crh–/– mice exhibiting compromised myocar-dial function at basal levels, which further deteriorates upon LPS administration. In this study, given the emerging evidence on the metabolic dysregulation underlying cardiovascular dysfunction and associated disease development, we investigated whether the effects of Crh deficiency are due to metabolic changes.
Methods: Crh+/+ and Crh–/– mice±LPS (n=10/group) were treated with
bezafibrate and metformin. Mice were subjected to 2D M-mode echocardio-graphy. Markers were analyzed by qPCR and western blotting.
Results: Compared to Crh+/+ mice, Crh–/– had reduced expression of
genes involved in lipid and glucose utilization, reminiscent of starving myo-cardium. These included the reduced expression of the β-oxidation markers ACO (80%) and Hadha (52%) and the glycolysis marker HexokinaseII (82%). In addition, PPARα (37%) and its target genes, as well as PPARγ (62%) and its co-activator PGC1α (66%) were downregulated in Crh–/–, as well as AMPKα2 (58%). LPS administration led to further decrease in the expression
levels of these markers. Administration of the PPAR agonist bezafibrate in
Crh–/– mice partially ameliorated the detrimental effects of LPS
administra-tion. Metformin treatment of Crh–/– mice prior to and after LPS led to further improvement of the LPS effects and significant extension of their survival time, via upregulation of AMPK and ACC.
Conclusion: Our results indicate a novel role for CRH in cardiac function
via impaired myocardium FA metabolism. The specific factors mediating the effects of CRH on cardiomyocytes and the relevance of these findings to human conditions are under investigation.
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Intratissular procoagulant microparticles do not contribute to enhanced thrombotic status in diabetes mellitus patients. Role in intraplaque hemorrhage and vulnerability?
Soraya El-Ghannudi (1), Malak Abbas (2), Fatia Zobairi (2), Béatrice Heschler (3), Pierre Mangin (3), Nabil Chakfe (1), Fabien Thaveau (1), Patrick Ohlmann (4), Florence Toti (5), Christian Gachet (3), Olivier Morel (1)
(1) NHC, Pôle d'activité médico-chirurgicale cardiovasculaire, Stras-bourg, France – (2) EA 7293 Stress vasculaire et tissulaire en Transplan-tation, Univeristé de Strasbourg, Strasbourg, France – (3) EFS, Strasbourg, France – (4) Hôpitaux Universitaires de Strasbourg, Fédera-tion de Cardiologie, Strasbourg, France – (5) INSERM 770 et Université de Strasbourg, Strasbourg, France
Background: Recent data have suggested the central importance of the
tissue factor-microparticle-mediated pathway in diabetic thrombophilia and cardiovascular complications. Advanced human atherosclerotic plaques contain large amounts of microparticles (MPs) and MPs isolated from human atherosclerotic lesions are highly thrombogenic. In the present study, we sought to investigate whether intra tissular procoagulant MPs trapped with carotid atheromatous plaque contribute to an enhanced thrombotic propensity in type 2 diabetes mellitus (T2DM) patients
Patients, materiel and methods: 77 consecutive patients undergoing carotid
artery endarterectomy were prospectively enrolled. The population was split in 4 sub-groups according to the diabetes status (T2DM, NTDM) and to the clin-ical presentation (symptomatic (SYM) or not (NSYM). MPs were isolated from the atherosclerotic plaque using serial centrifugation. MPs prothrombinase and tissue factor activities, cellular origins were determined. Plaque thrombogenicity using chamber perfusion model at various shear stress was determined.
Results: Regardless of the diabetic status, high concentrations of procoagulant
MPs mainly of leukocyte and endothelium origins could be detected within the plaque. Prothrombinase and tissue factor activities harbored by MPs appeared to
be in the same range in plaques obtained from T2DM or NT2DM patients. Like-wise, chamber perfusion data failed to evidence enhanced thrombogenic status associated with diabetic plaque. Conversely, high levels of platelet and erythro-cytes MPs could be evidenced within the plaque of SYM T2DM patients.
Conclusion: The present study did not substantiate the view of a specific
increased of thrombotic potential displayed by MPs within carotid atheroscle-rotic plaque in diabetes mellitus patients. Conversely, high levels of platelet and erythrocytes MPs, appear to be a characteristic feature of the diabetic vulnerable plaque. Whereas these MPs are solely a marker of intra plaque haemorrhage or act as true effector involved in matric proteolysis and angio-genesis remain to be established.
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Effects of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) on lamb fetus pulmonary circulation
Dyuti Sharma (1), Estelle Aubry (2), Tavernek Ouk (1), Ali Houeijeh (2), Rémi Besson (3), Véronique Houfflin-Debarge (4), Philippe Deruelle (2), François Godart (2), Laurent Storme (2)
(1) Laboratoire EA 4489, université Lille 2, Lille, France – (2) CHRU Lille, Hôpital Cardiologique, Cardiologie infantile, Lille, France – (3) Hôpital Jeanne de Flandre, Chirurgie pédiatrique, Lille, France – (4) Hôpital Jeanne de Flandres, Clinique d'Obstétrique, Lille, France
Persistant pulmonary hypertension is responsible for significant morbidity and mortality in neonates. We showed that, compared with Intralipid, the Omegaven® (including EPA and DHA) has pulmonary vasodilator properties during the perinatal life.
Objectives: To determine the active agent. We studied the circulatory
effects of EPA and DHA in model of lamb fetus and in vitro in isolated vas-cular rings to clarify the mechanism of action.
Methods: sheep were operated at 128 days of gestation, catheters were
placed in the systemic and pulmonary circulation, Doppler probe around the left pulmonary artery (LPA). Pulmonary artery pressure (PAP) and the LPA flow (LPAF) were measured during the injection of EPA or DHA for 120 minutes, allowing calculation of pulmonary vascular resistance (PVR). Blood gases were taken before and after injection of EPA or DHA. In vitro experiments studied effect-dose of EPA in isolated vascular rings incubated with increasing concentrations after vasoconstriction by agonist of Throm-boxane A2. Vascular rings were separated into two groups E+ (intact endothe-lium) and E- (damaged endotheendothe-lium), according to response to acetylcholine.
Results: EPA induced significant and prolonged drop of 22% in pulmonary
vascular resistance (n=8, p <0.001). DHA did not alter LPAF, or PVR. Heart rate, PAP and arterial blood gases did not change during experiments. Injection of ethanol, their solvent, had no significant effect on LPAF and RVP.
Incubation of EPA caused a maximum relaxation of 60% in group E + (n=7), whereas vessels tone did not change in group E- (n=5, p<0.001). Response to sodium nitroprusside was not different between 2 groups, indi-cating integrity of smooth muscle.
Conclusion: EPA induces a pulmonary vasorelaxation in lamb fetus
without adverse effects. This effect is endothelium- and dose-dependent. These results suggest that EPA should be used in clinical situations associated with pulmonary hypertension in the newborn
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Association of homocysteine thiolactonase activity and PON1 poly-morphisms with the severity of acute coronary syndrome
Mohamed Hammami (1), Ndia Koubaa (1), Sonia Hammami (2), Amel Nakbi (1), Khaldoun Ben Hamda (2), Faouzi Maatouk (2)
(1) Faculté de Médecine de Monastir, Biochimie Biologie Molécualire, Monastir, Tunisie – (2) Hôpital Fattouma Bourguiba, Cardiologie, Monastir, Tunisie
Introduction: Excess of total homocysteine (tHcy) and decrease of
thio-lactonase activities (HTase) have been proposed as risk factors for coronary artery diseases (CAD).
Objectives: We evaluated the relationship of tHcy and HTase with
paraox-onase 1 (PON1) gene polymorphism according to CAD severity.
Design and methods: 118 healthy volunteers and 91 CAD patients were
compared.
Results: Serum levels of tHcy and oxidized LDL (ox-LDL) increased
significantly by 26% and 48% in CAD patients and were associated with significantly lower levels of HDL cholesterol (p= 0.02) and 42% of decrease in HTase activities (pb0.05). In these patients the HTase activity was negatively associated with tHcy and Hs CRP levels (r=−0.622, p=0.00 and r=−0.355, p=0.007 respectively) but positively associated with apoB and triglyceride levels (r=0.35, p=0.042 and r=0.308, p=0.003 res-pectively). HTase activity decreased inversely to the number of affected
vessels and according to PON1 polymorphism. PON1 Q192R RR and PON1 L55M MM genotypes were associated with higher HTase activities. Only PON1 L55M (MM) genotype frequency was significantly higher in CAD patients than in controls (Pb0.05), while its frequency was similar between the two subgroups according to CAD severity. In a multivariate analysis, tHcy levels were the only independent factor affecting the severity of cardiovascular disease (p= 0.029).
Conclusions: High tHcy levels are associated with the severity of
cardio-vascular disease and may be partly explained by the diminished HTase acti-vities in these patients.
Keywords: Homocysteine; Thiolactonase; Coronary artery disease;
