Angela M. Carter

Rheumatoid arthritis association at 6q23

The Wellcome Trust Case Control Consortium (WTCCC) identified nine single SNPs putatively associated with rheumatoid arthritis at P = 1 × 10−5 − 5 × 10−7 in a genome-wide association screen. One, rs6920220, was unequivocally replicated (trend P = 1.1 × 10−8) in a validation study, as described here. This SNP maps to 6q23, between the genes oligodendrocyte lineage transcription factor 3 (OLIG3) and tumor necrosis factor-α–induced protein 3 (TNFAIP3).

Identification, classification, and expression of RAGE gene splice variants

The receptor for advanced glycation end‐products (RAGE) is a single‐transmembrane, mul tiligand receptor of the immunoglobulin superfamily. RAGE up‐regulation is implicated in numerous patho logical states including vascular disease, diabetes, can cer, and neurodegeneration. The understanding of the regulation of RAGE is important in both disease patho genesis and normal homeostasis. Here, we demonstrate the characterization and identification of human RAGE splice variants by analysis of RAGE cDNA from tissue and cells. We identified a vast range of splice forms that lead to changes in the protein coding region of RAGE, which we have classified according to the Human Gene Nomenclature Committee (HGNC). These resulted in protein changes in the ligand‐binding domain of RAGE or the removal of the transmembrane domain and cytosolic tail. Analysis of splice variants for premature termination codons reveals~50% of identified variants are targeted to the nonsense‐mediated mRNA decay pathway. Expression analysis revealed the RAGE_v1 variant to be the primary secreted soluble isoform of RAGE. Taken together, identification of functional splice variants of RAGE underscores the biological diversity of the RAGE gene and will aid in the under standing of the gene in the normal and pathological state.—Hudson, B. I., Carter, A. M., Harja, E., Kalea, A. Z., Arriero, M., Yang, H., Grant, P. J., Schmidt, A. M. Identification, classification, and expression of RAGE gene splice variants. FASEB J. 22, 1572–1580 (2008)

Association between polymorphisms in the Clock gene, obesity and the metabolic syndrome in man.

Objective:Accumulating evidence raises the hypothesis that dysregulation of intrinsic clock mechanisms are involved in the development of the metabolic syndrome, type 2 diabetes mellitus and cardiovascular disease. The aim of the present study was to investigate the relationship between three known common polymorphisms in the Clock gene and features of the metabolic syndrome in man.Methods:Genotype and haplotype analysis was carried out in a cohort of 537 individuals from 89 families characterized for inflammatory, atherothrombotic and metabolic risk associated with insulin resistance.Results:Heritability of the metabolic syndrome, defined according to International Diabetes Federation criteria, was 0.40. Haplotype analysis indicated three common haplotypes: CAT, TGT and CGC (rs4864548-rs3736544-rs1801260) with frequencies of 31, 33 and 28%, respectively. The CGC haplotype was less prevalent in subjects with the metabolic syndrome (P=0.0015) and was associated with lower waist circumference (P=0.007), lower hip circumference (P=0.023), lower body mass index (P=0.043) and lower leptin levels (P=0.028). The CAT haplotype was significantly associated with the presence of the metabolic syndrome (P=0.020).Conclusions:These findings suggest that the Clock gene CGC haplotype may be protective for the development of obesity and support the hypothesis that genetic variation in the Clock gene may play a role in the development of the metabolic syndrome, type 2 diabetes and cardiovascular disease.

Genetic regulation of fibrin structure and function: complex gene-environment interactions may modulate vascular risk

BACKGROUND Polymorphisms in the fibrinogen and factor XIII genes are associated with atherothrombotic risk, but clinical studies have produced inconsistent results and laboratory studies have not explained these findings. We aimed to investigate interactions between polymorphisms in the factor XIII and fibrinogen genes, fibrinogen concentrations, and other cardiovascular risk factors in relation to fibrin structure and function. METHODS We used permeation analysis and electron microscopy to investigate interactions between fibrin structure, factor XIII Val34Leu, fibrinogen Aalpha Thr312Ala, fibrinogen Bbeta Arg448Lys, and fibrinogen concentrations in plasma and purified systems. FINDINGS Increased fibrinogen concentrations were associated with decreases in permeability, with tighter clot structures in the presence of factor XIII 34Val alleles compared with those in the presence of 34Leu alleles. Findings were confirmed by scanning electron microscopy of fibrin. Similar changes in permeability were noted for Aalpha fibrinogen 312Ala compared with that for 312Thr. INTERPRETATION Our results show interactions between coding polymorphisms in fibrinogen and factor XIII and fibrinogen concentrations that modify fibrin and explain the apparent paradox between epidemiological studies of factor XIII 34Leu and reported in-vitro effects on fibrin structure and function. We suggest a potential complexity of gene-gene and gene-environment interactions in determining cardiovascular risk.

Association of the Platelet PlA Polymorphism of Glycoprotein IIb/IIIa and the Fibrinogen Bβ 448 Polymorphism With Myocardial Infarction and Extent of Coronary Artery Disease

Background Platelets and fibrinogen play an integral role in the development of thrombosis and are implicated in the process of atherosclerosis. The fibrinogen Bβ 448 polymorphism and the PlA polymorphism of platelet glycoprotein IIIa are reported to be independently associated with coronary artery disease. The aim of this study was to determine the association of the fibrinogen Bβ 448 and the platelet glycoprotein IIIa PlA polymorphisms in relation to extent of coronary atheroma as characterized by angiography and a past history of myocardial infarction (MI) as assessed by World Health Organization criteria. Methods and Results Caucasian patients (n=405) admitted for routine angiography for investigation of chest pain or suspected coronary artery disease were recruited. Caucasian control subjects (n=216) were recruited from local Family Health Services Authority general practice registers. Fibrinogen levels were higher (P=.04) in male patients (3.24 g/L; CI, 3.14 to 3.35) than male control subjects (3.06...

Platelet GP IIIa PlA and GP Ib Variable Number Tandem Repeat Polymorphisms and Markers of Platelet Activation in Acute Stroke

A number of polymorphisms of the platelet glycoprotein (GP) Ib-V-IX and IIb/IIIa complexes have been described, and the PlA polymorphism of GP IIIa has been associated with coronary thrombosis. We determined the levels of beta-thromboglobulin (beta-TG) and platelet factor 4 (PF4) and the genotype distributions of PlA and a variable number tandem repeat (VNTR) polymorphism of GP 1b in subjects with acute stroke (n=609) and healthy control subjects (n=435). Levels of beta-TG were higher in patients both initially (47.4 [44.7 to 50.2] ng/mL, P<0.0001) and after 3 months (42.9 [40.3 to 45.7] ng/mL, P=0.03) compared with control subjects (39.4 [37.7 to 41.2] ng/mL). Initial levels of beta-TG were significantly higher in those who subsequently died (58.7 [52.3 to 65.8] ng/mL) compared with those still alive (42.7 [40.1 to 45.5] ng/mL, P<0.0001). In a logistic regression model, beta-TG remained an independent predictor of poststroke mortality, with an odds ratio for an increase in 10 ng/mL of 1.12 (1.03 to 1.21, P=0.006). In subjects who had never smoked, there was a significant difference in the genotype distributions of patients with atherothrombotic stroke (A1/A1=147, A1/A2=70, and A2/A2=2) compared with controls (A1/A1=165, A1/A2=47, and A2/A2=5, P=0.03). The PlA distribution of subjects with atherothrombotic stroke before the age of 50 years (A1/A1=19 and A1/A2+A2/A2=18) was also significantly different from age- and sex-matched controls (A1/A1=54 and A1/A2+A2/A2=20, P=0.02). We found no association of VNTR with stroke or poststroke mortality. These data indicate that there is a persistent state of enhanced platelet activation in subjects with acute stroke, which is associated with poststroke mortality. The increased frequency of the PlA2 allele in young subjects with atherothrombotic stroke lends further support for a role of the PlA polymorphism in acute thrombosis.

Factor XIII Val 34 Leu A Novel Association With Primary Intracerebral Hemorrhage

BACKGROUND AND PURPOSE A common G-to-T point mutation (Val 34 Leu) in exon 2 of the alpha-subunit of the factor XIII is strongly negatively associated with the development of myocardial infarction. This result suggests that factor XIII Val 34 Leu is interfering with the formation of cross-linked fibrin. The role of factor XIII Val 34 Leu in the pathogenesis of cerebral infarction and primary intracerebral hemorrhage is unknown. METHODS Six hundred twelve patients with acute stroke, defined by World Health Organization criteria and cranial CT, and 436 age-matched control subjects free of cerebrovascular disease were genotyped for the factor XIII Val 34 Leu mutation. Venous blood was drawn for the determination of hemostatic variables and lipids. Factor XIII genotype was determined through a single-stranded conformational polymorphism technique and plasminogen activator inhibitor (PAI)-1 4G/5G promoter genotype by allele-specific polymerase chain reaction. RESULTS The mutation was more frequent in patients with primary intracerebral hemorrhage (n=62) (54.8%; P=.05) than in control subjects (41.7%) or in patients with cerebral infarction (n=529) (46.5%; P=.22). There was no relationship between PAI-1 levels and the PAI-1 4G/5G genotype. CONCLUSIONS There was a slightly higher incidence of factor XIII Val 34 Leu in patients with PICH. This may be related to impaired cross-linking of fibrin and/or coagulation proteins.

Factor V Leiden Gene Mutation and Thrombin Generation in Relation to the Development of Acute Stroke

To determine the prevalence of the factor V Leiden gene mutation in relation to the phenotypes of cerebral infarction and cerebral hemorrhage, we studied 386 randomly selected cases of acute stroke and 247 control subjects. Factor V genotype was determined by amplification of a 267-bp sequence of exon/intron 10 of the factor V gene. Levels of prothrombin fragment F(1 + 2), a marker of thrombin generation, were determined in both acute and convalescent stroke and related to factor V genotype. Prothrombin fragment F(1 + 2) was assessed by using an enzyme-linked immunosorbent assay. Sixteen stroke cases (4.1%) were identified as having the mutation compared with 14 (5.6%) control subjects. Prothrombin fragment F(1 + 2) levels were estimated in 191 cases and found to be elevated both acutely and after 3 months, but they were not related to factor V genotype. Prothrombin fragment F(1 + 2) is elevated in acute stroke and requires further evaluation in relation to cerebrovascular disease. These results suggest that the factor V Leiden gene mutation is not a risk factor for arterial thrombosis causing stroke.

Angiotensin-Converting Enzyme Insertion/Deletion Polymorphism and Cerebrovascular Disease

BACKGROUND AND PURPOSE There is evidence that an allelic variation in the angiotensin-converting enzyme (ACE) gene may confer an increased risk of vascular disease. The roles of the ACE insertion/deletion polymorphism and circulating ACE levels are unknown in cerebrovascular disease. METHODS We studied an insertion/deletion polymorphism within intron 16 of the ACE gene by polymerase chain reaction and plasma ACE activity in 467 cases of stroke, the pathological type of which was established by cranial CT, and 231 control subjects. ACE genotype and activity were related to stroke type and mortality at 4 weeks and 3 months. RESULTS No difference in genotype frequency was observed between all subjects with stroke and control subjects or between control subjects and subjects with cerebral infarction or cerebral hemorrhage. Plasma ACE activity was significantly lower in stroke patients at presentation (64.1 IU/L) than in control subjects (79.6 IU/L; P<.0001). Twenty-one patients (4.5%) with cerebral infarction died within 4 weeks and 56 patients (12%) within 3 months. These patients had significantly lower plasma ACE activity than patients who survived. There was some evidence that risk of death within 4 weeks increased with the number of D alleles (P=.02). Among survivors, plasma ACE activity showed a mean increase of 6.9 IU/L (95% confidence interval, 3.0 to 10.8) between levels at presentation and at 3 months (73.6 IU/L), the latter being similar to ACE activity in control subjects. CONCLUSIONS Low ACE activity at sroke presentation and possession of the D allele may be associated with increased risk of early death from acute cerebral infarction.

Heritability of Clot Formation, Morphology, and Lysis: The EuroCLOT Study

Objective—The relative balance between clot formation and fibrinolysis is considered to reflect thrombotic potential following vascular injury. The aims of the present study were to (1) to determine the contribution of genetic and environmental factors to variance in measures of clot structure/function in the Leeds Family Study, and (2) to determine the relationship between measures of clot structure/function and cardiovascular risk. Methods and Results—Using high throughput turbidimetric assays, heritabilities of measures of clot formation, clot structure, and clot lysis were ≈0.30. Fibrinogen contributed to variance in all measures and plasminogen activator inhibitor-1 to variance in lysis variables. Subjects at increased cardiovascular risk due to the presence of the metabolic syndrome (MetS) had increased clot density (MaxAbsC: 0.358 [0.340, 0.375]au) and prolonged lysis times (LysT: 510 [6569, 7939]s) compared with those without MetS (MaxAbsC: 0.319 [0.310, 0.328]au, P=0.003; LysT: 7221 [4884, 5328]s, P<0.001). Furthermore, measures of clot structure/function increased progressively with increasing number of MetS components. Conclusions—This study indicates that genetic factors contribute modestly to variance in clot structure/function and that clot structure/function is related to presence of the MetS and number of MetS components. Identification of the genetic and environmental factors influencing clot structure/function may further our understanding of the underlying factors predisposing to cardiovascular disease.