Development of a Genotype 325–Specific proCPU/TAFI ELISA

Gils, Ann; Alessi, Marie‐Christine; Brouwers, Els; Peeters, Miet; Marx, Pauline F.; Leurs, Judith; Bouma, Bonno N.; Hendriks, Dirk; Juhan‐Vague, I.; Declerck, Paul

doi:10.1161/01.atv.0000074145.58172.bd

Cited by 85 publications

(108 citation statements)

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“…[11][12][13][14][15] Alternatively, clot lysis experiments 31 or the use of fluorescently labeled plasminogen 32 may also be used to quantitate TAFI levels or TAFIa activity, respectively. Apart from a previous study 16 demonstrating that the reactivity of a majority of available assays is partially dependent on the Thr 325 polymorphism, it should be realized that interpretation of TAFI antigen levels as determined with noncharacterized immunologic assays is also quite ambiguous because TAFI can occur in different forms (TAFI, TAFIa, TAFIai, fragmentation products of TAFIai, released AP). Therefore, there is currently a strong need for well-characterized immunoassays for the evaluation of the levels of different TAFI forms.…”

Section: Discussionmentioning

confidence: 98%

“…15 A possible explanation for these contradictory results was that the different studies used different methods for TAFI determination. 16 Silveira et al used quantitative activation of the zymogen followed by determination of the total enzymatic activity with a chromogenic assay. Van of TAFI antigen levels, as a putative risk marker for cardiovascular events, has been mainly focused on "total" antigen levels (either by ELISA or by activity assays after full activation of TAFI), reporting levels of 4 to 15 g/mL.…”

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Development of ELISAs Measuring the Extent of TAFI Activation

Ceresa

Brouwers

Peeters

et al. 2006

ATVB

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Objectives-To date, quantitation of TAFI antigen levels has been mainly focused on "total" antigen levels and has been shown to yield ambiguous results because of the existence of different isoforms and various degrees of activation. Our objective was to develop assays that allow measuring the extent of TAFI activation. Methods and Results-A variety of enzyme-linked immunosorbent assays (ELISAs) were evaluated for their preferential reactivity toward TAFI before and after activation, and toward the recombinantly expressed activation peptide. Three ELISAs with distinct reactivities were selected: recognizing either exclusively nonactivated TAFI, the released activation peptide, or exclusively TAFIa (activated TAFI). Evaluation of TAFI activation during clot lysis revealed that decreases of TAFI levels are associated with increases of the released activation peptide and TAFIa levels. In addition, antigenic measurement of TAFIa parallels activity measured by chromogenic assay. Analyzing plasma samples revealed that subjects with hyperlipidemia had significantly higher plasma levels of both the activation peptide (109.2 versus 95.5; PϽ0.001) and TAFIa (112.1 versus 103.3; Pϭ0.03), and not of TAFI antigen (92.5 versus 87.9; Pϭ0.07) (results in % of plasma pooled from normolipidemic subjects). Conclusion-ELISAs that allow to measure the extent of TAFI activation were developed. These ELISAs constitute more sensitive markers in studies on the relationship between TAFI and cardiovascular diseases. ; 36 kDa). Subsequently, TAFIa is inactivated through a conformational change to an inactive form (TAFIai), followed by proteolytic cleavage resulting in fragments of 25 and 11 kDa. 5 TAFIa exerts an antifibrinolytic effect by removing C-terminal lysine residues from fibrin, resulting in a decreased plasmin formation and a retardation of clot lysis. 6 TAFIa is highly unstable in vitro (half-life at 37°C between 8 to 15 minutes). [7][8][9][10] Different clinical studies have investigated the possible relationship between TAFI and cardiovascular events. A positive correlation was found between TAFI levels and the risk for coronary artery disease, 11 venous thrombosis, 12 angina pectoris, 13 and ischemic stroke. 14 However, another study revealed that elevated TAFI may be protective against myocardial infarction. 15 A possible explanation for these contradictory results was that the different studies used different methods for TAFI determination. 16 Silveira et al used quantitative activation of the zymogen followed by determination of the total enzymatic activity with a chromogenic assay. Van of TAFI antigen levels, as a putative risk marker for cardiovascular events, has been mainly focused on "total" antigen levels (either by ELISA or by activity assays after full activation of TAFI), reporting levels of 4 to 15 g/mL. 16,20 -22 However, it should be stressed that in most of these studies the used immunologic assays were not validated with respect to putative differential reactivities toward different isoforms and fragments...

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Development of ELISAs Measuring the Extent of TAFI Activation

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Brouwers

Peeters

et al. 2006

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“…Several different antigen assays as well as distinct activity assays have been used in epidemiological studies. With the discovery that some enzyme linked immunosorbent assays of TAFI antigen are dependent on the 1040C/T polymorphism, revealing a lack of reactivity with the 1040T variant, 21 even more caution is required in the interpretation of results.…”

Section: Introductionmentioning

confidence: 99%

Low thrombin activatable fibrinolysis inhibitor activity levels are associated with an increased risk of a first myocardial infarction in men

Meltzer¹,

Doggen²,

Groot³

et al. 2009

Haematologica

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BackgroundStudies on the relation between thrombin activatable fibrinolysis inhibitor (TAFI) and arterial thrombosis have produced conflicting results.TAFI regulates fibrinolysis, but other roles of this inhibitor, including anti-inflammatory properties, have also been demonstrated. Design and MethodsWe investigated the association between TAFI activity and the risk of myocardial infarction. Additionally, we studied the association of common single nucleotide polymorphisms in the TAFI gene with levels of the TAFI protein and risk of myocardial infarction.We included 554 men under 70 years old with a first myocardial infarction and 643 controls participating in the Study of Myocardial Infarctions Leiden (SMILE), a case-control study. ResultsWe found odds ratios (95% confidence intervals) of a first myocardial infarction of 2.4 (1.6-3.6), 3.2 (2.1-4.7) and 3.4 (2.3-5.1) for subjects whose TAFI levels were in the third, second and first quartiles (lowest TAFI levels), respectively, compared with the fourth quartile, after adjusting for arterial disease risk factors. The rare -438A and 1040T alleles were associated with lower, and the rare 505G allele with higher TAFI levels than the common alleles. Carriers of the -438A allele had an increased risk of myocardial infarction (odds ratio 1.6 (1.0-2.5) for AA; odds ratio 1.2 (0.9-1.5) for AG compared with GG). The other single nucleotide polymorphisms were not associated with myocardial infarction. ConclusionsLow TAFI activity levels are associated with increased risk of a first myocardial infarction in men. The results on the association between TAFI single nucleotide polymorphisms and myocardial infarction were inconsistent.Key words: TAFI, myocardial infarction, genetics, epidemiology.Citation: Meltzer ME, Doggen CJM, de Groot PG, Meijers JCM, Rosendaal FR, and Lisman T. Low thrombin activatable fibrinolysis inhibitor activity levels are associated with an increased risk of a first myocardial infarction in men. Haematologica 2009; 94:811-818. doi:10.3324/haematol.2008 This is an open-access paper. Low thrombin activatable fibrinolysis inhibitor activity levels are associated with an increased risk of a first myocardial infarction in men

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“…10 -12 Furthermore, a combined segregation-linkage analysis suggested the existence of 2 quantitative trait loci that would explain 78% of the variance in plasma TAFI Ag. 13 However, it was demonstrated that some ELISAs used for TAFI Ag determination had different assay sensitivity between isoforms, 14,15 leading to overestimations of the effects associated with TAFI gene polymorphisms. Results from recent studies using genotype-independent assays have confirmed a genetic effect although of lesser magnitude.…”

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Thrombin Activatable Fibrinolysis Inhibitor Activation Peptide Shows Association With All Major Subtypes of Ischemic Stroke and With TAFI Gene Variation

Ladenvall

Gils

Jood

et al. 2007

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Objective-Thrombin activatable fibrinolysis inhibitor (TAFI) attenuates fibrinolysis. The aim of the present study was to investigate the possible association between TAFI and overall ischemic stroke and ischemic stroke subtypes. Methods and Results-The Sahlgrenska Academy Study on Ischemic Stroke (SAHLSIS) comprises 600 cases (18 to 69 years) and 600 matched population controls. Stroke subtype was defined by the Trial of Org 10172 in Acute Stroke Treatment (TOAST) classification. TAFI was investigated at the protein level, by analyzing plasma levels of intact TAFI and released activation peptide [AP], and at the genetic level, by genotyping a selection of eleven single nucleotide polymorphisms. After adjustment for traditional risk factors, both TAFI measurements showed association with overall ischemic stroke (AP: odds ratio, 2.22; 95% confidence interval, 1.89 to 2.61; intact TAFI: odds ratio, 1.21; 95% confidence interval, 1.06 to 1.38; for 1-SD increase in AP and intact TAFI, respectively). AP showed associations with all 4 major subtypes of ischemic stroke and intact TAFI to large vessel disease and cryptogenic stroke. TAFI genotypes and haplotypes showed significant associations with both TAFI measurements. In contrast, no association was observed between genetic variants and overall ischemic stroke. Conclusion-TAFI levels show independent association with overall ischemic stroke. This association is stronger for released AP than for intact TAFI, and for released AP, it is present in all ischemic stroke subtypes. Key Words: thrombin activatable fibrinolysis inhibitor Ⅲ polymorphism Ⅲ genetics Ⅲ ischemic stroke subtype T hrombin activatable fibrinolysis inhibitor (TAFI), also denoted procarboxypeptidase B 1 and procarboxypeptidase U, 2 is a zymogen present in human plasma. It can be activated by trypsin-like enzymes such as thrombin, plasmin, or the thrombin/thrombomodulin complex. [3][4][5] On activation of intact TAFI, the activation peptide (AP) (Phe1-Arg92; 20 kDa) is released from the catalytic domain (TAFIa) (Ala93-Val401; 36 kDa). 4 Both in vitro and in vivo experiments show that TAFIa retards fibrinolysis. 6 TAFIa operates by continuously removing C-terminal lysine residues on plasmin-modified partially degraded fibrin, thus attenuating the rate of plasminogen activation and fibrinolysis. 7 Because thrombin and thrombin/thrombomodulin complex can activate TAFI, and because TAFIa suppresses fibrinolysis, this pathway has been proposed to be a molecular link between coagulation and fibrinolysis.The TAFI gene (named CPB2) maps to chromosome 13q14.11, spans approximately 48 kb and contains 11 exons. 8 A number of TAFI gene polymorphisms have been identified, 2 of which result in amino acid substitutions, Thr147Ala and Thr325Ile. The Ile325 variant was shown to have a longer half-life and increased antifibrinolytic properties compared with the 325Thr variant. 9 Initial studies reported association between several TAFI gene single nucleotide polymorphisms (SNPs) and circulating levels of TAFI antigen (TA...

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Development of a Genotype 325–Specific proCPU/TAFI ELISA

Cited by 85 publications

References 32 publications

Development of ELISAs Measuring the Extent of TAFI Activation

Development of ELISAs Measuring the Extent of TAFI Activation

Low thrombin activatable fibrinolysis inhibitor activity levels are associated with an increased risk of a first myocardial infarction in men

Thrombin Activatable Fibrinolysis Inhibitor Activation Peptide Shows Association With All Major Subtypes of Ischemic Stroke and With TAFI Gene Variation

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