-
Journal of Thrombosis and Haemostasis :... Aug 2016Essentials We investigated the molecular base of antithrombin deficiency in cases without SERPINC1 defects. 27% of cases presented hypoglycosylation, transient in 62%...
UNLABELLED
Essentials We investigated the molecular base of antithrombin deficiency in cases without SERPINC1 defects. 27% of cases presented hypoglycosylation, transient in 62% and not restricted to antithrombin. Variations in genes involved in N-glycosylation underline this phenotype. These results support a new form of thrombophilia. Click here to listen to Dr Huntington's perspective on thrombin inhibition by the serpins
SUMMARY
Background Since the discovery of antithrombin deficiency, 50 years ago, few new thrombophilic defects have been identified, all with weaker risk of thrombosis than antithrombin deficiency. Objective To identify new thrombophilic mechanisms. Patients/methods We studied 30 patients with antithrombin deficiency but no defects in the gene encoding this key anticoagulant (SERPINC1). Results A high proportion of these patients (8/30: 27%) had increased hypoglycosylated forms of antithrombin. All N-glycoproteins tested in these patients (α1-antitrypsin, FXI and transferrin) had electrophoretic, HPLC and Q-TOF patterns indistinguishable from those of the congenital disorders of glycosylation (rare recessive multisystem disorders). However, all except one had no mental disability. Moreover, intermittent antithrombin deficiency and hypoglycosylation was recorded in five out of these eight patients, all associated with moderate alcohol intake. Genetic analysis, including whole exome sequencing, revealed mutations in different genes involved in the N-glycosylation pathway. Conclusions Our study provides substantial and novel mechanistic insights into two disease processes, with potential implications for diagnosis and clinical care. An aberrant N-glycosylation causing a recessive or transient antithrombin deficiency is a new form of thrombophilia. Our data suggest that congenital disorders of glycosylation are probably underestimated, especially in cases with thrombosis as the main or only clinical manifestation.
Topics: Adolescent; Adult; Aged; Antibodies; Anticoagulants; Antithrombin III; Antithrombins; Chromatography, High Pressure Liquid; Exome; Female; Genetic Variation; Genotype; Glycoproteins; Glycosylation; Humans; Male; Middle Aged; Mutation; Spain; Thrombophilia; Thrombosis; Young Adult
PubMed: 27214821
DOI: 10.1111/jth.13372 -
The Journal of Thoracic and... Jun 2017
Topics: Antithrombin III; Extracorporeal Membrane Oxygenation; Heart Failure; Humans; Liver Failure, Acute
PubMed: 28283240
DOI: 10.1016/j.jtcvs.2017.02.002 -
Polish Archives of Internal Medicine Oct 2020Antithrombin is a key endogenous anticoagulant that also plays other roles in inflammation, immunity, and other processes. Congenital antithrombin deficiency is the most...
Antithrombin is a key endogenous anticoagulant that also plays other roles in inflammation, immunity, and other processes. Congenital antithrombin deficiency is the most severe type of thrombophilia, yet characterized by a remarkable clinical heterogeneity. Here, as a primer for internists, we present a practical review of data regarding this disorder, focused on its molecular basis, diagnostic procedures, prognostic implications, and clinical management of patients suffering from this severe, and probably underdiagnosed, type of thrombophilia.
Topics: Anticoagulants; Antithrombin III; Antithrombin III Deficiency; Antithrombins; Humans; Thrombophilia
PubMed: 32426958
DOI: 10.20452/pamw.15371 -
The Journal of Biological Chemistry May 2006We previously showed that conformational activation of the anticoagulant serpin, antithrombin, by heparin generates new exosites in strand 3 of beta-sheet C, which...
Residues Tyr253 and Glu255 in strand 3 of beta-sheet C of antithrombin are key determinants of an exosite made accessible by heparin activation to promote rapid inhibition of factors Xa and IXa.
We previously showed that conformational activation of the anticoagulant serpin, antithrombin, by heparin generates new exosites in strand 3 of beta-sheet C, which promote the reaction of the inhibitor with the target proteases, factor Xa and factor IXa. To determine which residues comprise the exosites, we mutated strand 3C residues that are conserved in all vertebrate antithrombins. Combined mutations of the three conserved surface-accessible residues, Tyr253,Glu255, and Lys257, or of just Tyr253 and Glu255, but not any of these residues alone, was sufficient to reproduce the exosite defects of a strand 3C antithrombin-alpha1-proteinase inhibitor chimera in reactions of the heparin-activated variants with both factor Xa and factor IXa. Importantly, the exosite-defective antithrombins bound heparin with nearly wild-type affinities, and the heparin-activated mutants showed near normal reactivities with thrombin, a protease that does not utilize the exosite. Mutation of the conserved but partially buried strand 3C residue, Gln254, the reactive loop P6' residue, Arg399, which interacts with Glu255, or a residue proposed to constitute the exosite from modeling studies, Glu237, all produced minimal effects on antithrombin reactivity with thrombin, factor Xa, and factor IXa in the absence or presence of heparin. Together, these results indicate that Tyr253 and Glu255 are key exosite determinants responsible for promoting the reactions of conformationally activated antithrombin with both factor Xa and factor IXa.
Topics: Amino Acid Sequence; Antithrombin III; Binding Sites; Factor IXa; Factor Xa; Factor Xa Inhibitors; Glutamic Acid; Heparin; Humans; Kinetics; Models, Molecular; Mutation; Protein Binding; Protein Structure, Secondary; Thrombin; Tyrosine
PubMed: 16517611
DOI: 10.1074/jbc.M600415200 -
American Journal of Hematology Jul 1998Various hemostatic and vascular endothelial cell markers were measured in patients with disseminated intravascular coagulation (DIC), non-DIC, or thrombotic...
Various hemostatic and vascular endothelial cell markers were measured in patients with disseminated intravascular coagulation (DIC), non-DIC, or thrombotic thrombocytopenic purpura (TTP) and in healthy volunteers to examine the relationships between the hemostatic abnormalities or vascular endothelial cell injuries and the patients' outcomes. Although the plasma levels of soluble fibrin monomer, thrombin-antithrombin complex, plasmin-plasmin inhibitor complex, and D-dimer were significantly increased in the DIC patients, there were no significant differences in these markers between the DIC patients who survived and those who died, suggesting that these markers might not be directly related to the patient outcome. The plasma thrombomodulin (TM) levels in the DIC and TTP patients were significantly higher than those in the healthy volunteers, and the plasma TM levels in the patients who died were significantly higher than those in the patients who survived. These findings showed that the TM level reflected the outcome, and that the outcome of the diseases underlying DIC and TTP might depend on vascular endothelial cell injuries. The plasma protein C and antithrombin activities were markedly reduced in the DIC, non-DIC, and TTP patients who died compared to those who survived. These findings suggest that reduced plasma antithrombin and protein C activities are useful markers of systemic vascular endothelial injuries. Although the plasma tissue factor (TF) levels were significantly increased in the DIC patients, there was no significant difference in the plasma TF levels between the DIC patients who died and those who survived. In conclusion, we found that the outcome of the diseases underlying DIC and TTP is related to vascular endothelial cells, and that plasma TM, antithrombin, and protein C are useful markers for systemic vascular endothelial cell injury.
Topics: Antifibrinolytic Agents; Antithrombin III; Antithrombins; Disseminated Intravascular Coagulation; Endothelium, Vascular; Fibrin Fibrinogen Degradation Products; Fibrinolysin; Fibrinolytic Agents; Hemolytic-Uremic Syndrome; Humans; Partial Thromboplastin Time; Peptide Hydrolases; Protein C; Prothrombin Time; Purpura, Thrombotic Thrombocytopenic; Survival Rate; Thrombomodulin; alpha-2-Antiplasmin
PubMed: 9662269
DOI: 10.1002/(sici)1096-8652(199807)58:3<189::aid-ajh5>3.0.co;2-n -
Glycobiology Apr 2021Heparan sulfate (HS) is a heterogeneous, extracellular glycan that interacts with proteins and other molecules affecting many biological processes. The specific binding...
Heparan sulfate (HS) is a heterogeneous, extracellular glycan that interacts with proteins and other molecules affecting many biological processes. The specific binding motifs of HS interactions are of interest, but have not been extensively characterized. Glycan microarrays are valuable tools that can be used to probe the interactions between glycans and their ligands while relying on relatively small amounts of samples. Recently, chemoenzymatic synthesis of HS has been employed to produce specific HS structures that can otherwise be difficult to produce. In this study, a microarray of diverse chemoenzymatically synthesized HS structures was developed and HS interactions were characterized. Fluorescently labeled antithrombin III (AT) and fibroblast growth factor-2 (FGF2) were screened against 95 different HS structures under three different printing concentrations to confirm the utility of this microarray. Specific sulfation patterns were found to be important for binding to these proteins and results are consistent with previous specificity studies. Furthermore, the binding affinities (KD,surf) of AT and FGF2 to multiple HS structures were determined using a microarray technique and is consistent with previous reports. Lastly, the 95-compound HS microarray was used to determine the distinct binding profiles for interleukin 12 and platelet factor 4. This technique is ideal for rapid expansion and will be pivotal to the high-throughput characterization of biologically important structure/function relationships.
Topics: Antithrombin III; Binding Sites; Carbohydrate Conformation; Carbohydrate Sequence; Fibroblast Growth Factor 2; Heparitin Sulfate; Humans; Microarray Analysis
PubMed: 32681173
DOI: 10.1093/glycob/cwaa068 -
Scientific Reports Oct 2023Antithrombin (AT) deficiency increases the risk for venous thromboembolism, therefore, a highly sensitive assay to identify this condition is crucial. The aim of this... (Meta-Analysis)
Meta-Analysis
Antithrombin (AT) deficiency increases the risk for venous thromboembolism, therefore, a highly sensitive assay to identify this condition is crucial. The aim of this paper was to perform a meta-analysis comparing AT activities measured by different AT activity assays in patients with heparin binding site AT deficiency. In addition, the diagnostic sensitivity of selected assays was compared depending on the available data. An extensive literature search was performed considering results with publication date up to July 10, 2021. Seven relevant English-language observational studies, comparing AT activity measured by different AT activity assays in Caucasian Europeans with either the AT Budapest III or AT Padua I mutation were included in meta-analyses. There was no significant difference in AT activity between Labexpert and Innovance in patients with AT Budapest III (P = 0.567) and AT Padua I (P = 0.265), while AT activity determined by HemosIL was significantly higher compared to Innovance for both mutations (AT Budapest III: P < 0.001; AT Padua I: P < 0.001). These results are in line with the results of comparison of diagnostic sensitivity. In patients with AT Budapest III, the AT activity was also higher when measured with Berichrom compared to Innovance (P = 0.002), however, the results of comparison of diagnostic sensitivity across studies were variable. No significant difference (P = 0.117) in AT activity as well as diagnostic sensitivity was observed between Sta-Stachrom and Innovance. The results of our study suggest that Innovance, Labexpert and Sta-Stachrom are the most sensitive activity assays for detection of AT Budapest III and AT Padua I, whereas HemosIL showed considerably lower sensitivity for these two variants. As revealed in our study, the diagnostic sensitivity of AT activity assays to type II heparin binding site AT deficiency is different, and in some assays mutation dependent.
Topics: Humans; Heparin; Anticoagulants; Blood Coagulation Tests; Antithrombin III Deficiency; Binding Sites; Antithrombins
PubMed: 37794095
DOI: 10.1038/s41598-023-43941-x -
Proceedings of the National Academy of... Jul 1979Heparin preparations from pig intestinal mucosa and from bovine lung were separated by chromatography on antithrombin-Sepharose into a high-affinity fraction (with high...
Heparin preparations from pig intestinal mucosa and from bovine lung were separated by chromatography on antithrombin-Sepharose into a high-affinity fraction (with high anticoagulant activity) and a low-affinity fraction (with low anticoagulant). Antithrombin-binding heparin fragments (12-16 monosaccharide units) were prepared, either by digesting a high-affinity heparin-antithrombin complex with bacterial heparinase or by partial deaminative cleavage of the unfractionated polysaccharide with nitrous acid followed by affinity chromatography on immobilized antithrombin. Compositional analysis based on separation and identification of deamination products reduced with sodium boro[3H]hydride showed that nonsulfated L-iduronic acid occurred in larger amounts in high-affinity heparin than in low-affinity heparin; furthermore, this component was concentrated in the antithrombin-binding regions of the high-affinity heparin molecules, amounting to approximately one residue per binding site. It is suggested that nonsulfated L-iduronic acid is essential for the anticoagulant activity of heparin. The location of the non-sulfated uronic acid in the antithrombin-binding site was determined by periodate oxidation of antithrombin-binding fragments containing a terminal 2,5-anhydro-D-[1-3H]mannitol unit. Tentative structures for antithrombin-binding sequences in heparin are proposed, including some structural variants believed to be compatible with, but not required for, activity.
Topics: Animals; Antithrombin III; Binding Sites; Carbohydrate Sequence; Cattle; Heparin; Iduronic Acid; Intestinal Mucosa; Lung; Periodic Acid; Swine
PubMed: 226960
DOI: 10.1073/pnas.76.7.3198 -
Ethiopian Journal of Health Sciences Oct 2015One of the rare causes of venous thromboembolism in pregnancy is antithrombin III deficiency. Antithrombin III deficiency is estimated to carry a 30% risk of venous...
BACKGROUND
One of the rare causes of venous thromboembolism in pregnancy is antithrombin III deficiency. Antithrombin III deficiency is estimated to carry a 30% risk of venous thrombotic complication during each pregnancy and postpartum.
CASE DETAILS
We present thea case of a A 21-year-old pregnant woman (Para 1+) with a history of large atrial septal defect repair at our hospital (Imam Ali Hospital, 2 May 2014). The patient, with unknown history of antithrombin III deficiency, was admitted at our emergency center with dyspnea and chest pain for the rule out of tamponade. She presented with a right atrial thrombosis in the second trimester of pregnancy despite the use of therapeutic doses of heparin and warfarin in the postoperative period as thromboembolic prophylaxis. The risk of warfarin emberyopaty led to termination of pregnancy, and successful redo-cardiac surgery outcome was achieved with the combined use of therapeutic anticoagulation and regular plasma-derived antithrombin concentrate infusions to normalize her antithrombin levels.
CONCLUSION
She recovered from the operation uneventfully, and wad discharged in the 12(th) postoperative day. In the 6(th) month of follow-up, antithrombin III increased to 70% in more stable level and transethoracic echocardiography showed no recurrence of right atrial thrombus formation. This case leads to further debate regarding whether full anticoagulation should be a worthy preventive measure for venous thromboembolic prophylaxis after an open heart surgery complicated by pregnancy in a women with inherited antithrombin III deficiency. This point may become more relevant as further experience is gained with the use of recombinant human antithrombin in known cases during open cardiac surgery.
Topics: Adult; Anticoagulants; Antithrombin III; Antithrombin III Deficiency; Antithrombins; Cardiac Surgical Procedures; Female; Heart Atria; Heart Septal Defects, Atrial; Humans; Postoperative Complications; Pregnancy; Pregnancy Complications; Pregnancy Trimester, Second; Venous Thrombosis; Warfarin; Young Adult
PubMed: 26949306
DOI: 10.4314/ejhs.v25i4.15 -
Blood May 1992A cDNA containing the complete open-reading frame encoding rabbit antithrombin III (AT-III) was isolated from a rabbit liver cDNA expression library, using a specific...
A cDNA containing the complete open-reading frame encoding rabbit antithrombin III (AT-III) was isolated from a rabbit liver cDNA expression library, using a specific antibody as a probe. Sequence analysis showed 84% identity between the deduced amino acid sequences of the rabbit and human proteins. A previously described cell-free expression system was used to verify the identity of the clone. The full-length cDNA was inserted into an expression vector, and messenger RNA (mRNA) transcripts generated. In vitro translation of these transcripts, in the presence of [35S]methionine, in an mRNA-dependent rabbit reticulocyte lysate system resulted in the synthesis of a 51-Kd polypeptide, as shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). This nonglycosylated protein was capable of forming SDS-stable complexes with human alpha-thrombin. Complex formation was significantly enhanced following the deletion of nucleotides encoding the signal peptide, and the resultant generation of a 47-Kd nonglycosylated mature protein product. When the template DNA giving rise to this product was internally truncated, two rabbit AT-III deletion mutants were generated that lacked the ability to interact with thrombin, but retained the ability to bind heparin. Cell-free expression plasmids encoding the human and rabbit AT-III mature molecules were manipulated to produce two interspecies fusion proteins. For the first, human codons were used to replace rabbit codons from residue 369-433, while in the second human codons replaced rabbit codons from residue 217-433. Both fusion proteins exhibited less efficient thrombin-complexing ability than the original cell-free-derived mature rabbit AT-III. Thus, portions of AT-III molecules from the two species, despite their high degree of homology, are not interchangeable. Knowledge of the structure of rabbit AT-III, combined with the availability of the rabbit cDNA, will permit defined experimentation aimed at understanding antithrombin III structure relative to its function in vivo.
Topics: Amino Acid Sequence; Animals; Antithrombin III; Base Sequence; Blotting, Northern; Cell-Free System; Cloning, Molecular; Heparin; Humans; Molecular Sequence Data; Rabbits; Thrombin
PubMed: 1571546
DOI: No ID Found