Genetic Variability of the CLEC4M Endothelial Lectin Receptor Modulates Binding and Internalization of Von Willebrand Factor and Contributes to Variance in Plasma VWF Levels
Notice bibliographique
Résumé
Abstract Abstract 16 Type 1 von Willebrand's Disease (VWD) can result from decreased synthesis or accelerated clearance of von Willebrand Factor (VWF), resulting in partial quantitative deficiency. Approximately 35% of individuals with Type 1 VWD do not have a putative mutation in their VWF gene, suggesting that genes other than VWF may contribute to the pathophysiology of this disease. Recently, the CHARGE GWAS meta-analysis identified single nucleotide polymorphisms in the gene encoding the C-type lectin domain family 4 member M (CLEC4M) as being significantly associated with plasma VWF levels in normal individuals. CLEC4M is a pathogen recognition receptor with a polymorphic extracellular neck region consisting of a variable number of tandem repeats (VNTR) (3 – 9 repeats). We hypothesize that CLEC4M binds to and clears VWF from the circulation, and that different CLEC4M VNTR alleles may contribute to differences in plasma levels of VWF in normal subjects and patients with Type 1 VWD. Previously, genotyping of 555 subjects (196 cases with type 1 VWD, and 362 family members) for the CLEC4M VNTR number showed that the most frequently documented alleles were VNTR 5 (29%), 6 (15%), and 7 (53%). Family-based association analysis on kindreds with type I VWD has demonstrated a significant excess transmission of VNTR 6 to the type I VWD phenotype (p=0.005) and an association of this VNTR allele with VWF:RCo (p=0.037). In the present studies, we have complemented this genetic association data with experiments to directly evaluate the ability of CLEC4M to bind and internalize VWF. Further, we characterized the ability of different CLEC4M VNTR alleles to facilitate VWF clearance. Binding of VWF to CLEC4M was assessed with a modified ELISA using a recombinant CLEC4M-Fc chimera. CLEC4M-Fc bound to Humate P (plasma-derived VWF-FVIII) in a dose-dependent manner. CLEC4M-Fc also bound to recombinant human VWF, and factor VIII-free plasma-derived VWF. CLEC4M-Fc demonstrated a 70% increase in binding to de-O-glycosylated Humate P (p=0.041), and a 75% decrease in binding to de-N-glycosylated Humate P (p=0.046) relative to controls. Additionally, pre-incubation of CLEC4M-Fc with the polysaccharide mannan attenuated binding to all VWF preparations by approximately 50%. Binding and internalization of VWF by HEK 293 cells stably expressing CLEC4M (VNTR allele 7) was assessed with immunofluorescence and ELISA. Binding of VWF co-localized with CLEC4M expression on HEK 293 cells. CLEC4M and VWF co-localized with early endosomal antigen-1, suggesting that CLEC4M participates in receptor-mediated endocytosis of VWF. CLEC4M-expressing cells bound and internalized VWF in a dose- and time-dependent manner relative to controls. Preincubation of CLEC4M expressing cells with mannan inhibited VWF binding and internalization by 50% (p=0.0088). The contribution of CLEC4M genetic variability to VWF binding and internalization was measured using HEK 293 cells expressing CLEC4M with 4, 6, 7, and 9 tandem repeats. Decreased binding and internalization of VWF was observed with cells expressing CLEC4M 4 and 9 tandem repeat constructs as compared to CLEC4M with 7 tandem repeats (CLEC4M 4 – 60% reduction, p < 0.001; CLEC4M 9 – 45% reduction, p=0.006). Cells expressing the CLEC4M VNTR combination 4 and 9, had a 55% decrease in binding and internalization of VWF relative to cells expressing CLEC4M with 7 VNTRs (p < 0.001). These VNTR associated differences in VWF binding/internalization were not accounted for by variances in the CLEC4M expression levels in the transfected HEK 293 cells. These studies demonstrate that the C-type lectin CLEC4M binds to and internalizes VWF through an N-glycan-dependent mechanism. Additionally, it provides further evidence that polymorphisms in the CLEC4M gene contribute to quantitative VWF deficiency. Disclosures: Montgomery: Gen-Probe/GTI Diagnostics: Consultancy; CSL Behring: Consultancy; Octapharma: Consultancy. James:CSL-Behring, Baxter, Bayer: Honoraria, Research Funding.
Récupéré en direct depuis OpenAlex et désinversé. Les résumés ne sont pas conservés dans cette base de données : les index inversés représentent 8,6 Go des 9,3 Go de texte de la base, et le serveur dispose de 13 Go libres.
Comment cette classification a été obtenuedéplier
Prédiction machine sur la base complète
Imitation des enseignantsNi prévalence calibrée, ni vérité terrain. Validation humaine à venir. Le volet Gemma est une étiquette directe du modèle pour chaque travail de la base, lue sur la notice réduite au titre. Le volet Codex est un classifieur appris des 10 348 étiquettes directes de Codex et calibré sur les taux pondérés de l'échantillon; les champs sans appui suffisant ne portent aucun appel Codex. Le mode candidate est l'union des deux volets; le consensus est leur intersection. Ces sorties portent le statut machine_predicted_unvalidated et ne sont pas des étiquettes humaines.
Scores du classifieur distillé par catégorie (deux têtes)
| Catégorie | Codex | Gemma |
|---|---|---|
| Métarecherche | 0,001 | 0,002 |
| Méta-épidémiologie (sens strict) | 0,000 | 0,000 |
| Méta-épidémiologie (sens large) | 0,000 | 0,001 |
| Bibliométrie | 0,001 | 0,001 |
| Études des sciences et des technologies | 0,000 | 0,000 |
| Communication savante | 0,000 | 0,000 |
| Science ouverte | 0,000 | 0,000 |
| Intégrité de la recherche | 0,001 | 0,000 |
| Charge utile insuffisante (le modèle a refusé de juger) | 0,002 | 0,000 |
Scores machine (provisoires)
Les deux têtes enseignantes du modèle étudiant, lues sur ce travail. Un score ordonne la base pour la relecture; il n'affirme jamais une catégorie, et le statut de validation accompagne chaque rangée tel quel.
Scores de référence d'un modèle non mature (critères de maturité non atteints, 7 itérations). Un score ordonne; il n'affirme jamais une catégorie.
score_only:v0-immature-baseline · tel quel depuis la passe de notation : score_only signifie que le nombre peut ordonner les travaux, et qu'aucune étiquette de catégorie n'en découleClassification
machine, non validéePrédiction automatique; un appel candidat d’une seule source (Gemma direct ou Codex distillé), pas un consensus.
Le détail, modèle par modèle et score par score, se trouve en fin de page sous « Comment cette classification a été obtenue ».