Biology subjects

Bergmeier, W.

Publications and source records attributed to Bergmeier, W..

6 recordsLinked to original sources

bioRxiv2026

Megakaryocyte emperipolesis arms neutrophils via intracellular provisioning

Neutrophils are phenotypically heterogenous cells that mediate host defense and tissue homeostasis. Here, we identify emperipolesis - the evolutionarily conserved process by which neutrophils pass through megakaryocytes - as a phenotypically transformative route of egress from bone marrow. By intravital microscopy and 3-D histology, we show that the rapid form emperipolesis is markedly enhanced under inflammatory conditions. Neutrophils exit from megakaryocytes directly to the blood, acquiring exosomes enriched in proteins related to metabolism, migration, and immune function. This transfer induces a distinct neutrophil phenotype characterized by enhanced glycolysis, oxidative phosphorylation, cytokine release, and longevity. Correspondingly, emperipolesis-educated neutrophils display accelerated migration in vitro and in vivo. Disrupting emperipolesis does not alter circulating neutrophil abundance but impairs neutrophil infiltration into inflamed tissues, including Pseudomonas aeruginosa-infected lung. These findings establish emperipolesis as a mechanism by which megakaryocytes amplify neutrophil-mediated immunity.

immunology↗

bioRxiv2026

Lack of canonical PAR4 activation is associated with reduced arterial and venous thrombosis in mice.

BackgroundProtease-activated receptor 4 (PAR4) is the only functional thrombin receptor on mouse platelets. Expression and activation of platelet PAR4 was shown to influence hemostatic plug stability and contributes to thrombosis in different murine arterial and venous thrombosis models. PAR4 activation by thrombin and other serine proteases occurs at the canonical activation site at Arginine (Arg) 59 in mice and Arg47 in humans. If murine PAR4 has functional non-canonical activation sites as shown for other PARs is unknown. ObjectiveTo investigate canonical and potentially non-canonical PAR4 signaling in mice, we generated a mouse model expressing a functional, thrombin-cleavage resistant PAR4 by changing Arg59 to Alanine (Ala) 59 in murine PAR4 (PAR4R59A). PAR4R59A mice were used to assess the impact of impaired canonical (thrombin)-dependent PAR4 signaling on hemostasis and thrombosis in mice. MethodsWe analyzed platelet aggregation, platelet integrin activation and -granule release ex vivo. Hemostasis and thrombosis in PAR4R59A and their control mice was compared using the jugular vein needle puncture injury-induced hemostasis model, and the ferric chloride-induced carotid artery and electrolytic injury-induced femoral vein thrombosis models. ResultsPlatelets of PAR4R59A mice did not response to thrombin but responded normally to PAR4 agonist peptide (PAR4AP) stimulation in aggregation assays. Platelets of PAR4R59A and their control mice exhibited comparable responses to ADP, convulxin or PAR4AP regarding integrin activation and -granular release. PAR4R59A mice exhibited impaired hemostasis in the jugular vein needle puncture model, and were protected from ferric chloride-induced arterial thrombosis and from electrolytic injury induced thrombosis in the femoral vein. ConclusionThe novel PAR4R59A mouse expresses a thrombin-insensitive but still functional PAR4. We propose that the new mouse line will increase the in vivo investigation of canonical PAR4 signaling pathways and may reveal unknown non-canonical PAR4 signaling in different pathologies.

pathology↗

bioRxiv2025

RAP1-RHO small GTPase cross-talk mediates integrin-dependent and -independent platelet procoagulant response

Platelet adhesion and procoagulant activity are critical for primary and secondary hemostasis, respectively. The small GTPase RAP1 is a central regulator of platelet aggregation as it controls IIb{beta}3 integrin activation through direct interaction with the integrin adapter protein, TALIN-1 (Tln-1). In addition to their aggregation defect, activated platelets lacking RAP1 (Rap1mKO) exhibited a marked impairment in surface exposure of phosphatidylserine (PtdSer), a negatively charged phospholipid with procoagulant activity. However, the mechanisms by which RAP1 regulates PtdSer exposure are unclear. Here we investigated the hypothesis that RAP1 regulates platelet PtdSer exposure through cross-talk with small GTPases of the Rho family. Consistent with their defect in PtdSer exposure, Rap1mKO platelets showed reduced procoagulant activity in vitro and in vivo when compared to controls. Stimulated Rap1mKO platelets exhibited elevated RHOA-GTP levels, and inhibition of the RHOA effector, Rho associated coiled-coil kinase (ROCK), partially restored PtdSer exposure in these cells. A milder defect in PtdSer exposure was observed for platelets from Tln-1mR35/118E mice, i.e. mice with impaired RAP1-Tln-1 interaction but otherwise intact RAP1 signaling. ROCK inhibition fully restored PtdSer exposure in Tln-1mR35/118E platelets. Opening of the mitochondrial permeability transition pore, a cellular response critical to PtdSer exposure, was impaired in Rap1mKO platelets and restored by pretreatment of cells with the ROCK inhibitor. Our study provides first evidence that platelet RAP1 signaling affects hemostatic plug formation independent of its key role in platelet adhesion. Additionally, our studies strongly suggest that RAP1 regulates PtdSer exposure and procoagulant activity in a RHOA/integrin-dependent and -independent manner.

cell biology↗

bioRxiv2024

Platelets are Protective in Early Abdominal Aortic Aneurysm Formation

BackgroundAbdominal aortic aneurysm (AAA) is a disease associated with the pathophysiologic degradation of the tunica media resulting in aortic dilatation, systemic inflammation, and dysregulated hemostasis. Beyond role its role in initiating primary hemostasis, platelets are a source of ROS, inflammatory cytokines and growth factors necessary for angiogenesis and vascular remodeling. Although platelets contribute to the progression of established aneurysms, their role in the initiation of AAA remains undefined. MethodsLow density lipoprotein receptor deficient (Ldlr-/-) mice were examined for platelet accumulation in the angiotensin II (AngII) model of AAA utilizing in vivo labeling techniques. Two platelet antagonists (clopidogrel and aspirin), a thrombin inhibitor (dabigatran) or genetic deficiencies (protease-activated receptor 4, P2Y12, Lnk) were administered to AngII-infused mice to determine the role of platelets in initiation of AAA. The effect of platelet depletion was examined in multiple mouse strains of AngII-induced AAA and two additional aneurysm models. PheWAS and meta-analysis was analyzed in humans for platelet gene SNPs associated with AAA. ResultsWe show that platelets are recruited rapidly to the aorta after the initiation of AngII infusion. Genetic deficiency of platelet receptors had no effect on abdominal aortic diameter, but augmented rupture-induced death in littermate versus placebo controls during AngII-induced AAA. Moreover, Ldlr-/- mice receiving anti-platelet inhibitors or a thrombin inhibitor also had augmented rupture-induced death. Platelet depletion preceding aneurysm formation resulted in pervasive rupture-induced death in several mouse strains and with three different mouse models of AAA. ConclusionsInhibition of platelet function is detrimental in an early expanding aortic lumen resulting in catastrophic rupture and hemodynamic failure in murine AAA models.

pathology↗

bioRxiv2024

Utility of thromboelastography with platelet mapping (TEG-PM) for monitoring platelet transfusion in qualitative platelet disorders

Patients with pathogenic variants in RASGRP2 (inherited platelet disorder (IPD)-18) have normal platelet counts but show impaired platelet aggregation due to diminished activation of IIb{beta}3 integrin. This defect results in moderate to severe bleeding episodes, especially following surgical procedures, which require patients to be transfused with platelets and/or pro-hemostatic agents. We recently demonstrated that the hemostatic efficacy of transfused platelets is limited by dysfunctional endogenous platelets in a mouse model of IPD-18 (Rasgrp2-/- mice), as dysfunctional platelets were recruited to the forming hemostatic plug but did not participate in clot contraction. Consequently, higher amounts of transfused platelets were required to outcompete these dysfunctional cells and to reverse bleeding. We here studied the usefulness of thromboelastography with platelet mapping (TEG-PM), a method to evaluate platelet-dependent clot contraction, for ex vivo monitoring of the hemostatic potential in Rasgrp2-/- mice transfused with various amounts of wild-type (WT) platelets. Rasgrp2-/- whole blood samples did not contract in TEG-PM, consistent with a critical role of this protein in IIb{beta}3 activation. Addition of WT platelets improved TEG parameters (K time, -angle, MA) in a ratio dependent manner, consistent with our recent in vivo studies showing impaired hemostasis at a 5:1, but not at a 2:1 ratio of mutant to WT platelets. Interestingly, K and values were identified as better predictors of transfusion efficacy than MA, the most platelet-dependent TEG parameter. In conclusion, this proof-of-concept study supports the use of TEG-PM to monitor platelet transfusion ratios and hemostatic potential in IPD-18 and potentially other platelet disorders.

cell biology↗

bioRxiv2023

4D intravital imaging studies iden1fy platelets as the predominant cellular procoagulant surface in a mouse model of hemostasis

Interplay between platelets, coagulation/fibrinolytic factors, and endothelial cells (ECs) is necessary for effective hemostatic plug formation. This study describes a novel four-dimensional (4D) imaging platform to visualize and quantify hemostatic plug components with high spatiotemporal resolution. Fibrin accumulation following laser-induced endothelial ablation was observed at the EC-platelet plug interface, controlled by the antagonistic balance between fibrin generation and breakdown. Phosphatidylserine (PS) was first detected in close physical proximity to the fibrin ring, followed by exposure across the endothelium. Impaired PS exposure in cyclophilinD-/- mice resulted in a significant reduction in fibrin accumulation. Adoptive transfer and inhibitor studies demonstrated a key role for platelets, but not ECs, in fibrin generation during hemostatic plug formation. Inhibition of fibrinolysis with tranexamic acid (TXA) led to increased fibrin accumulation in WT mice, but not in cyclophilinD-/- mice or WT mice treated with antiplatelet drugs. These studies implicate platelets as the functionally dominant procoagulant surface during hemostatic plug formation. In addition, they suggest that impaired fibrin formation due to reduced platelet procoagulant activity is not reversed by TXA treatment.

cell biology↗