Beyond basic characterization and omics: Immunomodulatory roles of platelet-derived extracellular vesicles unveiled by functional testing.

Renowned for their role in haemostasis and thrombosis, platelets are also increasingly recognized for their contribution in innate immunity, immunothrombosis and inflammatory diseases. Platelets express a wide range of receptors, which allows them to reach a variety of activation endpoints and grants them immunomodulatory functions. Activated platelets release extracellular vesicles (PEVs), whose formation and molecular cargo has been shown to depend on receptor-mediated activation and environmental cues.

Integrin α2 is an early marker for osteoclast differentiation that contributes to key steps in osteoclastogenesis.

Introduction: Osteoclasts determine bone tissue turnover. Their increased activity causes osteoporosis, their dysfunction osteopetrosis.

Methods And Results: Murine monocytic ER-Hoxb8 cells differentiate into OCs upon treatment with M-CSF and RANKL and upregulate the collagen-binding integrin α2β1 distinctly earlier than other OC markers, such as the OC-associated receptor, OSCAR.

Correction: Flammang et al. Tumor-Suppressive miR-192-5p Has Prognostic Value in Pancreatic Ductal Adenocarcinoma. 2020, , 1693.

In the original publication [...

Structure-Activity Relationship of Synthetic Linear KTS-Peptides Containing Meta-Aminobenzoic Acid as Antagonists of α1β1 Integrin with Anti-Angiogenic and Melanoma Anti-Tumor Activities.

To develop peptide drugs targeting integrin receptors, synthetic peptide ligands endowed with well-defined selective binding motifs are necessary. The snake venom KTS-containing disintegrins, which selectively block collagen α1β1 integrin, were used as lead compounds for the synthesis and structure-activity relationship of a series of linear peptides containing the KTS-pharmacophore and alternating natural amino acids and 3-aminobenzoic acid (MABA). To ensure a better stiffness and metabolic stability, one, two and three MABA residues, were introduced around the KTS pharmacophore motif.

Characterization of the molecular composition and in vitro regenerative capacity of platelet-based bioproducts and related subfractions.

The use and demand of platelet-based bioproducts in regenerative medicine is steadily increasing. However, it is very difficult to establish the real clinical benefits of these therapies, as the lack of characterization and detailed production methods of platelet-based bioproducts persists in the literature and precludes cross-study comparisons. We characterized the molecular composition and in vitro regenerative capacity of platelet-rich plasma (PRP) produced in a closed-system.

A change of rhodocytin's suprastructure turns the agonist into an antagonist of tumor cell induced platelet aggregation.

Cancer cells circulating in the blood attach to platelets by direct cell-cell interactions via several receptor-counterreceptor contacts and indirectly by fibrin bridges which connect the two cell types by distinct integrin receptors. In the microenvironment of these tumor cell platelet aggregates (TCPAs), the tumor cells are shielded from the shear stress of the blood flow and from attack by the immune system. This supports hematogenous metastasis and tumor cell induced thrombosis.

Reversible photoregulation of cell-cell adhesions with opto-E-cadherin.

E-cadherin-based cell-cell adhesions are dynamically and locally regulated in many essential processes, including embryogenesis, wound healing and tissue organization, with dysregulation manifesting as tumorigenesis and metastasis. However, the lack of tools that would provide control of the high spatiotemporal precision observed with E-cadherin adhesions hampers investigation of the underlying mechanisms. Here, we present an optogenetic tool, opto-E-cadherin, that allows reversible control of E-cadherin-mediated cell-cell adhesions with blue light.

Multiparametric chemical exchange saturation transfer MRI detects metabolic changes in breast cancer following immunotherapy.

Background: With metabolic alterations of the tumor microenvironment (TME) contributing to cancer progression, metastatic spread and response to targeted therapies, non-invasive and repetitive imaging of tumor metabolism is of major importance. The purpose of this study was to investigate whether multiparametric chemical exchange saturation transfer magnetic resonance imaging (CEST-MRI) allows to detect differences in the metabolic profiles of the TME in murine breast cancer models with divergent degrees of malignancy and to assess their response to immunotherapy.

Methods: Tumor characteristics of highly malignant 4T1 and low malignant 67NR murine breast cancer models were investigated, and their changes during tumor progression and immune checkpoint inhibitor (ICI) treatment were evaluated.

Platelet functional abnormalities in pediatric patients with kaposiform hemangioendothelioma/Kasabach-Merritt phenomenon.

Kaposiform hemangioendothelioma (KHE) is a rare vascular tumor of infancy that is commonly associated with a life-threatening thrombocytopenic condition, Kasabach-Merritt phenomenon (KMP). Platelet CLEC-2, tumor podoplanin interaction is considered the key mechanism of platelet clearance in these patients. Here, we aimed to assess platelet functionality in such patients.

Divalent nanobodies to platelet CLEC-2 can serve as agonists or antagonists.

CLEC-2 is a target for a new class of antiplatelet agent. Clustering of CLEC-2 leads to phosphorylation of a cytosolic YxxL and binding of the tandem SH2 domains in Syk, crosslinking two receptors. We have raised 48 nanobodies to CLEC-2 and crosslinked the most potent of these to generate divalent and tetravalent nanobody ligands.

Rhomb-I, a P-I metalloproteinase from Lachesis muta rhombeata venom degrades vessel extra cellular matrix components and impairs platelet aggregation.

Rhomb-I, a 23-kDa metalloproteinase was isolated from L. m. rhombeata venom.

Protein Disulfide Isomerase and Extracellular Adherence Protein Cooperatively Potentiate Staphylococcal Invasion into Endothelial Cells.

Invasion of host cells is an important feature of Staphylococcus aureus. The main internalization pathway involves binding of the bacteria to host cells, e.g.

Profiling specific cell populations within the inflammatory tumor microenvironment by oscillating-gradient diffusion-weighted MRI.

Background: The inflammatory tumor microenvironment (TME) is formed by various immune cells, being closely associated with tumorigenesis. Especially, the interaction between tumor-infiltrating T-cells and macrophages has a crucial impact on tumor progression and metastatic spread. The purpose of this study was to investigate whether oscillating-gradient diffusion-weighted MRI (OGSE-DWI) enables a cell size-based discrimination between different cell populations of the TME.

Stimulation of platelet aggregation by affinity captured rhodocytin from the Malayan pit viper Calloselasma rhodostoma.

The receptor protein CLEC-2 on platelet membranes is the target of the endogenous ligand podoplanin found on cancer cells and of rhodocytin, a snake venom component of the Malayan pit viper Calloselasma rhodostoma. Ligand binding results in platelet activation, increased blood coagulation and thrombosis. In an effort to isolate rhodocytin, we have purified CLEC-2 as bait from E.

Experimental validation of computerised models of clustering of platelet glycoprotein receptors that signal via tandem SH2 domain proteins.

The clustering of platelet glycoprotein receptors with cytosolic YxxL and YxxM motifs, including GPVI, CLEC-2 and PEAR1, triggers activation via phosphorylation of the conserved tyrosine residues and recruitment of the tandem SH2 (Src homology 2) domain effector proteins, Syk and PI 3-kinase. We have modelled the clustering of these receptors with monovalent, divalent and tetravalent soluble ligands and with transmembrane ligands based on the law of mass action using ordinary differential equations and agent-based modelling. The models were experimentally evaluated in platelets and transfected cell lines using monovalent and multivalent ligands, including novel nanobody-based divalent and tetravalent ligands, by fluorescence correlation spectroscopy.

The Current State-of-the-Art Identification of Unknown Proteins Using Mass Spectrometry Exemplified on De Novo Sequencing of a Venom Protease from .

(1) Background: The amino acid sequence elucidation of peptides from the gas phase fragmentation mass spectra, de novo sequencing, is a valuable method for the identification of unknown proteins complementary to Edman sequencing. It is increasingly used in shot-gun mass spectrometry (MS)-based proteomics experiments. We review the current state-of-the-art and use the identification of an unknown snake venom protein targeting the human tissue factor (TF) as an example to describe the analysis process based on manual spectrum interrogation.

Comparison of inhibitory effects of irreversible and reversible Btk inhibitors on platelet function.

All irreversible Bruton tyrosine kinase (Btk) inhibitors including ibrutinib and acalabrutinib induce platelet dysfunction and increased bleeding risk. New reversible Btk inhibitors were developed, like MK-1026. The mechanism underlying increased bleeding tendency with Btk inhibitors remains unclear.

A redox-dependent thiol-switch and a Ca binding site within the hinge region hierarchically depend on each other in α7β1 integrin regulation.

Integrin-mediated cell contacts with the extracellular matrix (ECM) are essential for cellular adhesion, force transmission, and migration. Several effectors, such as divalent cations and redox-active compounds, regulate ligand binding activities of integrins and influence their cellular functions. To study the role of the Ca binding site within the hinge region of the integrin α7 subunit, we genetically abrogated it in the α7hiΔCa mutant.

Interaction of Bartonella henselae with Fibronectin Represents the Molecular Basis for Adhesion to Host Cells.

Bacterial adhesion to the host is the most decisive step in infections. Trimeric autotransporter adhesins (TAA) are important pathogenicity factors of Gram-negative bacteria. The prototypic TAA adhesin A (BadA) from human-pathogenic Bartonella henselae mediates bacterial adherence to endothelial cells (ECs) and extracellular matrix proteins.

Dissecting platelet proteomics to understand the pathophysiology of immune thrombocytopenia: studies in mouse models.

Immune thrombocytopenia (ITP) is an autoimmune disease characterized by enhanced platelet clearance and defective platelet production. Diagnosis by exclusion and trial-and-error treatment strategies is common practice, and despite the advancement in treatment options, many patients remain refractory. Although the existence of different pathophysiological entities is acknowledged, we are still far from stratifying and understanding ITP.

Platelet number and function alterations in preclinical models of sterile inflammation and sepsis patients: implications in the pathophysiology and treatment of inflammation.

Platelets are the blood cells in charge of maintaining the body hemostasis, recognising the damaged vessel wall, and providing the appropriate cellular surface for the coagulation cascade to act locally. Additionally, platelets are active immunomodulators. At the crossroads of hemostasis and inflammation, platelets may exert either beneficial actions or participate in pathological manifestations, and have been associated with the prothrombotic nature of multi-organ failure in systemic inflammation.

Katacine Is a New Ligand of CLEC-2 that Acts as a Platelet Agonist.

Background:  CLEC-2 is a platelet receptor with an important role in thromboinflammation but a minor role in hemostasis. Two endogenous ligands of CLEC-2 have been identified, the transmembrane protein podoplanin and iron-containing porphyrin hemin, which is formed following hemolysis from red blood cells. Other exogenous ligands such as rhodocytin have contributed to our understanding of the role of CLEC-2.

Matrix Metalloproteinases Shape the Tumor Microenvironment in Cancer Progression.

Cancer progression with uncontrolled tumor growth, local invasion, and metastasis depends largely on the proteolytic activity of numerous matrix metalloproteinases (MMPs), which affect tissue integrity, immune cell recruitment, and tissue turnover by degrading extracellular matrix (ECM) components and by releasing matrikines, cell surface-bound cytokines, growth factors, or their receptors. Among the MMPs, MMP-14 is the driving force behind extracellular matrix and tissue destruction during cancer invasion and metastasis. MMP-14 also influences both intercellular as well as cell-matrix communication by regulating the activity of many plasma membrane-anchored and extracellular proteins.