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Principal Investigator: THOMAS H BUGGE
Organization: NATIONAL INSTITUTE OF DENTAL & CRANIOFACIAL RESEARCH
Fiscal Year: 2024
Award: $1,942,173
Funding agency: National Institute of Dental and Craniofacial Research
Cellular orchestration of extracellular matrix degradation
Background: Our Section has a long-standing interest in pathways of extracellular matrix (ECM) degradation and the pathological consequences of insufficient or excessive ECM turnover.
Research accomplished:
Identification of the mannose receptor (MR) as a clearance receptor for thrombospondins
The mannose receptor (MR) is a large transmembrane glycoprotein that rapidly shuttles between the cell surface and the endocytic compartment glycoprotein and is expressed by M2-polarized tissue repair/remodeling macrophages. We have previously shown that MR functions as a clearance receptor for a diverse array of collagens, as well as for members of the large collectin family of pattern recognition receptors with collagen triple helical-like domains. To search for additional ligands for MR, in collaboration with Henrik Jessen Jurgensen, Lars Engelholm, and Niels Behrendt, University of Copenhagen, Denmark, we performed an unbiased proteomics-based analysis of lung epithelial lining fluid from normal or MR-deficient mouse lungs. Of the more than 1200 proteins identified, six showed significantly different expression, including four previously established MR ligands. One protein, thrombospondin-4 (TSP-4), was not previously identified as an MR ligand, but displayed strikingly increased abundance in lung epithelial fluid from MR-deficient mice. Internalization experiments using recombinant TSP-4 and MR-transfected CHO cells or MR-positive alveolar macrophages revealed that TSP-4 is, indeed, a novel ligand for MR. Mechanistic studies showed that the interaction between TSP-4 and MR is dependent on both the lectin activity of MR and on structural motifs in the collagen-binding fibronectin type-II domain of the receptor. Furthermore, cellular uptake studies with different thrombospondin family members in MR-transfected cells uncovered the closely-related thrombospondin-5, TSP-5, as an additional novel ligand for MR. Furthermore, two other members of the thrombospondin family, TSP-1 and -2 were identified as ligands for a close homologue of MR, the widely-expressed collagen receptor, urokinase plasminogen activator receptor-associated protein (uPARAP). Taken together, the study demonstrates that MR takes part in the regulation of TSP-4, an important inflammatory component, in the injured lung, and that two closely related endocytic receptors, MR and uPARAP, undertake the selective endocytosis of distinct members of the thrombospondin family.
Lipid nanoparticles and siRNA targeting plasminogen provide lasting inhibition of fibrinolysis in mouse and dog models of hemophilia A.
Fibrin is a provisional ECM protein that is formed by the polymerization of thrombin-cleaved fibrinogen, followed by cross-linking of the fibrin polymer by coagulation factor XIII. Fibrin is deposited into the extravascular space in response to tissue injury, where it serves to stem the loss of blood, immobilize bacteria, and provide a provisional matrix for tissue regeneration. Antifibrinolytic drugs, such as tranexamic acid, are used extensively for treatment of severe acute bleeding. Controlling fibrinolysis could also be an effective strategy to prevent and lessen chronic recurring bleeding in bleeding disorders, such as hemophilia A (HA). In a large collaboration, spearheaded by Dr. Christian Kastrup, Department of Surgery, Medical College of Wisconsin, Wauwatosa, Wisconsin, USA, we have assisted in evaluating the therapeutic potential of a long-lasting antifibrinolytic small interfering RNA (siRNA) encapsulated by clinically-used lipid nanoparticles (LNPs). Following siRNA treatment, circulating plasminogen, the zymogen of the principal fibrinolytic protease, plasmin, was selectively depleted, and fibrinolysis was suppressed in wildtype and HA mice and dogs. Treated wildtype mice did not display signature phenotypes of plasminogen deficiency, such as periodontal disease, suggesting that residual plasminogen expression was sufficient to maintain long-term health. In HA mice, hemostatic efficacy depended on the injury model; plasminogen knockdown significantly improved hemostasis after a saphenous vein injury- identifying this as a murine bleeding model particularly sensitive to the contribution of fibrinolysis. In dogs with HA, siRNA targeting plasminogen was as effective at stabilizing clots as clinical antifibrinolytics, and in a pilot study with two dogs, the incidence of spontaneous or excess bleeding was reduced during four months of prolonged knockdown. Collectively, these data show that long-acting antifibrinolytic therapy can be achieved, has hemostatic benefit in multiple animal models of HA, and this is likely to be effective across multiple bleeding disorders and pathologies.
Terms: <AMCA><AMCHA><Acute><Alveolar Macrophages><Animal Model><Animal Models and Related Studies><Antifibrinolysins><Antifibrinolytic Agents><Antifibrinolytics><Antithrombotic Agents><Antithrombotic Drugs><Bacteria><Binding><Biochemistry><Biological Chemistry><Biology><Bleeding><Blood Coagulation Disorders><Blood Coagulation Factor I><Blood Coagulation Factor One><Blood Coagulation Factor XIII><Blood Factor One><Blood Vessels><CD36 Antigens><CD36 Fatty Acid Transporter><CHO Cells><Cancers><Canine Species><Canis familiaris><Cell Body><Cell Surface Glycoproteins><Cell surface><Cell-Extracellular Matrix><Cells><Chinese Hamster Ovary Cell><Chronic><Clinical><Clotting><Coagulation><Coagulation Disorder><Coagulation Factor I><Coagulation Factor One><Coagulation Factor XIII><Coagulation Process><Coagulopathy><Cold-Insoluble Globulins><Collaborations><Collagen><Collagen Receptors><Collagenous Lectins><Collectins><Data><Denmark><Deposit><Deposition><Development><Dogs><Dogs Mammals><EC 3.4.21.7><ECM><Encapsulated><Endocytosis><Enzyme Precursors><Epithelium><Esteroproteases><Extracellular Matrix><Extracellular Matrix Degradation><Extracellular Matrix Proteins><FN1><Factor I><Factor One><Factor VIII Deficiency><Factor XIII><Factor XIII Transamidase><Family><Family member><Fibrin><Fibrin Stabilizing Factor><Fibrinase><Fibrinogen><Fibrinolyses><Fibrinolysin><Fibrinolysis><Fibrinolytic Agents><Fibrinolytic Drugs><Fibrinolytic Therapy><Fibronectin 1><Fibronectins><GPIV Platelet Glycoprotein><GeneHomolog><Glu-Plasmin><Glycoproteins><Health><Hemophilia><Hemophilia A><Hemorrhage><Hemostasis><Hemostatic Agents><Hemostatic function><Hemostatics><Homolog><Homologous Gene><Homologue><Immobilization><Incidence><Inflammatory><Injury><LETS Proteins><Laki-Lorand Factor><Large External Transformation-Sensitive Protein><Lectin><Ligands><Liquid substance><Lung><Lung Respiratory System><Lung damage><Macrophage><Malignant><Malignant - descriptor><Malignant Neoplasms><Malignant Tumor><Membrane Glycoproteins><Mice><Mice Mammals><Modeling><Molecular Interaction><Murine><Mus><Mφ><Natural regeneration><Operative Procedures><Operative Surgical Procedures><Opsonic Glycoprotein><Opsonic alpha(2)SB Glycoprotein><Parodontosis><Pathologic><Pathology><Pathway interactions><Pattern recognition receptor><Peptidases><Peptide Hydrolases><Periodontal Diseases><Phenotype><Pilot Projects><Plasmin><Plasminogen><Polymers><Proenzymes><Profibrinolysin><Protease F><Protease Gene><Proteases><Protein Cleavage><Proteinases><Proteins><Proteolysis><Proteolytic Enzymes><Proteomics><Pulmonary Macrophages><Receptor Protein><Recombinants><Recurrence><Recurrent><Regeneration><Regulation><Research><Residual><Residual state><Saphenous Vein><Short interfering RNA><Small Interfering RNA><Surface Glycoproteins><Surgical><Surgical Interventions><Surgical Procedure><TSP-1><TSP-4><TSP1><TSP4><Therapeutic><Therapeutic Thrombolysis><Thrombase><Thrombin><Thrombolytic Agents><Thrombolytic Drugs><Thrombolytic Therapy><Thrombospondin 1><Thrombospondin Receptors><Thrombospondins><Tranexamic Acid><Transfection><U-PA receptor><Universities><Urokinase Plasminogen Activator Receptor><Urokinase Receptor><Urokinase-type Plasminogen Activator Receptor><Wild Type Mouse><Wisconsin><Zymogens><alpha 2-Surface Binding Glycoprotein><antithrombotic medication><antithrombotics><bleeding disorder><blood loss><canine><canine animal model><canine model><clotting disorder><crosslink><developmental><dog model><domestic dog><experiment><experimental research><experimental study><experiments><fibrinogenase><fluid><improved><injuries><injury to tissue><interest><knock-down><knockdown><lipid based nanoparticle><lipid nanoparticle><liquid><lung injury><malignancy><mannose receptor><medical college><medical schools><member><model of animal><mouse model><murine model><neoplasm/cancer><novel><oral tissue><orthopedic freezing><pathway><periodontal disorder><periodontium disease><periodontium disorder><pilot study><polymer><polymeric><polymerization><prevent><preventing><pro-urokinase receptor><pulmonary><pulmonary damage><pulmonary injury><pulmonary tissue damage><pulmonary tissue injury><receptor><receptor function><regenerate><regenerate new tissue><regenerate tissue><regenerating damaged tissue><regenerating tissue><response><school of medicine><siRNA><stem><surgery><t-AMCHA><thrombospondin 4><tissue injury><tissue regeneration><tissue regrowth><tissue renewal><tissue repair><tissue specific regeneration><triple helix><uPAR receptor><uptake><vascular><wildtype mouse>