title: Neutrophil cathepsin G promotes prothrombinase and fibrin formation under flow conditions by activating fibrinogen-adherent platelets.
abstract: Human neutrophil proteases cathepsin G and elastase can directly alter platelet function and/or participate in coagulation cascade reactions on the platelet or neutrophil surface to enhance fibrin formation. The clotting of recalcified platelet-free plasma (PFP) or platelet-rich plasma (PRP) supplemented with corn trypsin inhibitor (to shut down contact activation) was studied in well-plates or flow assays. Inhibitors of cathepsin G or elastase significantly delayed the burst time (t(50)) of thrombin generation in neutrophil-supplemented PRP from 49 min to 59 and 77 min, respectively, in well-plate assays as well as reduced neutrophil-promoted fibrin deposition on fibrinogen-adherent platelets under flow conditions. In flow assays, purified cathepsin G was a far more potent activator of platelet-dependent coagulation than elastase. Anti-tissue factor had no effect on neutrophil protease-enhanced thrombin formation in PRP. The addition of cathepsin G (425 nm) or convulxin (10 nm) to PRP dramatically reduced the t(50) of thrombin generation from 53 min to 17 or 23 min, respectively. In contrast, the addition of elastase to PRP left the t(50) unaltered. Whereas perfusion of PFP (gamma(w) = 62.5 s(-1)) over fibrinogen-adherent platelets did not result in fibrin formation until 50 min, massive fibrin could be observed on cathepsin G-treated platelets even at 35 min. Cathepsin G addition to corn trypsin inhibitor-treated PFP produced little thrombin unless anionic phospholipid was present. However, further activation inhibition studies indicated that cathepsin G enhances fibrin deposition under flow conditions by elevating the activation state of fibrinogen-adherent platelets rather than by cleaving coagulation factors.
authors.affiliation: Institute for Medicine and Engineering, Department of Chemical Engineering, University of Pennsylvania, 1010 Vagelos Research Laboratories, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Mukul S
authors.initials: MS
authors.lastname: Goel
authors.forename: Scott L
authors.initials: SL
authors.lastname: Diamond
mesh_terms: D000328:Adult; D001792:Blood Platelets / Q000378:metabolism*; D056649:Cathepsin G; D002403:Cathepsins / Q000737:chemistry* / Q000378:metabolism; D002448:Cell Adhesion; D005337:Fibrin / Q000378:metabolism*; D005340:Fibrinogen / Q000737:chemistry*; D006439:Hemodynamics; D006801:Humans; D008856:Microscopy, Fluorescence; D018715:Microscopy, Video; D008954:Models, Biological; D009504:Neutrophils / Q000378:metabolism*; D010196:Pancreatic Elastase / Q000737:chemistry / Q000378:metabolism; D010477:Perfusion; D010743:Phospholipids / Q000737:chemistry; D017434:Protein Structure, Tertiary; D012697:Serine Endopeptidases; D013917:Thrombin / Q000378:metabolism; D013925:Thromboplastin / Q000378:metabolism*; D013927:Thrombosis; D013997:Time Factors
chemical_list: D010743:Phospholipids; D005337:Fibrin; D005340:Fibrinogen; D013925:Thromboplastin; D002403:Cathepsins; D012697:Serine Endopeptidases; C534094:CTSG protein, human; D056649:Cathepsin G; D010196:Pancreatic Elastase; D013917:Thrombin