Plasma kallikrein: Novel functions for an old protease

Abstract

Editorial on Bird et al. Thromb Haemost 2012; 107: 1141-1150. Plasma kallikrein (PK) is a plasma protease that drives procoagulantand proinflammatory reactions via the intrinsic pathway of coagulation and the kallikrein-kinin system, respectively. The structure and biochemistry of PK have been analysed in great detail; however, its in vivo functions are just beginning to emerge. In this issue of Thrombosis and Haemostasis, Bird et al. presents a mouse model with target ablation of the PK gene (1). Severe deficiency of PK abolishes experimental thrombosis in arterial and venous vascular beds. Despite the striking thromboprotective effects conferred by targeting the protease, PK deficiency has minor effect on the haemo static capacity of gene-deficient mice. This animal model provides a promising tool to study the pathogenetic role of the protease in models of thrombosis and inflammation with exciting therapeutic perspectives. In the original waterfall cascade model of plasma coagulation, fibrin formation may be initiated by two distinct pathways, triggered either by exposure of blood to tissue factor at an injured vessel wall or to blood-borne (intrinsic) factors. The intrinsic pathway of coagulation is initiated by factor (F)XII, in a reaction involving highmolecular-weight kininogen (HK) and PK, collectively referred to as the plasma contact system (2). Contact with negatively charged surfaces such as kaolin or polyphosphate activates FXII, and activated FXII (FXIIa) cleaves PK to generate active kallikrein, which in turn reciprocally activates FXII zymogen. FXIIa initiates fibrin formation through its principal substrate of the intrinsic coagulation pathway, FXI and also triggers liberation of the potent inflammatory mediator bradykinin (BK) by PK-mediated HK cleavage (3). Binding of BK to the kinin B2 receptor (B2R) causes an immediate nitric oxide-mediated vasodilation through an increase of cGMP in vascular smooth muscle cells, activates various pro-inflammatory signalling pathways that dilate vessels, induces chemotaxis of neutrophils and increases vascular permeability and fluid efflux (4). In a positive feedback loop, kallikrein also activates surfacebound FXII to FXIIa. Furthermore, PK has the capacity to convert pro-urokinase to urokinase, plasminogen to plasmin, and for triggering the renin-angiotensin system. Due to its BK-forming capacity, kallikrein has potent in vitro effects on leukocyte migration, stimulates monocytes, and induces neutrophil aggregation. Recent studies have suggested a role of PK for control of intracerebral haemorrhage in hyperglycaemia (5). In contrast, hereditary deficiencies in PK (Fletcher trait) and its substrate HK (Fitzgerald trait) were not associated with haemostatic abnormalities (6). The serpin C1 esterase inhibitor (C1INH) is the major plasma inhibitor of PK and controls enzymatic activity of the protease. Plasma prokallikrein (PPK) is the zymogen of the trypsin-like serine protease PK. PPK is expressed by hepatocytes in humans and mice and circulates in plasma at a concentration of 50 g/ml, predominantly in a noncovalent 1:1-complex with HK. PK is a paralog of FXI, and both genes have evolved from a common anchestor (7). The principal mechanism of PK is to cleave the high-molecular-weight precursor (HK) that liberates BK. Despite similar names, PK is not related to the large group of kininogenases called tissue-type (urinary or glandular) kallikreins. Analogous to FXI, PK is composed of four apple domains (Apple 1–4) that form the heavy chain of the molecule and a C-terminal protease domain (8). However, in contrast to human FXI, human PK is a monomer. PK migrates as a doublet with 86and 88-kDa on nonreducing SDS-polyacrylamide gels. The two bands represent differentially glycosylated proteins. Contact activation is the mechanism of contact-induced FXII zymogen activation. Contact activation provides the mechanistic basis for one of the most commonly used diagnostic clotting test the activated partial thromboplastin time (aPTT). The contact surface used in aPTT assays is typically a negatively charged non-physiological substance such as kaolin (aluminum silicate), celite (diatomaceous earth), or ellagic acid. The aPTT is widely used in clinical practice for preoperative screening and monitoring of anticoagulant therapy. Despite its important role in fibrin formation in the aPTT assay, PK-driven coagulation does not appear to have haemo static or other physiological functions in vivo. This notion is based on the observation that PK deficiency, a rare incidentally observed coagulation abnormality in humans, has not yet been associated with pathology, such as abnormal bleeding, despite causing a marked prolongation of the aPTT. Similar to PK-deficient individuals, humans lacking the other contact proteins, FXII or HK, do not have impaired haemo stasis. Recently, FXII-deficient (FXII-/-) mice were generated and phenotyped to study the function of coagulation FXII in vivo (9, 10). Similar to FXII-deficient humans, FXII-/mice have a normal haemostatic capacity as assessed by a tail-bleeding assay. Completely unexpected, intravital fluorescence microscopy and blood flow measurements in three distinct arterial beds revealed a severe defect in FXII-defi-