The emerging role of coagulation proteases in kidney disease

Abstract

A role of coagulation proteases in kidney disease beyond their function in normal haemostasis and thrombosis has long been suspected, and studies performed in the past 15 years have provided novel insights into the mechanisms involved. The expression of protease-activated receptors (PARs) in renal cells provides a molecular link between coagulation proteases and renal cell function and revitalizes research evaluating the role of haemostasis regulators in renal disease. Renal cell-specific expression and activity of coagulation proteases, their regulators and their receptors are dynamically altered during disease processes. Furthermore, renal inflammation and tissue remodelling are not only associated, but are causally linked with altered coagulation activation and protease-dependent signalling. Intriguingly, coagulation proteases signal through more than one receptor or induce formation of receptor complexes in a cell-specific manner, emphasizing context specificity. Understanding these cell-specific signalosomes and their regulation in kidney disease is crucial to unravelling the pathophysiological relevance of coagulation regulators in renal disease. In addition, the clinical availability of small molecule targeted anticoagulants as well as the development of PAR antagonists increases the need for in-depth knowledge of the mechanisms through which coagulation proteases might regulate renal physiology.

Figure 1. The coagulation system

The tissue factor (TF) and contact activation pathways lead to a common pathway that generates thrombin. In the tissue factor pathway, tissue injury or inflammatory cytokines induce cell-surface expression of TF. The TF/FVIIa complex activates FX, which converts FII to thrombin. In the contact activation pathway, negatively charged surfaces (such as phospholipids and polyphosphates from activated platelets) activate FXII, initiating a cascade leading to FX activation and thrombin generation. Thrombin triggers formation of blood clots, provides feed-back amplification or inhibition of the coagulation activation process and engages in receptor-dependent signalling. Localization of the coagulation cascade to cell surfaces ensures a spatial constraint on thrombin generation. Feed-back amplification is provided by activation of the non-catalytic cofactors FV and FVIII, platelet activation and thrombin-mediated activation of FXI. Excess coagulation activation is averted through several anticoagulant mechanisms, including inhibition of the TF/FVIIa/FXa complex by tissue factor pathway inhibitor (TFPI), inhibition of several coagulation factors by antithrombin (AT) and proteolytic inactivation of FVa and FVIIIa by activated protein C (aPC). As aPC is generated by the FIIa/thrombomodulin complex on undisturbed endothelial cells, local generation of FIIa following endothelial or vascular injury triggers aPC formation in a spatially and temporally limited fashion. Thrombin, aPC and other coagulation proteases interact with protease activated receptors, initiating cellular signalling that regulates inflammation and tissue remodelling. a, activated; F, factor; PC, protein C.

Figure 2. Initiation, amplification and propagation of coagulation

Upon vessel wall injury and/or activation of endothelial cells, tissue factor (TF) is exposed to blood and binds to FVII or FVIIa, promoting FVII activation or enhancing its catalytic activity. The TF–FVIIa (extrinsic tenase) complex activates small amounts of FIX and FX. FXa associates with FVa to form the prothrombinase complex, which cleaves FII to generate a small amount of thrombin. This initiation phase is followed by the amplification phase, in which thrombin activates cell-surface (predominately platelet) bound FV and FVIII and platelet-bound FXI. FIXa binds to FVIIIa on negatively charged surfaces (predominately platelet-derived phospholipids), activating FX (intrinsic tenase) and initiating a burst of thrombin generation—the propagation phase. Thrombin has pleotropic functions including feedback inhibition, fibrin formation, platelet activation and signalling through protease activated receptors. a, activated; F, factor.

Figure 3. Potential mechanisms of PAR activation by thrombin and aPC

An example scheme of protease activated receptors (PARs) and the N-terminal sequences of human PAR1 and PAR3 depicting distinct cleavage sites for thrombin and activated protein C (aPC; arrows). The qualitatively distinct signalling mechanisms of thrombin and aPC can be attributed to the distinct proteolytic activation mechanisms of the G protein-coupled receptor (GPCR) N-terminus, resulting in protease-specific tethered ligands (shown in red for thrombin and blue for aPC) or induction of distinct protease-specific signalling complexes. PAR3 is not considered to be signalling competent and the function of the tethered ligands remains incompletely resolved. Activation of PARs might elicit protease-specific classical GPCR signalling by activation of individual PAR receptors (that is protomers) or ligand-specific PAR–PAR heterodimers. In addition, coagulation-protease-dependent signalling might engage non-PAR receptors, enabling biased signalling and thus leading to signalling diversity. Other proteases can cleave PARs at different sites, for example matrix metalloproteinase-1 cleaves PAR1 at Asp39 and neutrophil elastase cleaves PAR-1 at Leu45.

Figure 4. Expression of coagulation protease receptors in renal cells

Protease activated receptor (PAR) 2 is expressed on all types of human renal cells but is not expressed on murine podocytes, whereas PAR3 expression is predominantly restricted to podocytes. Endothelial protein C receptor (EPCR), which promotes protein C activation and activated protein C signalling, is expressed by glomerular endothelial and tubular epithelial cells. Tissue factor (TF) is expressed by podocytes and mesangial cells in humans, whether it is expressed on these cells in other species remains unproven. In various disease models (for example sepsis and diabetic nephropathy) TF expression is induced, but expression of thrombomodulin and EPCR is impaired.

Figure 5. Coagulation regulators in acute kidney injury

a | In acute glomerular injury, inflammation induces tissue factor (TF) expression on inflammatory cells recruited into the glomeruli or potentially on glomerular cells themselves, resulting in coagulation activation. FVa and the FXa complex assemble on these cells and enhance thrombin generation. FXa and thrombin induce glomerular cell dysfunction via protease activated receptor (PAR) 2 and PAR1, respectively. Thrombin signalling via PAR1 might involve transactivation of PAR2. Increased fibrin deposition contributes to the formation of glomerular crescents, which is inhibited by plasmin-mediated fibrinolysis. Inhibition of plasmin by plasminogen activator inhibitor-1 (PAI-1), which might be induced via the renin–angiotensin–aldosterone system (RAAS) or activated thrombin activatable fibrinolysis inhibitor (TAFIa), abolishes this effect. b | In acute tubular injury inflammation, for example in the context of ischaemia–reperfusion injury (IRI), induces TF expression on inflammatory cells or potentially on tubular epithelial cells themselves, triggering coagulation activation within the tubular compartment. Thrombin signalling via PAR1 leads to transforming growth factor β (TGF-β)-dependent tissue remodelling and tubular injury, whereas activated protein C (aPC) signalling via PAR1–endothelial protein C receptor (EPCR) inhibits TGF-β-dependent tubular fibrosis and preserves expression of YB-1 by restricting its ubiquitin-dependent degradation, thereby preventing tubular injury. Excess fibrin formation induces tubular injury, whereas moderate fibrin generation triggers fibrinolysis, leading to extracellular matrix (ECM) degradation and generation of the fibrin-derived peptide Bβ15–42, which blocks the interaction of fibrin with intercellular adhesion molecule 1 (ICAM-1). Moderate fibrin deposition, therefore, contributes to renal recovery. Green inhibitory arrows indicate that inhibition promotes repair; red inhibitory arrow indicates that inhibition promotes injury. a, activated; AT, antithrombin; F, factor; TM, thrombomodulin.

Figure 6. Coagulation regulators in chronic kidney injury

a | In chronic kidney disease, subclinical inflammation induces expression of tissue factor (TF) on glomerular cells or potentially on inflammatory cells recruited into the glomeruli, thus triggering coagulation activation. FXa and thrombin induce glomerular cell dysfunction via protease activated receptor (PAR) signalling, fibrin induction and extracellular matrix (ECM) deposition. In chronic diabetic kidney disease, thrombomodulin (TM)-dependent protein C activation is impaired resulting in exacerbated glucose toxicity and glomerular cell dysfunction. Impaired protein C activation also triggers mitochondrial localization of Bax and p66Shc, resulting in mitochondrial dysfunction in glomerular cells. Reconstitution of activated protein C (aPC) signalling, for example by exogenous administration, restores cellular function via endothelial protein C receptor (EPCR)–PAR1 signalling in endothelial cells and PAR3-mediated signalling in podocytes, thus preventing diabetic nephropathy. Independent of aPC generation, thrombomodulin inhibits complement activation via its lectin-like domain and ameliorates diabetic nephropathy. b | Chronic inflammation triggers fibrin generation in the tubulointerstitial compartment, contributing to tubular injury. Plasmin-mediated degradation of fibrin and ECM inhibits this process, but this tissue-protective mechanism is inhibited by thrombin-mediated activation of thrombin activable fibrinolysis inhibitor (TAFI). Green inhibitory arrow indicates that inhibition promotes repair; red inhibitory arrow indicates that inhibition promotes injury. a, activated; EC, endothelial cells; F, factor; P, podocytes; PAI-1 plasminogen activator inhibitor-1; ROS, reactive oxygen species; TGF-β, transforming growth factor β; tPA, tissue-type plasminogen activator.