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Cathepsin S Elicits Itch and Signals via Protease-Activated Receptors

2010; Elsevier BV; Volume: 130; Issue: 5 Linguagem: Inglês

10.1038/jid.2009.430

1523-1747

Vemuri B. Reddy, Steven G. Shimada, Parul Sikand, Robert H. LaMotte, Ethan A. Lerner,

Food Allergy and Anaphylaxis Research

protease-activated receptor Cathepsin S is a cysteine protease linked to inflammatory processes, including atherosclerosis and asthma. The possibility that this or other cysteine proteases might evoke itch or be part of a classical ligand-receptor signaling cascade has not been considered previously. We show here that human cathepsin S evokes itch and activates human protease-activated receptors (PARs) 2 and 4. The sensation of itch is mediated by two distinct non-overlapping populations of cutaneous nerve fibers that evoke comparable degrees of itch (Johanek et al., 2008Johanek L.M. Meyer R.A. Friedman R.M. et al.A role for polymodal C-fiber afferents in nonhistaminergic itch.J Neurosci. 2008; 28: 7659-7669Crossref PubMed Scopus (156) Google Scholar; Namer et al., 2008Namer B. Carr R. Johanek L.M. et al.Separate peripheral pathways for pruritus in man.J Neurophysiol. 2008; 100: 2062-2069Crossref PubMed Scopus (207) Google Scholar). One set of fibers, the mechano-insensitive population, is more responsive to histamine than to cowhage. The other set is mechanosensitive and is more responsive to cowhage than to histamine (Johanek et al., 2008Johanek L.M. Meyer R.A. Friedman R.M. et al.A role for polymodal C-fiber afferents in nonhistaminergic itch.J Neurosci. 2008; 28: 7659-7669Crossref PubMed Scopus (156) Google Scholar; Namer et al., 2008Namer B. Carr R. Johanek L.M. et al.Separate peripheral pathways for pruritus in man.J Neurophysiol. 2008; 100: 2062-2069Crossref PubMed Scopus (207) Google Scholar). Histamine is a classical mediator of itch and is associated with a wheal and flare. As most clinical itches do not have a wheal or flare and do not respond to antihistamines, histamine is not thought to contribute to most itches (Ikoma et al., 2006Ikoma A. Steinhoff M. Ständer S. et al.The neurobiology of itch.Nat Rev Neurosci. 2006; 7: 535-547Crossref PubMed Scopus (748) Google Scholar). Cowhage refers to a tropical legume or, in this case, the loose hairs that cover the pods of Mucuna pruriens and evoke itch. The active component of cowhage is mucunain, a cysteine protease that serves as a ligand for PARs 2 and 4 (Reddy et al., 2008Reddy V.B. Iuga A.O. Shimada S. et al.Cowhage evoked itch is mediated by a novel cysteine protease — a ligand of protease activated receptors.J Neurosci. 2008; 28: 4331-4335Crossref PubMed Scopus (193) Google Scholar). The identification of an endogenous mediator with properties similar to cowhage could lead to insights into non-histamine-mediated itch. We focused on human cathepsin S because it shares active site sequence homology with mucunain and is selectively up-regulated in human keratinocytes upon stimulation with interferon-gamma, consistent with a possible pruritic role in inflammatory skin disease (Schwarz et al., 2002Schwarz G. Boehncke W.H. Braun M. et al.Cathepsin S activity is detectable in human keratinocytes and is selectively upregulated upon stimulation with interferon-γ.J Invest Dermatol. 2002; 119: 44-49Crossref PubMed Scopus (49) Google Scholar). There are 15 human cathepsins, including 11 cysteine, 2 aspartic, and 2 serine proteases. Cathepsins were traditionally considered lysosomal proteases. It is now recognized that the broad expression and range of pH dependence of some cathepsins reveal that they have many functional roles, including tissue remodeling, metastasis and inflammation. Examples of cysteine cathepsin activities include cleavage of collagen by cathepsin L to generate endostatin (Felbor et al., 2000Felbor U. Dreier L. Bryant R.A. et al.Secreted cathepsin L generates endostatin from collagen XVIII.EMBO J. 2000; 19: 1187-1194Crossref PubMed Scopus (403) Google Scholar), an endogenous inhibitor of angiogenesis, and cleavage of the invariant chain in antigen-presenting cells by cathepsin S (Nakagawa et al., 1999Nakagawa T.Y. Brissette W.H. Lira P.D. et al.Impaired invariant chain degradation and antigen presentation and diminished collagen-induced arthritis in cathepsin S null mice.Immunity. 1999; 10: 207-217Abstract Full Text Full Text PDF PubMed Scopus (367) Google Scholar) as part of the inflammatory cascade. There are four PARs and they are members of the G-protein-coupled receptor family. Their identified endogenous activators are all serine proteases. These proteases trigger the activation of PARs by unmasking 'tethered ligand' sequences near the N-termini of the receptors. Certain kallikrein-related peptidases and mast cell tryptase, which are serine proteases, can activate PAR2 in vitro (Oikonomopoulou et al., 2006Oikonomopoulou K. Hansen K.K. Saifeddine M. et al.Kallikrein-mediated cell signalling: targeting proteinase-activated receptors (PARs).Biol Chem. 2006; 387: 817-824PubMed Google Scholar, Stefansson et al., 2008Stefansson K. Brattsand M. Roosterman D. et al.Activation of proteinase-activated receptor-2 by human kallikrein-related peptidases.J Invest Dermatol. 2008; 128: 18-25Crossref PubMed Scopus (169) Google Scholar), but they have not been demonstrated to be endogenous pruritic agents. Serine proteases and PAR2 have also been linked to barrier function (Hachem et al., 2006Hachem J.P. Houben E. Crumrine D. et al.Serine protease signaling of epidermal permeability barrier homeostasis.J Invest Dermatol. 2006; 126: 2074-2086Abstract Full Text Full Text PDF PubMed Scopus (168) Google Scholar). The presence of PAR2 on free nerve endings in the skin, keratinocytes and dorsal root ganglia link this receptor to itch and pain (Steinhoff et al., 2000Steinhoff M. Vergnolle N. Young S.H. et al.Agonists of proteinase-activated receptor 2 induce inflammation by a neurogenic mechnism.Nat Med. 2000; 6: 151-158Crossref PubMed Scopus (800) Google Scholar, Steinhoff et al., 2003Steinhoff M. Neisius U. Ikoma A. et al.Proteinase-activated receptor 2 mediates itch: a novel pathway for pruritus in human skin.J Neurosci. 2003; 23: 6176-6180PubMed Google Scholar; Shpacovitch et al., 2008Shpacovitch V. Feld M. Hollenberg M.D. et al.Role of protease-activated receptors in inflammatory responses, innate and adaptive immunity.J Leukoc Biol. 2008; 83: 1309-1322Crossref PubMed Scopus (154) Google Scholar). Data on PAR4 are more limited, revealing that it is expressed in rat dorsal root ganglia and its activation, at least in this species, appears to be anti-nociceptive (Asfaha et al., 2007Asfaha S. Cenac N. Houle S. et al.Protease-activated receptor-4: a novel mechanism of inflammatory pain modulation.Br J Pharmacol. 2007; 150: 176-185Crossref PubMed Scopus (102) Google Scholar). A PAR4 hexapeptide agonist has recently been reported to cause scratching in mice (Tsujii et al., 2008Tsujii K. Andoh T. Lee J.B. et al.Activation of proteinase-activated receptors induces itch-associated responses through histamine-dependent and independent pathways in mice.J Pharmacol Sci. 2008; 108: 385-388Crossref PubMed Scopus (44) Google Scholar). Cathepsin S and mucunain are endogenous and exogenous cysteine proteases, evoke similar itch and nociceptive sensations, and serve to activate PARs 2 and 4 (Figures 1 and 2; Reddy et al., 2008Reddy V.B. Iuga A.O. Shimada S. et al.Cowhage evoked itch is mediated by a novel cysteine protease — a ligand of protease activated receptors.J Neurosci. 2008; 28: 4331-4335Crossref PubMed Scopus (193) Google Scholar). The relative contribution of PAR2 versus PAR4 activation in cathepsin S-induced sensations is not addressed by these data. Cathepsin S has been implicated in neuropathic pain, manifested by increased gene expression in rat dorsal root ganglia following peripheral nerve injury. Such pain was lessened by a cathepsin S inhibitor (Barclay et al., 2007Barclay J. Clark A.K. Ganju P. et al.Role of the cysteine protease cathepsin S in neuropathic hyperalgesia.Pain. 2007; 130: 225-234Abstract Full Text Full Text PDF PubMed Scopus (109) Google Scholar). These observations suggest that cathepsin S may have an excitatory effect on neurons. As keratinocytes have been demonstrated to express cathepsin S (Schwarz et al., 2002Schwarz G. Boehncke W.H. Braun M. et al.Cathepsin S activity is detectable in human keratinocytes and is selectively upregulated upon stimulation with interferon-γ.J Invest Dermatol. 2002; 119: 44-49Crossref PubMed Scopus (49) Google Scholar), a role for this protease in keratinocyte-neuronal communication may be expected. Cathepsin S may contribute to the pruritus of inflammatory skin conditions, including atopic dermatitis and psoriasis, and could have a role in barrier function. Cathepsin S and other endogenous or exogenous cysteine proteases may activate PARs as a part of additional inflammatory processes. For example, Der p1, a mite cysteine protease associated with asthma, activates PAR2 (Asokananthan et al., 2002Asokananthan N. Graham P.T. Stewart D.J. et al.House dust mite allergens induce pro-inflammatory cytokines from respiratory epithelial cells: the cysteine protease allergen, Der p 1, activates protease-activated receptor (PAR)-2 and inactivates PAR-1.J Immunol. 2002; 169: 4572-4578Crossref PubMed Scopus (284) Google Scholar). Taken together, we suggest that exogenous and endogenous cysteine proteases interact with PARs as a part of signaling cascades in homeostasis and disease.Figure 2Recombinant human cathepsin S activates human protease-activated receptors (PARs). (a) Single-cell imaging of cathepsin S (2μM) induced responses in HeLa cells transfected with either PAR 2 or PAR 4 as measured by ratiometric calcium imaging in cells loaded with fura-2 as in Reddy et al., 2008Reddy V.B. Iuga A.O. Shimada S. et al.Cowhage evoked itch is mediated by a novel cysteine protease — a ligand of protease activated receptors.J Neurosci. 2008; 28: 4331-4335Crossref PubMed Scopus (193) Google Scholar. The responses to PARs 2 and 4 were blocked by the protease inhibitor E64 (10μM) as a control. Other controls included vector alone and PAR1. Cathepsin S was added as indicated. (b) Concentration–effect responses of cathepsin S in HeLa cells transiently transfected with PARs 2 and 4 as determined by ratiometric imaging of 20 cells per data point. Salmon sperm (SS) DNA as control. For both (a) and (b), the averages from at least three separate experiments were then combined±SD (see Supplementary material online).View Large Image Figure ViewerDownload (PPT) The authors state no conflict of interest. This work was supported in part by a Clinical and Translational Science Award, National Center for Research Resources (NCRR) Grant UL1 RR0249139 (a component of the National Institutes of Health (NIH)), NIH Grant P01 NS047300 (RHL, PI), and an agreement between Massachusetts General Hospital and Shiseido (Tokyo, Japan). Psychophysical experiments were approved by the Yale University Human Investigation Committee. The study was conducted according to the Declaration of Helsinki Principles. Participants gave their written informed consent. We thank Aurel Iuga and Barry Green for their input and technical assistance. Supplementary material is linked to the online version of the paper at http://www.nature.com/jid Download .pdf (.03 MB) Help with pdf files Supplementary Material and Methods