Rheumatoid arthritis regulation of synovial inflammation

The International Journal of Biochemistry&Cell Biology36(2004)372–378Medicine in focusRheumatoid arthritis:regulation of synovial inflammationSusan E.Sweeney,Gary S.Firestein∗Division of Rheumatology,Allergy,and Immunology,UCSD School of Medicine,9500Gilman Drive,La Jolla,CA92093,USAReceived1May2003;received in revised form26June2003;accepted16July2003AbstractRheumatoid arthritis(RA)is a systemic,inflammatory autoimmune disorder that presents as a symmetric polyarthritis as-sociated with swelling and pain in multiple joints,often initially occurring in the joints of the hands and feet.Articular inflam-mation causes activation and proliferation of the synovial lining,expression of inflammatory cytokines,chemokine-mediated recruitment of additional inflammatory cells,as well as B cell activation with autoantibody production.A vicious cycle of altered cytokine and signal transduction pathways and inhibition of programmed cell death contribute to synoviocyte and os-teoclast mediated cartilage and bone destruction.A combination of targeted interventions at various stages in the pathogenesis of RA will likely be required to control symptoms in certain patients with this complex and potentially disabling disease.The regulation of rheumatoid synovial inflammation will be reviewed,followed by a brief summary of the therapeutic implications of these advances,including strategies targeting key cytokines,signal transduction molecules,co-stimulatory molecules,B cells,chemokines,and adhesion molecules.©2003Elsevier Ltd.All rights reserved.Keywords:Rheumatoid arthritis;Inflammation;Cytokines1.IntroductionRheumatoid arthritis(RA)is a synovial inflamma-tory disease of unknown etiology that occurs in about 1%of the population.The onset is typically between ages30and55years with women nearly twice as likely to develop the disease.Clinically,RA manifests as a symmetric polyarthritis associated with swelling and pain in multiple joints,often initially occurring in the joints of the hands,wrists,and feet.It has been proposed that the initiating event in RA involves an infectious agent or other environmental exposure.Lo-cal induction of innate immunity may activate cells in ∗Corresponding author.Tel.:+1-858-822-0480;fax:+1-858-534-2606.E-mail address:gfirestein@(G.S.Firestein).the synovial lining and prepare the synovium for the subsequent adaptive immune response in genetically susceptible individuals(Firestein&Zvaifler,1990). The resultant inflammatory arthritis can cause signifi-cant joint deformity and disability.Although the joint is the main target of the pathologic immune response, RA can also affect many other organs.Systemic man-ifestations,including subcutaneous nodules,pleuritis, pericarditis,and vasculitis,contribute to morbidity and mortality in long-standing disease.Over the past decade,significant advances in understanding the pathogenesis of RA and synovial inflammation have led to the identification of novel therapeutic targets.A combination of targeted interventions at various stages of RA will likely be required to control symptoms in certain patients with this complex and disabling disease.1357-2725/$–see front matter©2003Elsevier Ltd.All rights reserved. doi:10.1016/S1357-2725(03)00259-0S.E.Sweeney,G.S.Firestein /The International Journal of Biochemistry &Cell Biology 36(2004)372–3783732.Pathogenesis2.1.Cellular infiltration in the rheumatoid synoviumThe initiating event in RA is followed by the in-duction of an immune response that results in in-flammation in the lining of the joint,known as the synovial membrane.The normal synovium is a rela-tively acellular structure with an intimal lining com-prised of macrophage-and fibroblast-like cells known as synoviocytes.In RA,the intimal lining becomes hyperplastic due to the ingress of the former and in-creased proliferation of the latter (Fig.1).Mononu-clear cells,most notably CD4+T cells,infiltrate the sublining of the synovium.B cells and dendritic cells form aggregates with T cells and tissue macrophages and,in approximately 20%of patients,form struc-tures that resemble lymph node follicles.Local im-munoglobulin production of autoantibodies occurs and results in immune complex formation.The autoanti-bodies recognize articular antigens,such as typeIIFig.1.Rheumatoid arthritis synovial tissue.The synovial intimal lining is hyperplastic due to an increase in macrophages and fibroblasts.The sublining region contains increased numbers of mononuclear cells,including T cells that can form lymphoid aggregates.Increased numbers of blood vessels is also observed.collagen and proteoglycans,or bind to the Fc portion of normal IgG,also known as rheumatoid factor.The T cells that accumulate in the synovium are relatively quiescent,although some products like interleukin-17(IL)and interferon gamma (IFN ␥)are produced in physiologically relevant concentrations.The majority of T cell products,however,are present at relatively low levels compared with typical antigen-dependent in vivo responses.In contrast,fibroblast-like synovio-cytes (FLS)and macrophage (M )products like IL-1,tumor necrosis factor alpha (TNF ␣),IL-8,and many others are abundant in the RA synovium and likely contribute to synovial inflammation.2.2.Cytokine networks in RABased on the cytokine profile of RA,paracrine and autocrine networks likely participate in disease perpetuation.M and FLS in the intimal lining pro-duce factors that activate adjacent cells and enhance synovial inflammation.For instance,IL-1and TNF ␣,both produced by the synovial M ,can stimulate374S.E.Sweeney,G.S.Firestein /The International Journal of Biochemistry &Cell Biology 36(2004)372–378fibroblast proliferation and increase production of IL-6,granulocyte-macrophage colony-stimulating factor (GM-CSF),IL-8,and other chemokines,as well as enzymes that result in joint destruction.These cytokines,in turn,can activate macrophages in the en-vironment and lead to continued cytokine production.This creates a positive feedback mechanism between the FLS and M that perpetuates synovial inflamma-tion (Fig.2).Although the basic rules of the cytokine profile remain essentially the same today,many addi-tional factors have been added to the network in recent years.For instance,a new member of the IL-1family,IL-18,results in M activation,IFN ␥induction,and local Th1differentiation of T cells (Gracie,Forsey,Chan,&et al.,1999).IL-15,produced by the M ,can induce TNF ␣production through activation of syn-ovial T cells (Ziolkowska,Koc,Luszczykiewicz,&et al.,2000).Modern molecular techniques have clari-fied the T cell contribution to the local milieu.Synovial T cells are often biased toward the Th1phenotype,regulate delayed type hypersensitivity,and express the chemokine receptors CCR5and CXCR3(Loetscher,Uguccioni,Bordoli,&et al.,1998).IL-17is an exam-ple of a Th1cytokine produced in the rheumatoid joint that synergizes with IL-1and TNF ␣to causefibroblastFig.2.Cytokine networks in rheumatoid arthritis.Fibroblast-and macrophage-like synoviocytes (FLS and M ,respectively)are located in close proximity in the synovial intimal lining.Activation occurs via paracrine pathways,autocrine pathways,as well as direct cell contact.Synergy between cytokines in the environment results in increased joint destruction.In RA,positive (+)feedback loops predominate,but inhibitory (−)feedback has also been demonstrated.activation and bone destruction (Chabaud,Durand,Buchs,&et al.,1999).Cell–cell contact dependent interactions between T cells and other cell types in RA can also enhance cytokine and metalloproteinase production resulting in increased joint destruction.Synergistic interactions between the various cytokines in the inflamed joint also result in enhancement of metalloproteinase production and joint destruction.2.3.Activation of signal transduction pathways and transcription factors in RAInflammatory cytokines activate intracellular sig-nal transduction and transcription factor pathways in the synovium (Fig.3)(Firestein,2003).NF-␬B,mitogen-activated protein (MAP)kinases,and transcription factor activator protein-1(AP-1)are especially important in the regulation of synovial in-flammation.NF-␬B is activated in the RA synovium and controls transcription of inflammatory genes im-plicated in RA,including TNF ␣,IL-1,IL-6,IL-8,cyclooxygenase-2(COX-2),inducible nitric oxide sy-thetase (iNOS),and intercellular adhesion molecule-1(ICAM-1)(Han,Boyle,Manning,&et al.,1998).Ex-posure to inflammatory cytokines or other activationS.E.Sweeney,G.S.Firestein/The International Journal of Biochemistry&Cell Biology36(2004)372–378375Fig.3.Signal transduction cascades in rheumatoid arthritis.Sig-nal transduction through the MAP kinase and NF-␬B pathways is activated by extracellular stimuli such as cytokines or inflam-matory mediators.In the MAP kinase cascade(left),this results in activation and phosphorylation of MAP kinase kinase kinases (MAPKKK),followed by phosphorylation of MAP kinase kinases (MAPKK),which in turn results in MAP kinase phosphoryla-tion of JNK,p38,or ERK.This results in transcription factor activation and subsequent production of cytokines and/or matrix metalloproteinases(MMPs).NF-␬B is engaged after extracellular stimuli result in activation of I␬B kinases that phosphorylate I␬B, resulting in its degradation.Subsequently,NF-␬B translocates to the nucleus and results in transcription of various cytokines and inflammatory mediators.signals leads to phosphorylation of the natural in-hibitor of NF-␬B,I␬B,via activation of the I␬B kinase(IKK1and IKK2)signal complex in the syn-oviocyte.IKK2activation of FLS results in phos-phorylation of I␬B,which is subsequently degraded, allowing the nuclear localization of NF-␬B.Synovio-cyte IKK2activation is both necessary and sufficient for NF-␬B mediated induction of IL-6,IL-8,and ICAM-1gene expression.A second pathway impli-cated in RA,the MAP kinases,includes the extracel-lular signal-regulated kinase(ERK),c-Jun N-terminal kinase(JNK),and p38families.All three are ex-pressed and phosphorylated in rheumatoid synovial tissue and can increase cytokine and metallopro-teinase production(Schett,Tohidast-Akrad,Smolen, &et al.,2000).Exposure to inflammatory cytokines induces rapid phosphorylation of these kinases in cul-tured FLS.Kinases such as MKK3,MKK4,MKK6, and MKK7,located upstream of the MAP kinases, are also activated in rheumatoid synovium and form signaling complexes that respond to specific environ-mental stimuli with an appropriate change in gene expression.One of the key transcription factors activated by the MAP kinase JNK is activator protein(AP-1).The AP-1binding site is present in the promoter regions of many genes that participate in the pathogenesis of RA, including TNF␣and various metalloproteinases.Af-ter synoviocytes are stimulated with pro-inflammatory cytokines,the c-Jun component of AP-1is phospho-rylated by JNK and the activated transcription factor induces massive expression of many proteases and cy-tokines.Expression of c-Jun and c-Fos is increased in RA synovium and is localized to the nuclei of synovial lining cells.2.4.Recruitment of inflammatory cells into the rheumatoid synoviumOne of the cardinal features of rheumatoid synovitis is the recruitment and retention of inflammatory cells in the sublining region.This process is mediated by attractant chemokines as well as adhesion molecules on vascular endothelium and counter-receptors on circulating leukocytes that draw cells to sites of in-flammation(Szekanecz&Koch,2001).As a result, the cytokine and chemokine milieu in RA contributes to the specific cell phenotypes that collect in syn-ovial tissue.For instance,T cells exhibiting the Th1 phenotype are abundant in the joint.This is,in part, accounted for by their expression of the chemokine receptors,CXCR3and CCR5,and response to MCP-1 and RANTES produced by the synovium(Szekanecz, Kunkel,Strieter,&et al.,1998).IL-8is a potent chemotactic agent for neutrophils and is perhaps the most abundant chemokine in RA synovial effusions. This results in the marked influx of neutrophils into the rheumatoid joint.As noted above,chemoattractants are only part of the cell recruitment story(Szekanecz&Koch, 2001).In addition,circulating leukocytes must bind to specific adhesion receptors expressed on syn-ovial blood vessels.This process is mediated by the pro-inflammatory milieu,with induction of adhesion proteins like ICAM-1,E-selectin,and vascular cell adhesion molecule-1(VCAM-1)on the synovial mi-crovasculature.ICAM-1expression on the synovial M and FLS is also increased by stimulation with376S.E.Sweeney,G.S.Firestein/The International Journal of Biochemistry&Cell Biology36(2004)372–378TNF␣or IL-1and can help retain cells in the tissue by anchoring them to the extracellular matrix.ICAM-1 binds to counter receptor LFA-1and mediates cos-timulation and adhesion.VCAM-1is expressed on rheumatoid endothelial cells,lining cells,and cultured FLS.Levels are also increased by exposure to inflam-matory cytokines such as TNF␣,IL-1,IL-4,and IFN␥. Immigration of inflammatory cells into the joint could not occur without an exuberant vasculature to both supply the tissue with nutrients and provide sufficient endothelial cell surface to bind circulat-ing leukocytes.Neovascularization of rheumatoid synovium plays a major role in both of these pro-cesses and occurs under the influence of hypoxia and inflammatory stimulation.Certain factors that con-tribute to angiogenesis,including vascular endothelial growth factor(VEGF),fibroblast growth factor(FGF), TNF␣,and soluble adhesion receptors(e.g.soluble VCAM-1),are expressed by the intimal lining.In ad-dition to supporting the growth of new blood vessels, these factors also can suppress involution of synovial microvasculature(Szekanecz&Koch,2001).2.5.Destruction of cartilage and bone Destruction of the extracellular matrix leads to significant disability in patients with RA.Current concepts of joint destruction suggest that distinct mechanisms contribute to bone and cartilage dam-age.Cartilage destruction is mediated in large part through the elaboration of proteases by synoviocytes and cellular invasion into the matrix.Metallopro-teinases and aggrecanases,induced by IL-1,TNF␣, and IL-17,play a key role in this process.In addition, RA FLS can be permanently altered in the inflam-matory synovial environment.Oxidative stress and altered DNA repair potentially cause mutations in cer-tain key genes,such as the p53tumor suppressor,and can increase synoviocyte proliferation and cartilage invasion(Tak,Zvaifler,Green,&et al.,2000).Al-though synoviocytes and chondrocytes are involved in cartilage destruction,it is now known that bone ero-sion is mediated by the RANK/RANKL system.The RANKL receptor,RANK,is expressed on osteoclasts, osteoclast precursors,and chondrocytes.RANKL is expressed by various cells in rheumatoid synovium, including T cells and synoviocytes,and induces os-teoclast maturation and activation.The soluble decoy receptor to RANKL,osteoprotegerin(OPG),inhibits bone resorption and osteoclast function.Adminis-tration of OPG to rats with adjuvant arthritis blocks bone destruction.In most animal models,OPG has minimal effect on inflammation or cartilage damage (Kong,Feige,Sarosi,&et al.,1999).3.TherapyOne of the most exciting aspects of translational research in RA is that these studies have directly led to the introduction of new therapies(Moreland, 2001).For instance,elaboration of the cytokine net-works contributed to the development of the TNF inhibitors(Moreland,&et al.,1997)(etanercept, infliximab,and adalimumab)and use of the IL-1re-ceptor antagonist(Gabay&Arend,1998).The TNF inhibitors mainly act by neutralizing ambient TNF␣in the blood and tissues.About a third of patients have a dramatic response to these agents;of the re-maining patients,approximately half exhibit a partial response and the remaining patients have minimal improvement.Additional promising cytokine directed therapeutic targets include IL-6,IL-15,and IL-18. Preliminary studies involving an anti-IL-6receptor antibody(MRA)suggest that it is approximately as effective as the TNF-directed approaches(Wendling, Racadot,&Wijdenes,1993).IL-18can be targeted with IL-18binding protein that could inhibit M ac-tivation and the Th1bias of the synovium.Blockade of IL-15is being targeted to decrease T cell activation and macrophage-mediated TNF␣production. Drawbacks of cytokine targeted therapy include the expense(usually US$10,000–20,000per year)as well as the requirement for parenteral administration of proteins.Identification of intracellular molecules important in synovial inflammation in RA has led to the targeting of the signal transduction MAP ki-nase and NF-␬B pathways(Firestein,2003).Orally bioavailable small molecule inhibitors of p38MAPK have demonstrated promising results in various ani-mal models of arthritis with both anti-inflammatory effects and decreased joint damage.A selective JNK inhibitor decreased inflammation but more impres-sively provided protection against bone and carti-lage destruction.Targeting the NF-␬B pathway via intra-articular therapy with adenoviral IKK2dominantS.E.Sweeney,G.S.Firestein/The International Journal of Biochemistry&Cell Biology36(2004)372–378377negative constructs has also shown promise in an-imal models.Small molecule inhibitors of IKK2 are effective in adjuvant arthritis in rats as well as collagen-induced arthritis in mice.The primary con-cern with this approach is whether host defense will be unacceptably crippled by NF-␬B blockade.T cell directed therapy has been targeted to block costimulatory molecules such as CTLA4/B7with CTLA4-Ig fusion protein.Preliminary studies in hu-Table1Potential biologic targets for RA therapyTarget Development status andefficacyT cellCTLA4-Ig Phase2:ACR20,60%;ACR50,37%aB cellRituximab Phase2:ACR20,80%;ACR50,50% CytokinesTNF inhibitors ApprovedIL-1Ra ApprovedIL-6receptor antibody Phase2:ACR20,78%;ACR50,40%IL-15N/AIL-18binding protein N/ASignal transductionp38,JNK,NF-␬B inhibitors p38inhibitors in phase1 Osteoprotegerin N/AChemokine or chemokine receptorsCCR1Phase1CCR5N/AIL-8inhibitors N/AAdhesion molecule blockadeICAM-1Phase2:lack of efficacy Inhibitors of angiogenesisAnti␣v␤3N/A Metalloproteinase inhibitorsTrocade Phase3:lack of efficacy Induction of apoptosisFas ligand N/ATRA8(anti-TRAIL-R2/DR5mAb)N/AOtheriNOS N/ACOX-2inhibitors ApprovedN/A:not available.a Percentage of patients with ACR20and ACR50response rates(i.e.20and50%decrease in disease activity by American College of Rheumatology criteria).mans are promising and suggest that this approach might be additive with TNF inhibitors.B cell tar-geted therapy with rituximab,which binds to CD20, was effective in a phase II clinical trial,although it required concomitant treatment with methotrex-ate or cyclophosphamide(Vita,2002).Cell adhe-sion molecules ICAM-1,␣4/␤1integrins,and CD11 are also targets for interruption of co-stimulation and migration of inflammatory cells into the joint. Chemokine receptor antagonists have been designed to block CCR1,RANTES,and IL-8for the purpose of decreasing migration of inflammatory cells into the bined angiogenesis and adhesion in-hibitors such as anti-␣v/␤3are also under evaluation. Metalloproteinase inhibitors have been tested in an attempt to prevent joint damage.Unfortunately, these drugs have thus far demonstrated limited effi-cacy and have been plagued by side effects,in part related to excessivefibrosis.Enzymes such as iNOS and COX-2represent additional therapeutic targets to inhibit generation of inflammatory mediators and joint destruction.OPG might prevent or heal bone erosions by blocking the differentiation and activation of osteoclasts.Medical synovectomy could be used to effectively debulk the synovium in RA by induc-ing apoptosis.Induction of cell death with Fas ligand or TNF related apoptosis inducing ligand receptor2 (TRAIL-R2)represents future approaches to alter the course of this disease.A summary of the potential fu-ture therapeutic targets for RA is outlined in Table1.4.ConclusionThe extraordinary complexity of RA provides many opportunities for therapeutic intervention.Disease heterogeneity and variable mechanisms of synovitis occurring over a decade-long process suggest that combinations or individually tailored therapy might be needed to control disease.While daunting,application of translational research to the development of new treatment paradigms based on disease mechanisms is a rational approach to this unmet medical need. ReferencesChabaud,M.,Durand,J.M.,&Buchs,N.et al.,(1999).Human interleukin-17:A T cell derived proinflammatory378S.E.Sweeney,G.S.Firestein/The International Journal of Biochemistry&Cell Biology36(2004)372–378cytokine produced by the rheumatoid synovium.Arthritis and Rheumatism,42,963–970.Firestein,G.S.(2003).Evolving concepts of rheumatoid arthritis.Nature,423,356–361.Firestein,G.S.,&Zvaifler,N.J.(1990).How important are T cells in chronic rheumatoid synovitis?Arthritis and Rheumatism,33, 768–773.Gabay, C.,&Arend,W.P.(1998).Treatment of rheumatoid arthritis with IL-1inhibitors.Springer Seminars in Immunopathology,20,229–246.Gracie,J.A.,Forsey,R.J.,Chan,W.L.,Gilmour,A.,Leung,B.P.,Greer,M.R.,Kennedy,K.,Carter,R.,Wei,X.Q.,Xu,D., Field,M.,Foulis,A.,Liew,F.Y.,&McInnes,I.B.(1999).A proinflammatory role for IL-18in rheumatoid arthritis.Journal of Clinical Investigation,104,1393–1401.Han,Z.,Boyle, D.L.,Manning, A.M.,&Firestein,G.S.(1998).AP-1and NF-␬B regulation in rheumatoid arthritis and murine collagen-induced arthritis.Autoimmunity,28,197–208.Kong,Y.Y.,Feige,U.,Sarosi,I.,Balon,B.,Tafuri,A.,Morony, S.,Capparelli,C.,Li,J.,Elliott,R.,McCabe,S.,Wong,T., Campagnuolo,G.,Moran,E.,Bogoch,E.R.,Van,G.,Nguyen, L.T.,Ohashi,P.S.,Lacey,D.L.,Fish,E.,Boyle,W.J.,& Penninger,J.M.(1999).Activated T cells regulate bone loss and joint destruction in adjuvant arthritis through osteoprotegerin ligand.Nature,402,304–309.Loetscher,P.,Uguccioni,M.,Bordoli,L.,Baggiolini,M.,Moser,B.,Chizzolini,C.,&Dayer,J.M.(1998).CCR5is characteristicof Th1lymphocytes.Nature,391,344–345.Moreland,L.W.,Baumgartner,S.W.,Schiff,M.H.,Tindall,E.A., Fleischmann,R.M.,Weaver,A.L.,Ettlinger,R.E.,Cohen,S., Koopman,W.J.,Mohler,K.,Widmer,M.B.,&Blosch,C.M.(1997).Treatment of rheumatoid arthritis with a recombinanthuman tumor necrosis factor receptor(p75)-Fc fusion protein.The New England Journal of Medicine,337,141–147. Moreland,L.W.(2001).Potential biologic agents for treating rheumatoid arthritis.Rheumatic Diseases Clinics of North America,27,445–491.Schett,G.,Tohidast-Akrad,M.,Smolen,J.S.,Schmid, B.J., Steiner,C.W.,Bitzan,P.,Zenz,P.,Redlich,K.,Xu,Q.,& Steiner,G.(2000).Activation,differential localization,and regulation of the stress-activated protein kinases,extracellular signal-regulated kinase,c-Jun N-terminal kinase,and p38 mitogen-activated protein kinase,in synovial tissue and cells in rheumatoid arthritis.Arthritis and Rheumatism,43,2501–2512. Szekanecz,Z.,&Koch,A.E.(2001).Update on Synovitis.Current Rheumatology Reports,3,53–63.Szekanecz,Z.,Kunkel,S.L.,Strieter,R.M.,&Koch,A.E.(1998).Chemokines in rheumatoid arthritis.Springer Seminars in Immunopathology,20,115–132.Tak,P.P.,Zvaifler,N.J.,Green,D.R.,&Firestein,G.S.(2000).Rheumatoid arthritis and p53:How oxidative stress might alter the course of inflammatory diseases.Immunology Today,21, 78–82.Vita,S.D.et al.,(2002).Efficacy of selective B cell blockade in the treatment of rheumatoid arthritis:Evidence for a pathogenetic role of B cells.Arthritis and Rheumatism,46,2029–2033. Wendling,J.M.,Racadot,E.,&Wijdenes,J.(1993).Treatment of severe rheumatoid arthritis by anti-interleukin6monoclonal antibody.Journal of Rheumatology,20,259–262. Ziolkowska,M.,Koc, A.,Luszczykiewicz,G.,Ksiezopolska-Pietrzak,K.,Klimczak, E.,Chwalinska-Sadowska,H.,& Maslinski,W.(2000).High levels of IL-17in rheumatoid arthritis patients:IL-15triggers in vitro IL-17production via cyclosporin A-sensitive mechanism.Journal of Immunology, 164,2832–2838.。