69 Etiology and Pathogenesis of Rheumatoid Arthritis
Evidence of autoimmunity, including high serum levels of autoantibodies such as rheumatoid factors and anticitrullinated protein antibodies, can be present for many years before the onset of clinical arthritis.
Rheumatoid arthritis (RA) is the most common inflammatory arthritis, affecting from 0.5% to 1% of the general population worldwide. Although the prevalence is surprisingly constant across the globe, regardless of geographic location and race, there are some exceptions. For instance, in China the occurrence of RA is somewhat lower (≈0.3%), whereas it is substantially higher in other groups such as the Pima Indians in North America (≈5%). Because of its prevalence and the ready accessibility of joint samples for laboratory investigation, RA has served as a useful model for the study of all inflammatory and immune-mediated diseases. As such, the information gleaned from these studies provides new and unique insights into the mechanisms of normal immunity.
Although RA is primarily considered a disease of the joints, abnormal systemic immune responses are evident and can cause a variety of extra-articular manifestations. These manifestations clearly show that RA has features of a systemic disease that can involve many organs. In some cases, autoantibody production with the formation of immune complexes that fix complement contribute to these extra-articular findings. One of the mysteries of RA is why the synovium is the primary target, although the unique structure of its vascular bed could provide an environment that is ideal for innate and adaptive immune responses.
Although the precise causes of RA remain uncertain, environmental and genetic influences clearly participate. Clues have been provided by detailed immunogenetic studies and the observation that underlying autoimmunity antedates onset of arthritis by up to a decade. Progress in understanding the pathogenesis has been even more robust: The roles of small-molecule mediators of inflammation (e.g., arachidonic acid metabolites), autoantibodies, cytokines, growth factors, chemokines, adhesion molecules, and matrix metalloproteinases (MMPs) have been carefully defined. Synovial cells can exhibit behavior resembling a localized tumor that invades and destroys articular cartilage, subchondral bone, tendons, and ligaments. Irreversible loss of articular cartilage and bone begins soon after the onset of RA, and early interventions can probably improve long-term outcomes. Increased appreciation of how comorbidities, especially cardiovascular disease and accelerated atherosclerosis, can affect mortality has also led to attempts to suppress synovial and systemic inflammation.
The etiology and pathogenesis of RA are complex and multifaceted. A variety of predetermined (genes) and stochastic (random events and environment) factors contribute to susceptibility and pathogenesis. As an introduction, a summary of how these mechanisms interact to create and perpetuate RA is shown in Figure 69-1. Individual mechanisms are discussed in greater detail throughout the chapter.
Figure 69-1 Schematic diagram of disease mechanisms that likely occur in rheumatoid arthritis. Innate immunity could activate fibroblast-like synoviocytes (FLS), dendritric cells (DC), and macrophages (MΦ) in the earliest phases in individuals with underlying immune hyper-reactivity as evidenced by the production of autoantibodies. The genetic makeup of an individual including the presence of certain gene polymorphisms in genes that regulate immune responses and environmental exposures are both required. Chronic inflammation leads to citrullination of proteins in a variety of sites including mucosal surfaces such as the lungs or the joint. In a genetically susceptible individual, a breakdown of tolerance can occur with the formation of anticitrullinated protein antibodies. DCs can migrate to the central lymphoid organs to present antigen and activate T cells, which can in turn activate B cells. These lymphocytes can migrate back to the synovium and enhance adaptive immune responses in the target organ. In addition, repeated activation of innate immunity can directly lead to chronic inflammation and possibly antigen presentation in the synovium. In the latter phases of disease, many cell types activate osteoclasts (OC) through the receptor activator of nuclear factor κB (NFκB)/receptor activator of NFκB ligand (RANK/RANKL) system, although FLS and T cells likely provide the greatest stimulus. Autonomous activation of FLS might also contribute to this process.
The initiation of RA probably begins years before the onset of clinical symptoms. This process involves certain specific genes that can help break tolerance and lead to autoreactivity. It is likely that the earliest phases are marked by repeated activation of innate immunity (see Figure 69-1).1 Cigarette smoke, bacterial products, viral components, and other environmental stimuli can contribute to these responses. This process probably occurs often in normal individuals but is self-limited. In individuals, a predetermined propensity for immune hyper-reactivity or autoreactivity might lead to a different outcome. The genome of these individuals might encode for a variety of genes implicated in RA including class II major histocompatibilty complex (MHC) genes, protein tyrosine phosphatase-22 (PTPN22), cytokine promoter polymorphisms, signal transduction gene polymorphisms, population-specific genes (e.g., PADI4 in Japanese or Koreans), and other undefined genes. Abnormal T cell selection could also contribute by allowing autoreactive T cells to escape deletion. The environmental stresses can lead to post-transcriptional modification of proteins, especially citrullination of arginine residues, in mucosal surfaces or the synovium. Although this commonly occurs without sequelae in normal individuals, people with a propensity for RA can develop antibodies against these modified proteins with production of rheumatoid factors (RFs) and anticitrullinated protein antibodies (ACPAs).
Activation of synovial innate immunity can also increase vascular leakage in the synovium, production of chemoattractants that recruit immune cells to the joint, and processing of antigens by dendritic cells. Antigen presentation can potentially occur in the synovial germinal centers or, more commonly, in central lymphoid organs after the loaded dendritic cells migrate via the lymphatics. Naïve T cells can then be activated through interactions with the T cell receptor and co-stimulatory signals. T cells can help B cells produce pathogenic antibodies and/or migrate to the joint, where they can influence other cells through the production of cytokines such as interleukin (IL)-17 or through cell contact mechanisms that do not require a specific antigen. Although it is uncertain what transforms subclinical inflammation to symptomatic arthritis, this process can take up to a decade before it reaches fruition.
Ultimately, a destructive phase proceeds, which can have antigen-dependent and -independent mechanisms and is mediated by mesenchymal elements such as fibroblasts and synoviocytes. Bone erosions are subsequently caused by osteoclasts, whereas cartilage dissolution results from proteolytic enzymes produced by synoviocytes in the pannus or synovial fluid neutrophils. Anti-inflammatory mechanisms such as soluble TNF receptors, suppressive cytokines, cytokine binding proteins, protease inhibitors, lipoxins, antioxidants, antiangiogenic factors, and natural cytokine antagonists are not present in sufficient concentrations to truncate the inflammatory and destructive process. The only way to suppress this response is through therapeutic interventions that either modulate pathogenic cells or neutralize the effector molecules produced by the rheumatoid process, or restore tolerance.
The heterogeneity of mechanisms provides an explanation for the unpredictable response to therapeutic agents and also allows clinicians to consider new therapeutic targets to either prevent RA or interfere with the immunologic, inflammatory, or destructive components as separate but interrelated entities. Each of these mechanisms is discussed in detail later. Brief summaries are also provided intermittently to help guide the reader through this complex maze.
Although the etiology of RA remains unknown, a variety of studies suggest that the interaction of environmental and genetic factors is responsible; either one is necessary but not sufficient for full expression of the disease. The most compelling example for a genetic component is in monozygotic twins, in whom the concordance rate is perhaps 12% to 15% when one twin is affected, compared with 1% for the general population. The fact that concordance is not higher provides key evidence that other influences such as the environment, epigenetics, or even microchimerism from maternal-fetal transfer might be as important as or even more important than the genetic component. The risk for a fraternal twin of a patient with RA is also high (≈2% to 5%) but similar to the rate for other first-degree relatives.
Although the immunogenetics is, at best, incompletely understood, one of the best-studied and perhaps most influential genetic risk factor is the class II MHC haplotype of an individual. PTPN22 and PADI4 increase risk in some racial and ethnic groups, but not all. Genome-wide screens have implicated at least 35 genes, many of which are involved with immune function. However, most have a relatively modest contribution and the susceptibility polymorphism confers only a 1.1- or 1.2-fold increase. Combinations of genes can clearly interact with one another, and a 45-fold increase in risk is conferred by a combination of HLA-DR, PTPN22, and the TRAF1-C5.1 This combination, however, is found in less than 1% of individuals with RA. The RA-associated alleles identified to date contribute approximately 40% of total genetic susceptibility. Additional progress in understanding the role of genes in RA including rare variants that might be more important than some common polymorphisms will require sophisticated bioinformatics to clarify how individual alleles contribute to susceptibility, severity, and response to targeted therapies.
The structure of class II MHC molecules on antigen-presenting cells is associated with increased susceptibility and severity of RA and accounts for about 40% of the genetic influence. A genetic link between HLA-DR and RA was initially described in the 1970s with the observation that HLA-DR4 occurred in 70% of RA patients, compared with about 30% of controls, giving a relative risk of having RA to those with HLA-DR4 of approximately 4 to 5.
The susceptibility to RA is associated with the third hypervariable region of DRβ-chains, from amino acids 70 through 74. The epitope is glutamine-leucine-arginine-alanine-alanine (QKRAA), a sequence found in DR4 and DR14, in addition to some DR1β-chains. The “susceptibility epitope” (SE) on DR4β-chains with the greatest association with RA are DRB*0401, DRB*0404, DRB*0101, and DRB*1402 (Table 69-1). Up to 96% of patients with RA exhibit the appropriate HLA-DR locus in some populations.2 In certain ethnic and racial groups, however, the association with QKRAA is not as prominent or is not associated. The QKRAA epitope might also predict the severity of established RA, with a greater prevalence of extra-articular disease and erosions in patients with two copies. Other HLA genes such as DRB*1301 contain the DERAA sequence and are associated with decreased susceptibility to RA.3
|Old Nomenclature (HLA-DRB1 Alleles)||Current Nomenclature||Association with Rheumatoid Arthritis|
|HLA-DR2||1501, 1502, 1601, 1602||−|
|HLA-DR5||1101-1104, 1201, 1202||−|
|HLA-DRw13||1301-1304||1301 associated with protection|
Modified from Weyand CM, Hicok KC, Conn DL, Goronzy JJ: The influence of HLA-DRB1 genes on disease severity in rheumatoid arthritis, Ann Intern Med 117:801, 1992.
One intriguing possibility that could account for some patients who do not fit within this paradigm is microchimerism.4 Maternal cells expressing the SE can survive and persist in the circulation throughout adulthood. These noninherited maternal antigens (NIMAs) could then confer increased risk of disease in the children of SE-expressing women.
The region associated with RA (QKRAA) primarily faces away from the antigen-binding cleft of the DR molecule that determines the specificity of peptides presented to CD4+ helper T cells. Attempts to elute peptides from the binding pocket of RA-associated alleles have not revealed a specific antigen that is either unique to or associated with RA. The precise function of the SE is uncertain, but it could also play a role in shaping the T cell repertoire in the thymus or altering intracellular HLA-DR trafficking and antigen loading. QKRAA could serve as an autoantigen due to molecular mimicry in some situations because some xenoproteins such as gp110 from the Epstein-Barr virus also include this sequence.
The shared epitope might not be an independent risk factor for RA but instead a marker for immunoreactivity and anticitrullinated protein antibodies (ACPAs).5 In a large series of patients with early undifferentiated inflammatory arthritis, one-third of patients met criteria for RA within 1 year. Progress to RA occurred regardless of HLA-DR genotype if patients were anticitrullinated protein (anti-CP) positive. When patients were stratified according to ACPA, the shared epitope did not make an additional contribution to progression from undifferentiated arthritis to RA. The shared epitope probably contributes to immune hyper-reactivity, but ACPAs are more closely associated with RA. In other studies, however, the presence of the shared epitope and ACPAs together is associated with even greater disease severity.
The genetic influence on RA has also led to studies evaluating non-MHC genes. Single nucleotide polymorphisms (SNPs) in promoter regions, coding regions, or areas with no known function have been extensively investigated in RA with a variety of methods including genome-wide association studies. Table 69-2 shows some of the SNPs and microsatellites that have been associated with RA. The relative contribution for most is modest, and variations in technique, stage of disease, and patient populations result in some disagreement among various reports.
|Gene||Odds Ratio for Risk Alleles||Comment|
|PTPN22||≈2 fold||Not in Asian populations|
|PADI4||≈2 fold||Primarily in Asian populations|
|TRAF1-C5||>1.2 fold; <2 fold|
|STAT4||>1.2 fold; <2 fold|
|TNFAIP3||>1.2 fold; <2 fold|
|IL2/21||>1.2 fold; <2 fold|
Other genes with odds ratio >1.0 and <1.2: CTLA4, CD40, CCL21, CD244, IL2Rb, TNFRSF14, PRKCQ, PIP4K2C, IL2RA, AFF3, REL, BLK, TAGAP, CD28, TRAF6, PTPRC, FCGR2A, PRDM1, CD2-CD58, IRF5, CCR6, CCL21, IL6ST, RBPJ.
Given the importance of cytokines in RA (see following), it is not surprising that many studies have focused on these genes. The most intriguing evidence relates to tumor necrosis factor (TNF). This proinflammatory factor is a major cytokine in the pathogenesis of RA, and the TNF genes are located in the MHC locus on chromosome 6 in humans. Several polymorphisms of the TNF promoter including two at positions −238 and −308 can alter gene transcription. Associations among the TNF polymorphisms and RA susceptibility and radiographic progression have been reported, although there is not uniform agreement. In addition, certain polymorphisms in cytokines, especially TNF or Fc receptors, have been associated with differential response to therapy. For instance, substitution of a T for a C at position −857 in the TNF promoter might confer greater responsiveness to TNF inhibitors.6
Among the many noncytokine and non-MHC genetic linkages described for RA, the ones associated with peptidyl arginase deiminase (PADI) and PTPN22 have the strongest effect on susceptibility. The PADI genes are responsible for the post-translational modification of arginine to citrulline. Four isoforms have been identified, known as PADI1 through PADI4. In light of the striking associations of RA with ACPAs, several groups have investigated potential associations with these genes. The most promising is an extended haplotype in the PADI4 gene that can lead to increased levels of PADI4 protein due to enhanced messenger RNA (mRNA) stability.7 In a Japanese cohort, a twofold increase in risk of RA was observed with PADI4 SNPs. Confirmatory reports have been mixed because the association has been confirmed in other Asian populations but not in Western Europe. These studies suggest that the contribution of PADI4 to RA might be restricted, depending on the overall genetic background of the patient population.
Protein tyrosine phosphatase-22 (PTPN22) associations have been discovered in large-scale screening efforts to identify SNP associations in RA.8 Using 12,000 SNPs in the initial screens, a novel association was discovered at position 1858 in the PTPN22 gene that, like PADI4, conferred a twofold increase in risk. The allele containing thymidine leading to an amino acid substitution (R620W) was present in 8.5% of controls but was found in nearly 15% in patients with seropositive RA. Subsequent studies have demonstrated a similar association with systemic lupus erythematosus (SLE), type 1 diabetes, and several other autoimmune diseases. PTPN22 is a phosphatase that regulates the phosphorylation status of several kinases important to T cell activation including Lck and ZAP70. The R620W allele surprisingly results in a gain of function that alters the threshold for T cell receptor (TCR) signaling. Because the PTPN22 allele is rare in Japan, it is another gene (e.g., PADI4) where susceptibility is specific for particular ethnic or racial populations.
The list of genes associated with RA consistently involves immune regulation.9 Cytokine polymorphisms such as for TNF and the IL-1 inhibitor, IL-1Ra are not surprising. Genes that regulate adaptive immune responses in T cells such as PTPN22 and the co-stimulation receptor CTLA have also been associated with RA. Other genes associated with B cell function and/or antigen presentation such as BTLA (B- and T-lymphocyte attenuator), Fc receptors, and CD40 are also implicated. Polymorphisms have also been identified in signal transduction pathways that regulate immune function such as TRAF1-C5 and STAT4. The consistent thread in this analysis is that most gene associations for RA cluster to innate immunity, adaptive immunity, and inflammation. Aside from providing insight into the mechanisms of disease, they could also potentially contribute to responses to targeted therapies.
A number of environmental factors clearly contribute to RA susceptibility, although no specific exposure has been identified as the pivotal agent. Smoking is the best defined environmental risk factor for seropositive RA. The reason for its influence on the development of synovitis is not fully defined but could involve the activation of innate immunity and PADI in the airway. Citrullinated proteins have been detected in bronchoalveolar lavage samples of smokers, and this could provide a stimulus for generation of ACPAs in susceptible individuals.10 Repeated activation of innate immunity, especially in an individual with underlying genetically determined autoreactivity, could potentially contribute to autoreactivity and the initiation of synovitis. Other environmental factors such as oral contraceptives appear to modestly protect from RA, perhaps due to changes in the hormone milieu.11
The interaction between HLA-DR and tobacco exposure is perhaps the best example of how genes and the environment conspire to enhance risk. Although smoking and the SE alone modestly increase the likelihood of developing RA, the combination is synergistic.12 An individual with a history of cigarette smoking and two copies of the SE increases the odds of developing RA by up to 40-fold. The mechanism of the interaction is not known, but it could potentially relate to the increase in protein citrullination in smokers and increased ability of SE-containing HLA-DR molecules to bind some citrullinated proteins. The extent of smoking is also predictive, with the greatest risk seen with at least 20 pack-years. The risk declines slowly with cessation of smoking, taking more than a decade to begin approaching nonsmokers.13 Alcohol consumption can decrease this risk, and exposure to other inhaled irritants like silica dust increases risk, demonstrating the complexity of environment and human behavior on understanding disease susceptibility.
RA is one of many chronic autoimmune diseases that predominates in women. The ratio of female-to-male patients is 2 : 1 to 3 : 1, which in not as high as Hashimoto’s thyroiditis (25 : 1 to 50 : 1) or SLE (9 : 1). The gender effect is often observed in some animal models of autoimmunity such as the NZB/NZW model of SLE, in which female mice have more severe disease. Estrogens are one obvious explanation, and some data support the concept that these hormones modulate immune function.14 For example, autoantibody-producing B cells exposed to estradiol are more resistant to apoptosis, suggesting that autoreactive B cell clones might escape tolerance. The effect on T lymphocytes is harder to reconcile with the female preponderance in RA because estrogens tend to bias T cell differentiation toward the Th2 phenotype. The cytokines produced by this subset such as IL-4 and IL-13 are usually considered anti-inflammatory in animal models of arthritis and are present in only limited amounts in the RA synovium. Estrogen receptors are expressed on fibroblast-like synoviocytes (FLS) and increase production of metalloproteinases. In macrophage cell lines, estrogen can enhance production of TNF. Nulliparity has also been suggested as a risk factor in early studies, but more recent reports do not support this notion. Thus the effects of estrogens are complex, and the specific mechanisms responsible for the female preponderance of RA are not fully understood.
Pregnancy is often associated with remission of the disease in the last trimester. More than three quarters of pregnant patients with RA improve in the first or second trimester, but 90% of these experience a flare of disease associated with a rise in RF titers in the weeks or months after delivery. The mechanism of protection is not defined but might be due to the expression of suppressive cytokines such as IL-10 during pregnancy, production of α-fetoprotein, or alterations in cell-mediated immunity. One intriguing finding is that fetal DNA levels in the maternal peripheral blood correlate with the propensity for improved symptoms in pregnant RA patients. It is not certain whether the DNA itself contributes or whether it is a marker for increased leakage of fetal cells into the maternal circulation.15 Immune responses directed against paternal HLA antigens can occur and lead to the production of alloantibodies in the maternal circulation. Maternal-fetal disparity in human leukocyte antigen (HLA) class II phenotypes can correlate with pregnancy-induced remission. More than three-fourths of pregnant women with maternal-fetal disparity of HLA-DRB1, DQA, and DQB haplotypes have significant improvement, whereas disparity is only observed in one-fourth of women whose pregnancy is characterized by continuous active arthritis.16 Therefore suppression of maternal immune responses to paternal HLA haplotypes might be protective. This question remains unsettled because another study failed to find a correlation between the HLA disparity and clinical improvement during pregnancy.17
Epigenetics describes phenotypic or gene expression properties caused by mechanisms other than changes in the underlying DNA sequence. Modification of CpG DNA sequences by methylation, for example, can suppress gene expression, and it plays a role in cell differentiation. Histone acetylation also alters accessibility of DNA for transcription factors and RNA polymerases. microRNAs can bind to DNA and suppress expression of key genes involved with the inflammatory process. Some epigenetic information such as DNA methylation can be transferred from one generation to the next, thereby providing an alternative mechanism for rapidly altering disease susceptibility in a population due to the environment.
Most information on epigenetics in RA comes from studies of RA synovium or cultured synoviocytes. Some evidence of imprinting is available in the latter, with evidence of global DNA hypomethylation.18 Only low levels of a key DNA methylase, Dnmt1 are expressed in RA synovium, suggesting a molecular basis for this observation. Because this enzyme is carried by gametes, the methylation pattern can be transgenerationally maintained. One particular CpG site in the IL-6 promoter of peripheral blood mononuclear cells has decreased methylation, and this is associated with higher levels of IL-6 production.19 Dietary influences such as ingestion of methyl donors such as folate, or even exposure to methyl donors in utero, can profoundly alter DNA methylation and adaptive immune functions and affect susceptibility to autoimmune disease.
The histone deacetylase HDAC1 is overexpressed in RA FLS. When this gene is suppressed, synoviocyte proliferation decreases and expression of tumor suppressor proteins such as p53 increases. HDAC inhibitors are also effective in collagen-induced arthritis, markedly delaying the onset of disease and decreasing bone erosions. Finally, analysis of synovioyctes shows that some individual microRNAs such as microRNA-124a are decreased in RA compared with osteoarthritis (OA) cells. This particular microRNA can suppress cell cycling and chemokine genes. Increasing microRNA-124a levels in RA synoviocytes decreased the production of the chemokine MCP-1.20 Forced expression of microRNA-203 increases metalloproteinase and IL-6 expression by synoviocytes as well.21
The role of epigenetics in RA is not understood. It is not clear whether these observations occur before the onset of disease, are involved with the transition from asymptomatic autoimmunity to clinical disease, or participate in the destructive phase in established RA. Environmental stress plays a major role in disease susceptibility, perhaps greater than DNA polymorphisms. The mechanisms probably involve epigenetic deregulation of gene expression leading to decreased thresholds for autoreactivity in the adaptive immune system.
The history of RA reveals the surprising observation that it is a relatively new disease in Europe and Northern Africa. Examination of ancient skeletal remains in Europe and Northern Africa fails to reveal convincing evidence of RA, even though other rheumatic diseases such as OA, ankylosing spondylitis, and gout are readily discernable. In contrast, typical marginal erosions and rheumatoid lesions are present in the skeletons of Native Americans found in Tennessee, Alabama, and Central America from thousands of years ago. The first clear descriptions of RA in Europe appeared in the seventeenth century, and the disease was distinguished from gout and rheumatic fever by Garrod in the mid-nineteenth century. Although still controversial, the disease might have migrated from the New World to the Old World coincident with opening the trade and exploration routes. Because genetic admixture was relatively limited, an undefined environmental exposure potentially caused RA in susceptible Europeans. The most obvious explanation would, of course, be that an infectious agent is responsible. However, other environmental influences like tobacco smoking were also introduced to the Old World at the same time and could play a role.
Equally intriguing, the severity and incidence of RA appeared to decrease in the late twentieth century (Figure 69-2).22 In certain well-defined populations including Native Americans, the incidence of RA has gradually declined by as much as 50% over the past half of the twentieth century. Changes in hygiene and other lifestyle modifications related to industrialization might contribute, and an infectious agent might be less prevalent secondary to these societal changes, as with many other infectious diseases. Recent data from 1995 to 2007 suggest that the incidence might be rising again in women, but not in men. Dissecting the environmental exposures will be key to understanding how the susceptibility to disease varies over time.23
(From Doran MF, Pond GR, Crowson CS, et al: Trends in incidence and mortality in rheumatoid arthritis in Rochester, Minnesota over a forty-year period, Arthritis Rheum 46:625–631, 2002.)
Repeated inflammatory stress, especially through specialized receptors that recognize common molecules produced by pathogens, in a genetically susceptible individual might contribute to breakdown of tolerance and subsequent autoimmunity.
Considerable effort has been expended to assess the role of infectious agents in RA (Table 69-3). A potential pathogen could initiate disease through a variety of mechanisms including direct infection of the synovium, activation of innate immunity by pattern-recognition receptors that bind to components of the agent, or through molecular mimicry that induces an autoreactive adaptive immune response.
|Infectious Agent||Potential Pathogenic Mechanisms|
|Mycoplasma||Direct synovial infection; superantigens|
|Parvovirus B19||Direct synovial infection|
|Retroviruses||Direct synovial infection|
|Enteric bacteria||Molecular mimicry (QKRAA, e.g., in bacterial heat shock proteins)|
|Mycobacterium||Molecular mimicry (proteoglycans, QKRAA), immunostimulatory DNA (Toll-like receptor 9 activation)|
|Epstein-Barr virus||Molecular mimicry (QKRAA in gp110)|
|Bacterial cell walls||Toll-like receptor 2 activation|
Infectious agents could contribute to the initiation or perpetuation of RA through a variety of mechanisms. Some arthrotropic microorganisms could potentially infect the synovium and cause a local inflammatory response. There is increasing awareness that the innate immune system could also directly affect the onset and course of synovitis. Pathogen-associated molecular pattern receptors, especially the Toll-like receptors (TLRs), are expressed by sentinel cells in the host that provide a first line of defense. These receptors recognize preserved structures in bacteria and other infectious agents and permit rapid release of inflammatory mediators, activation of antigen-presenting cells, and enhancement of adaptive immune responses.
At least 11 TLRs exist in humans such as TLR2 (binds peptidoglycans), TLR3 (binds double-stranded RNA [dsRNA]), TLR4 (binds lipopolysaccharide), and TLR9 (binds bacterial DNA containing CpG motifs). Many of these pattern recognition receptors are expressed by rheumatoid synovial tissue and cultured FLS including TLR2, TLR3, TLR4, and TLR9. Exogenous TLR ligands such as bacterial peptidoglycan and DNA, as well as endogenous ligands (e.g., heat shock proteins, fibrinogen, and hyaluronan), are present in arthritic joints (see later). Engagement of these receptors participates in certain animal models of arthritis and can exacerbate synovial inflammation. TLR3, which recognizes viral dsRNA and activates the antiviral response, is also expressed by synovial cells in the intimal lining. Necrotic debris containing mRNA from RA synovial fluid cells activates TLR3 signaling and proinflammatory gene expression in synovitis.
The role of innate immunity in RA led to the notion that repeated engagement of TLRs in the synovium could help initiate disease. This hypothesis could explain why specific pathogens have been difficult to identify in the joint. In contrast, a genetically susceptible individual could potentially break tolerance if the TLRs are repeatedly engaged and permit autoimmune responses against articular antigens. Several animal models of disease require TLR ligands for initiations such as TLR9 in adjuvant arthritis. TLR2 is required for streptococcal cell wall arthritis, and the chronic T cell–dependent phase of that model requires TLR4. Mice lacking TLR4−/− have significantly less joint damage induced by IL-1 overexpression, even though synovial inflammation is still robust.24 These data suggest that endogenous TLR ligands play a key role in matrix regulation independent of inflammatory responses.
A second mechanism that regulates innate immunity involves a novel structure called the inflammasome. This complex includes several proteins involved in recognition of “danger signals” and pathogens such as muramyl dipeptides and uric acid. One central component is cryopyrin, also called NALP3, which is linked to caspase 1 (IL-1 convertase) by adapter proteins. When the inflammasome is engaged, caspase 1 is activated and IL-1 is produced. Mutations in this pathway, especially in cryopyrin, have been associated with autoinflammatory disorders such as Muckle-Wells syndrome and familial cold autoinflammatory disease. Inflammation induced by uric acid crystals or ATP uses this pathway and can be abrogated by IL-1 inhibitors. Cryopyrin is abundant in RA synovium and is constitutively expressed by FLS and macrophages. Expression in cultured FLS is markedly increased by TNF. Although the role of the inflammasome in RA has not been fully defined, its ability to induce cytokine production by exposure to bacterial products and other danger signals suggests that it participates in IL-1 and IL-18 regulation.
Active infection of synovial tissue by pyogenic bacteria is an unlikely cause of RA, and extensive searches for a unique or specific organism in synovial tissue or joint effusions have been negative. Antibodies to certain organisms such as Proteus are reportedly elevated in the blood of patients with RA, but this could represent an epiphenomenon or a nonspecific B cell activation. Most RA and reactive arthritis patients contain bacterial DNA sequences in their synovium. The bacteria identified are not unique and generally represent a cross-section of skin and mucosal bacteria including Acinetobacter and Bacillus spp. It is possible that the synovium functions as an adjunct to the reticuloendothelial system in arthritis, allowing local macrophages to accumulate circulating bacterial products.
In addition to prokaryotic DNA, bacterial peptidoglycans have been detected in RA synovial tissue (Figure 69-3). Antigen-presenting cells containing these products express TLRs and produce proinflammatory cytokines such as TNF. It is not known whether the peptidoglycans activate cells in situ or whether phagocytic cells from other sites or the blood engage the molecules and then migrate to the joint. In either case, it is not difficult to imagine how they can contribute to synovial inflammation.
Figure 69-3 Accumulation of bacterial peptidoglycan in rheumatoid synovium. A and B, Immunohistochemistry shows synovial cells containing peptidoglycan (red). C, Double staining studies show that bacterial peptidoglycan accumulates in synovial macrophages (arrow). These bacterial products can activate Toll-like receptors and stimulate cytokine production.
(From Schrijver IA, Melief MJ, Tak PP, et al: Antigen-presenting cells containing bacterial peptidoglycan in synovial tissues of rheumatoid arthritis patients coexpress costimulatory molecules and cytokines, Arthritis Rheum 43:2160, 2000.)
Several animal models of arthritis are dependent on TLR2, TLR3, TLR4, or TLR9. For instance, rodents injected with streptococcal cell walls (TLR2 ligand) develop severe polyarticular arthritis. The initial phase of disease resolves and is then followed by a chronic T cell–dependent phase that resembles RA. The arthritogenicity of complete Freund’s adjuvant in the rat adjuvant arthritis model is dependent on mycobacterial DNA that binds to TLR9 and activates an adaptive immunity. Endogenous TLR4 ligands such as heat shock proteins and fibrinogen also play a role in immune complex models such as passive K/BxN arthritis.25
Mycoplasma-derived superantigens such as from Mycoplasma arthritidis can directly induce T cell–independent cytokine production by macrophages and can exacerbate or trigger arthritis in mice immunized with type II collagen. There is also a higher prevalence of antimycoplasma pneumoniae IgG antibodies in RA patients than matched controls. Despite this and other circumstantial evidence, most efforts to identify Mycoplasma and Chlamydia organisms or DNA in joint samples have been negative, and there is no direct evidence to support these organisms as etiologic agents.
Epstein-Barr virus (EBV) is a polyclonal B lymphocyte activator that increases the production of RF, and rheumatoid macrophages and T cells have defective suppression of EBV-induced proliferation of human B cells. Rheumatoid patients have higher levels of EBV shedding in throat washings, an increased number of virus-infected B cells in the circulating blood, higher levels of antibodies to normal and citrullinated EBV antigens, and abnormal EBV-specific cytotoxic T cell responsiveness compared with controls. Defective elimination of EBV-transformed lymphocytes in RA has fueled speculation that a specific immune defect contributes to initiation of disease.
Additional intriguing data implicating EBV in RA are derived from sequence homology between the susceptibility cassette in HLA-DR proteins and the EBV glycoprotein gp110. Like DRB*0401, gp110 contains the QKRAA motif and patients with serologic evidence of a previous EBV infection have antibodies against this epitope. Hence T cell recognition of EBV epitopes in some patients with the SE might cause an immune response directed at innocent bystander cells through “molecular mimicry.” This hypothesis could potentially account for disease perpetuation in the absence of active infection in patients with a specific MHC genotype. However, the data are circumstantial and gp110 is only one of many xenoproteins such as the Escherichia coli heat shock protein dnaJ that contain QKRAA. RA T cells, especially synovial fluid T cells, have increased proliferative responses to gp110, perhaps supporting the molecular-mimicry link between a variety of QKRAA-containing proteins and arthritis.
Antecedent infection with parvovirus B19 has been implicated in some patients with RA based on serologic evidence including the nonstructural protein NS1. However, only about 5% of patients have evidence of recently acquired parvovirus B19 infection at the time of disease onset. Of interest, 75% of RA synovium samples contain B19 DNA compared with about 20% of non-RA controls. Immunohistochemical evidence of the B19 protein VP-1 was detected in patients with RA but not other forms of arthritis. However, evidence of the B19 genome in RA joint samples was not found in other studies.
The mechanisms of B19-induced synovitis, when it does occur, could be related to alterations in the function of FLS.26 In a cell-culture model of synoviocyte invasion into cartilage, infection with the parvovirus significantly increased the migration of cells into the matrix. Mice that are transgenic for the B19 protein NS1 were more susceptible to collagen-induced arthritis and developed high titers of anti–type II collagen antibodies. These data suggest that the B19 genome might not cause arthritis but can enhance an arthritogenic response to other environmental stimuli.
Because rubella virus and the rubella vaccine can cause arthritis in humans, the virus has attracted some attention as a possible triggering agent. Live rubella virus can be isolated from synovial fluid of some patients with chronic inflammatory oligoarthritis or polyarthritis without clinical evidence of rubella. However, most rubella patients do not have the classic polyarticular involvement and display an oligoarthritis involving large joints. As with B19 infection, it is possible that a small subset of patients with chronic polyarthritis that are called RA actually have direct infection with wild-type or attenuated rubella virus.
Studies of synovial tissue in a variety of inflammatory and noninflammatory arthropathies have also demonstrated DNA of other viruses such as cytomegalovirus and herpes simplex, but not adenovirus or varicella-zoster. As with bacterial DNA, parvovirus, and EBV, the localization of viral DNA to the inflamed joint might be related to the migration of inflammatory cells containing the viral genome or other nonspecific mechanisms rather than an active infection. Although the hypothesis that one or more of these viral infections might serve as a triggering agent in the genetically susceptible host is both appealing and intellectually satisfying, the pathogenic role of these agents is unlikely.
Retroviral infections have been suggested as a cause of RA. Extensive searches for potential agents have not been fruitful. Endogenous retroviruses are abundant in inflamed and normal synovium, and certain transcripts are expressed in RA cells. In one study, higher levels of HERV-K10 gag protein from a common endogenous retrovirus were detected more often in RA compared with OA and normal peripheral blood mononuclear cells. Some indirect studies are suggestive of retroviral infection such as the demonstration of zinc-finger transcription factors in cultured synoviocytes that can increase signaling through enzymes like p38 mitogen-activated protein (MAP) kinase. In addition, the pX domain of one human retrovirus, human T lymphotropic virus-1 (HTLV-1), causes synovitis in transgenic mice, and synoviocytes from patients infected with HTLV-1 have increased cytokine production. Other studies failed to demonstrate increased expression of human retrovirus-5 proviral DNA in rheumatoid synovium. There is still no direct evidence that retroviruses cause RA, but some viral products could activate TLR3 or TLR7 to enhance cytokine and chemokine production.
The idea that aberrant immune responses are directed toward self-antigens in RA was recognized with the discovery of RF in the blood of patients with the disease. Initially described by Waaler and later by Rose, it was not until the mid-1950s that Kunkel and colleagues firmly established that RF is an autoantibody. Although our understanding of autoantigens has changed over the years and the relative contributions of cellular and humoral immunity have been debated, emphasis on the role of autoantibodies in RA has enjoyed a resurgence over the past few years. Clinical improvement can be associated with decreases in levels of RFs or ACPAs, although the changes tend to be modest and are inconsistent.