OBJECTIVES

After completing this chapter, the reader should be able to

  • Describe the physiologic basis for rheumatic laboratory tests and the pathophysiologic processes that result in abnormal test results

  • Describe the appropriate clinical applications for laboratory tests used to diagnose or assess the activity of select rheumatic diseases

  • Interpret the results of laboratory tests used to diagnose or manage common rheumatic diseases

  • Use the results of rheumatic laboratory tests to make decisions about the effectiveness of pharmacotherapy

  • Employ laboratory tests to identify and prevent adverse reactions to drugs used to treat rheumatic diseases

The diagnosis and management of most rheumatic diseases depend primarily on patient medical history, symptoms, and physical examination findings. A variety of laboratory tests are used to assist in the diagnosis of rheumatic disorders, but many are nonspecific tests that are not pathognomonic for any single disease. However, the results of some specific laboratory tests may be essential for confirming the diagnosis of some diseases. Consequently, laboratory tests are important diagnostic tools when used in concert with the medical history and other subjective and objective findings. Some laboratory test results are also used to assess disease severity and to monitor the beneficial and adverse effects of pharmacotherapy.

The diagnostic utility of a laboratory test depends on its sensitivity, specificity, and predictive value (Chapter 1). Tests that are highly sensitive and specific for certain rheumatic diseases often have low predictive values because the prevalence of the suspected rheumatic disease is low. The most important determinant of a laboratory test’s diagnostic usefulness is the pretest probability of disease, or a clinician’s estimated likelihood that a certain disease is present based on history and clinical findings. As the number of disease-specific signs and symptoms increases and approaches diagnostic confirmation, the pretest probability also increases.

After briefly reviewing pertinent physiology of immunoglobulins, this chapter discusses various tests used to diagnose and assess rheumatic diseases, followed by interpretation of these test results in common rheumatic disorders. Tests used to monitor antirheumatic pharmacotherapy are also described.

STRUCTURE AND PHYSIOLOGY OF IMMUNOGLOBULINS

Many rheumatic laboratory tests involve detection of immunoglobulins (antibodies) that are directed against normal cellular components. The structure and functions of immunoglobulins are reviewed briefly here to facilitate an understanding of these tests.

When the immune system is challenged by a foreign substance (antigen), activated B lymphocytes differentiate into immunoglobulin-producing plasma cells. Immunoglobulins are Y-shaped proteins with an identical antigen-binding site (called Fab or fraction antigen-binding) on each arm of the Y (Figure 20-1). Each arm is composed of a light (L) amino acid chain covalently linked to a heavy (H) amino acid chain. The terms light and heavy refer to the number of amino acids in each chain. Because the heavy chain has more amino acids than the light chain, it is longer and has a higher molecular weight.

FIGURE 20-1.
FIGURE 20-1.

Schematic of the antibody molecule. (Source: Adapted with permission from Four Chain Structure of a Generic Antibody-IgG2 Structures by OpenStax College—Anatomy & Physiology, Connexions Website. http://cnx.org/content/col11496/1.6/, Jun 19, 2013. Licensed under CC BY 3.0 via Wikmedia Commons– https://commons.wikimedia.org/wiki/File:2220_Four_Chain_Structure_of_a_Generic_Antibody-IgG2_Structures.jpg.)

Both types of chains have a variable region and a constant region. The variable regions contain the antigen-binding sites and vary in amino acid sequence. The sequences differ to allow immunoglobulins to recognize and bind specifically to thousands of different antigens. Within the variable regions, there are four framework regions and three complementary-determining regions; together these make up the antigen-binding pocket. The constant region of the light chain is a single section. Immunoglobulins that have identical constant regions in their heavy chains are of the same class.

The five classes of immunoglobulins are IgA, IgD, IgE, IgG, and IgM. Depending on the immunoglobulin, the constant region of the heavy chain has either three domains and a hinge region (IgA, IgD, and IgG) that promotes flexibility, or four domains without a hinge region (IgE and IgM). Thus, the immunoglobulin’s heavy chain determines its class (α heavy chains, IgA; δ heavy chains, IgD; ε heavy chains, IgE; γ heavy chains, IgG; and μ heavy chains, IgM). Tests are available to measure the serum concentrations of the general types of immunoglobulins as well as immunoglobulins directed against specific antigens (viruses, other infectious agents, other allergens).

In Figure 20-1, the second and third domains of the heavy chain are part of the fraction crystallizable (Fn) portion of the immunoglobulin. This portion has two important functions: (1) activation of the complement cascade (discussed later) and (2) binding of immunoglobulins (which react with and bind antigen) to cell surface receptors of effector cells such as monocytes, macrophages, neutrophils, and natural killer cells.1

TESTS TO DIAGNOSE AND ASSESS RHEUMATIC DISEASES

Blood tests that are relatively specific for certain rheumatic diseases include rheumatoid factors (RFs), anticitrullinated protein antibodies (ACPAs), antinuclear antibodies (ANAs), antineutrophil cytoplasmic antibodies (ANCAs), and complement. Nonspecific blood and other types of tests include erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), analysis of synovial fluid, and others. Where applicable, the sections that follow discuss quantitative assay results (where normal values are reported as a range of concentrations), qualitative assay results (where assay results are reported as only positive or negative), and their use in common rheumatic and nonrheumatic diseases.

Rheumatoid Factor

Rheumatoid factors (RFs) are immunoglobulins (predominantly IgM but may also be IgG or IgA) that are abnormally directed against the Fc portion of IgG. These immunoglobulins do not recognize the IgG as being “self.” Therefore, the presence of RFs in the blood indicates an autoimmune process. The RF measured in most laboratories is IgM-anti-IgG (an IgM antibody that specifically binds IgG). Like all IgM antibodies, IgM RF is composed of five subunits whose Fc portions are attached to the same base. The variable regions of each IgM antibody can bind up to five IgG molecules at its multiple binding sites, making IgM RF the most stable and easiest to quantify.

Rheumatoid factors are most commonly associated with rheumatoid arthritis (RA) but are not specific for that disease. Other rheumatic diseases in which circulating RFs have been identified include systemic lupus erythematosus (SLE), systemic sclerosis (scleroderma), mixed connective tissue disease (MCTD), and Sjögren syndrome.3 The significance of RFs in these diseases is unknown.

The presence of RF is not conclusive evidence that a rheumatic disease exists. Patients with various acute and chronic inflammatory diseases as well as healthy individuals may be RF positive. Nonrheumatic diseases associated with RFs include mononucleosis, hepatitis, malaria, tuberculosis, syphilis, subacute bacterial endocarditis, cancers after chemotherapy or irradiation, chronic liver disease, hyperglobulinemia, and cryoglobulinemia.

The percentage of individuals with positive RF concentrations and the mean RF concentration of the population increase with advancing age but is 1% to 2% of the heathy population on average. Although RFs are associated with several rheumatic and many nonrheumatic diseases, the concentrations of RFs in these diseases are lower than those observed in patients with RA.

Quantitative Assay Results

Normal values: <1:16 or <15 International Units/mL

When a quantitative RF test is performed, results are reported as either a dilutional titer or a concentration in International Units per milliliter. RF titers are reported positive as a specific serum dilution; the ability to detect RF is tested at each dilution. The greatest dilution that results in a positive test is reported as the endpoint. A titer of >1:16 or a concentration >15 International Units/mL is generally considered to be positive. However, reference ranges vary depending on the method used, so it is important to use the limits provided by the individual laboratory.

Qualitative Assay Results

At a serum dilution at which 95% of the normal population is RF negative, 70% to 90% of RA patients will have a positive RF test. The remaining RA patients who have RF titers within the normal range may be described as seronegative.

Anticitrullinated Protein Antibodies

Citrullination is the process by which susceptible genes exposed to environmental factors undergo an abnormal change. As a result, citrullinated proteins become antigens and lead to the formation of autoantibodies. ACPAs bind to the nonstandard amino acid citrulline that is formed from removal of amino groups from arginine. Nonstandard amino acids are generally not found in proteins and often occur as intermediates in the metabolic pathways of standard amino acids. In the joints of patients with RA, proteins may be transformed to citrulline during the pathogenesis that leads to joint inflammation. When these antibodies are present, there is a 90% to 95% likelihood that the patient has RA. The combination of both positive RF and positive ACPA has 99.5% specificity for RA. Positive ACPAs may occur in other diseases, including SLE, systemic sclerosis, psoriasis, and tuberculosis.

Anticitrullinated protein antibodies are important considerations in the context of RA for multiple reasons: (1) presence is associated with more erosive forms of RA, (2) presence is considered one of the clinical features of RA that is associated with a worse long-term prognosis, and (3) presence is a positive predictor of future RA diagnosis in patients with undifferentiated arthritis. Therefore, ACPA-positive patients should receive aggressive treatment early and be followed closely throughout the course of their disease to control systemic inflammation.

Quantitative and Qualitative Assay Results

Normal values: <20 EU/mL (assay dependent)

Quantitative ACPAs are tested by enzyme-linked immunosorbent assay (ELISA) and are reported in ELISA units (EU). The relationship between these values and qualitative results are generally reported in the following way:

  • <20 EU: negative

  • 20 to 39 EU: weakly positive

  • 40 to 59 EU: moderately positive

  • >60 EU: strongly positive

Antinuclear Antibodies

Antinuclear antibodies (ANAs) are a heterogeneous group of autoantibodies directed against nucleic acids and nucleoproteins within the nucleus and cytoplasm. Intracellular targets of these autoantibodies include DNA, RNA, individual nuclear histones, acidic nuclear proteins, and complexes of these molecular elements (Table 20-1).4-7

TABLE 20-1.

Laboratory and Clinical Characteristics of Antibodies to Nuclear/Cytoplasmic Antigens

ANA

TARGETED CELLULAR MATERIAL

SENSITIVITY

SPECIFICITY

dsDNA

dsDNA

SLE: 70%

RA: 1%

Systemic sclerosis: <1%

SLE: high (>95%)

Drug-induced lupus: low (1% to 5%)

RA: low (1%)

Systemic sclerosis: low (<1%)

Sjögren syndrome: low (1% to 5%)

ssDNA

ssDNA

SLE: 80%

Drug-induced lupus: 80%

RA: 60%

SLE: low

Drug-induced lupus: low

RA: moderate

Sma

Nuclear ribonucleoproteins

SLE: 10% to 50%

Drug-induced lupus: 1%

RA: 1%

Systemic sclerosis: <1%

Sjögren syndrome: 1% to 5%

SLE: high (55% to 100%)

RNP

(or U1RNP)

Nuclear ribonucleoproteins

MCTD: 100%

SLE: 3% to 69%

Systemic sclerosis: <1%

RA: 25%

Polymyositis: <1%

MCTD: low

SLE: low

Systemic sclerosis: low

RA: low

Polymyositis: low

Histone

Chromatin and DNA-packing protein

SLE: 70%

Drug-induced lupus: 95%

RA: 15% to 20%

Systemic sclerosis: <1%

SLE: moderate

Drug-induced lupus: high

RA: low

Systemic sclerosis: low

Roa/SSAb

Nuclear ribonucleoproteins

Sjögren syndrome: 10% to 60%

SLE: 40%

Polymyositis: 18%

RA: 5%

Systemic sclerosis: 5%

Sjögren syndrome: moderate

SLE: low

Polymyositis: low

RA: low

Systemic sclerosis: low

Laa/SSBb

Nuclear ribonucleoproteins

Sjögren syndrome: 70% to 95%

SLE: 10% to 35%

Sjögren syndrome: high (94%)

SLE: low

Centromere

(ACA)

Chromatin and centromere

Systemic sclerosis: 25% to 30%

CREST: 50% to 90%

Raynaud phenomenon: 15% to 30%

Systemic sclerosis: high

CREST, Raynaud phenomenon: high

(>95%)

DNA topoisomerase I (Scl70)

Chromatin and DNA-catalyzing protein

Systemic sclerosis: 15% to 20%

Systemic sclerosis: high (>95%)

Jo-1

Cytoplasm and histidyl tRNA synthetase

SLE: low

Drug-induced lupus: low

RA: low

Systemic sclerosis: low

Sjögren syndrome: low

Polymyositis: 30%

Jo-1 syndrome: 50%

SLE: low

Drug-induced lupus: low

RA: low

Systemic sclerosis: low

Sjögren syndrome: low

Polymyositis with interstitial

lung disease: high

Jo-1 syndrome: high

ACA = anticentromere antibody; CREST = syndrome characterized by calcinosis, Raynaud phenomenon, esophageal motility disorder, sclerodactyly, and telangiectasias; La/SSB = La/Sjögren syndrome B antibody; RNP = ribonucleoprotein; Ro/SSA = Ro/Sjögren syndrome A antibody; Scl70 = scleroderma-70 or DNA topoisomerase I antibody; U1RNP = uridine-rich ribonuclear protein.

aRepresents the first two letters of the surname of the patient whose serum was used to identify the reaction in agar diffusion.

bSjögren syndrome A and B.

Source: From references 47.

The ANA test is included in the diagnostic criteria for idiopathic SLE, drug-induced lupus, and MCTD because of its high rate of positivity in these disorders. However, its low specificity makes it unsuitable for use as a screening test for rheumatic or nonrheumatic diseases in asymptomatic individuals. A positive ANA can also be found in otherwise healthy individuals. ANAs are also associated with various genetic and environmental factors (eg, intravenous drug abuse), hormonal factors, and increased age. They also are associated with nonrheumatic diseases, both immunologically mediated (eg, Hashimoto thyroiditis, idiopathic pulmonary fibrosis, primary pulmonary hypertension, idiopathic thrombocytopenic purpura, and hemolytic anemia) and nonimmunologically mediated (eg, acute or chronic bacterial, viral, or parasitic infections and neoplasm).

Antibody tests that have clinical use for diagnosis of SLE, drug-induced lupus, and other diseases are as follows.

Double-Stranded DNA (anti-dsDNA) Antibodies

Normal values: negative (ELISA), <1:10 titer (FANA)

These antibodies are relatively specific for SLE, which makes them useful for diagnosis. In some patients with SLE, the titers tend to rise with a disease flare and fall (usually into the normal range) when the flare subsides. Thus, anti–double-stranded DNA (dsDNA) titers may be helpful in managing disease activity in some patients with SLE. Anti-dsDNA antibodies have been found in low titers in many other autoimmune diseases (eg, RA, Sjögren syndrome, systemic sclerosis, Raynaud phenomenon, MCTD, discoid lupus, juvenile idiopathic arthritis [JIA], and autoimmune hepatitis).6 The presence of anti-dsDNA antibodies has also been reported in patients receiving some drugs used to treat rheumatic diseases (eg, minocycline, etanercept, infliximab, and penicillamine).

Single-Stranded DNA (Anti-ssDNA) Antibodies

These antibodies identify and react primarily with purine and pyrimidine bases within the β-helix of dsDNA. They may also bind with nucleosides and nucleotides. They are much less specific for SLE than anti-dsDNA antibodies. They are of limited clinical use.

Smith (Anti-Sm) Antibodies

Normal values: negative

These antibodies bind to a series of nuclear proteins complexed with small nuclear RNAs. These complexes are known as small nuclear ribonucleoprotein particles and are important in the processing of RNA transcribed from DNA.6 The anti-Sm antibody test has low sensitivity (10% to 50% depending on assay methodology) but high specificity (55% to 100%) for SLE. Titers usually remain positive after disease activity has subsided and titers of anti-DNA antibodies have declined to the normal range. Thus, the anti-Sm antibody titer may be a useful diagnostic tool, especially when anti-DNA antibodies are undetectable. There is currently no evidence that monitoring anti-Sm antibodies is useful for following the disease course or predicting disease activity.6

Ribonucleoprotein (Anti-RNP) or Uridine-Rich Ribonuclear Protein (Anti-U1RNP) Antibodies

Normal values: Negative

This antibody system reacts to antigens that are related to Sm antigens. However, these antibodies bind only to the U1 particle, which is involved in splicing nuclear RNA into messenger RNA. Ribonucleoprotein antibodies are found in many patients with SLE (3% to 69%), and low titers may be detected in other rheumatic diseases (eg, Raynaud phenomenon, RA, systemic sclerosis).6 Importantly, anti-U1RNP antibodies are a hallmark feature of MCTD. A positive test result in a patient with suspected MCTD increases the probability that this diagnosis is correct, even though the test is nonspecific. On the other hand, a negative anti-RNP result in a patient with possible MCTD virtually excludes this diagnosis.

Anti-Histone (Nucleosome) Antibodies

Normal values: Negative

These antibodies target the protein portions of nucleosomes, which are DNA–protein complexes comprising part of chromatin. These antibodies are present in virtually all cases of drug-induced lupus. In fact, the diagnosis of drug-induced lupus should be questioned in their absence. Most cases of drug-induced lupus are readily diagnosed because a commonly implicated drug (eg, hydralazine, isoniazid, procainamide) is being taken or a strong temporal relationship exists between drug initiation and the onset of SLE signs and symptoms. However, in some cases of potential drug-induced lupus, histone autoantibody testing can be helpful. Histone antibodies appear less commonly in other diseases, including RA, JIA, autoimmune hepatitis, scleroderma, and others. There is some evidence that histone antibodies correlate with disease activity in SLE.

Two closely related ANA tests are detected frequently in patients with Sjögren syndrome, but they are nonspecific; they may also be helpful for diagnosis of SLE.

  1. Ro/Sjögren Syndrome A (Anti-Ro/SSA) Antibody

    • Normal values: Negative

    • Autoantibodies directed against Ro/SSA recognize one of two cellular proteins, one in the nucleus (Ro60) and one in the cytoplasm (Ro52). The numbers represent the molecular weight of the protein in kilodalton.

  2. La/Sjögren Syndrome B (Anti-La/SSB) Antibody

    • Normal values: Negative

    • Autoantibodies directed against La/SSB targets a protein found in the nucleus that goes back and forth between the cytoplans and the nucleus.

The presence of either antibody in patients with suspected Sjögren syndrome strongly supports the diagnosis. It is unusual to detect the anti-La/SSB antibody in patients with SLE or Sjögren syndrome in the absence of the anti-Ro/SSA antibody. In women of childbearing age who have a known connective-tissue disease (eg, SLE, MCTD), a positive anti-Ro/SSA antibody is associated with an infrequent (1% to 2%) but definite risk of bearing a child with neonatal SLE and congenital heart block. Presence of the anti-Ro/SSA antibody also correlates with late-onset SLE and secondary Sjögren syndrome. In patients who are ANA negative but have clinical signs of SLE, a positive anti-Ro/SSA antibody may be useful in establishing a diagnosis of SLE.

It has been recommended that the anti-Ro/SSA antibody test be ordered in the following situations7: (1) women with SLE or Sjögren syndrome who are planning to become pregnant, (2) women with a history of giving birth to children with heart block or myocarditis, (3) women known to be ANA positive who wish to become pregnant, (4) patients suspected of having a systemic connective tissue disease with a negative ANA screening test, and (5) patients with xerostomia, keratoconjunctivitis sicca, and salivary and lacrimal gland enlargement.

Two ANAs are highly specific for systemic sclerosis (scleroderma), but the tests have low sensitivity:

  • Antikinetochore (centromere) antibody

  • Antitopoisomerase I (Scl70) antibody

In addition, these two antibodies are highly specific for CREST syndrome (associated with calcinosis, Raynaud phenomenon, esophageal dysmotility, sclerodactyly, and telangiectasias), Raynaud phenomenon, and occasionally SLE. When systemic sclerosis is suspected on clinical grounds, antibody testing for antikinetochore and Scl70 can be useful in making the diagnosis. However, negative results do not exclude the disease because of low test sensitivity.

The Jo-1 antibody (anti-Jo) is highly specific for idiopathic inflammatory myopathy, including polymyositis and dermatomyositis, or myositis associated with another rheumatic disease or interstitial lung disease. The titer or quantitative value should be considered when evaluating the clinical significance of ANA test results.

Quantitative Antinuclear Antibody Assay Results

Normal: Negative at 1:40 dilution (varies among laboratories)

The indirect immunofluorescence antinuclear antibody test (FANA) is a rapid and highly sensitive method for detecting the presence of ANAs.4 It is considered the gold standard for ANA testing. Although the FANA is positive in >95% of patients with SLE, it is also positive in some normal individuals and patients with drug-induced lupus and other autoimmune diseases. An ELISA also provides a rapid and highly sensitive method for detecting the presence of ANA; however, the sensitivity is lower than FANA. Many laboratories perform screening ANA tests by the ELISA technique because it can be automated and is less labor intensive; FANA testing is performed only in specimens testing positive by ELISA.5 It is important for the clinician to know the technique used at a specific laboratory to interpret the results correctly.

Laboratories usually report the ANA titer, which is the highest serum dilution that remains positive for ANAs. A high concentration (titer >1:640) should raise suspicion for an autoimmune disorder but is not in itself diagnostic of any disease. In the absence of clinical findings, these individuals should be monitored closely for the overt development of an autoimmune disorder. On the other hand, a high ANA titer is less useful in a patient who already has definite clinical evidence of a systemic autoimmune disease. The finding of a low antibody titer (<1:80) in the absence of signs or symptoms of disease is not of great concern, and such patients require less frequent follow-up than those with high titers. False-positive ANAs are common in the normal population and tend to be associated with low titers (<1:40). The positive antibody titers in healthy persons tend to remain fairly constant over time; this finding can also be seen in patients with known disease.

Qualitative Antinuclear Antibody Assay Results

The pattern of nuclear fluorescence after staining may reflect the presence of antibodies to one or more nuclear antigens. The nuclear staining pattern was used commonly in the past, but the pattern type is now recognized to have relatively low sensitivity and specificity for individual autoimmune diseases. For this reason, specific antibody tests have largely replaced use of patterns.5 The common immunofluorescent patterns are as follows:

  • Homogeneous: This pattern is seen most frequently in patients with SLE but can also be observed in patients with drug-induced lupus, RA, vasculitis, and polymyositis. This pattern reflects antibodies to the DNA-histone complex.

  • Speckled: This pattern is also seen most frequently in patients with SLE but can appear in patients with MCTD, Sjögren syndrome, progressive systemic sclerosis, polymyositis, and RA. This pattern is produced by antibodies to Sm, Ro/SSA, La/SSB, DNA topoisomerase I (Scl70), and other antigens.

  • Nucleolar: This pattern is infrequently observed in patients with SLE but is more frequently seen in patients with polymyositis, progressive systemic sclerosis, and vasculitis. It is produced by antibodies to RNA polymerase I and several other antigens.

  • Peripheral or nuclear rim: This is the only pattern that is highly specific for any rheumatic disease and is observed predominantly (98%) in SLE patients. It is produced by antibodies to DNA (dsDNA, ssDNA) and nuclear envelope antigens (antibodies to components of the nuclear envelope, such as certain glycoproteins).

Table 20-1 summarizes the most frequently identified ANAs, their corresponding targeted cellular material, and disease sensitivities and specificities.4-7

Antineutrophil Cytoplasmic Antibodies

As the name implies, antineutrophil cytoplasmic antibodies (ANCAs) are antibodies directed against neutrophil cytoplasmic antigens. Testing for ANCAs is important for the diagnosis and classification of various forms of vasculitis. In these disorders, the target antigens are proteinase 3 (PR3) and myeloperoxidase (MPO). Both antigens are located in the azurophilic granules of neutrophils and the peroxidase-positive lysosomes of monocytes. Antibodies that target PR3 and MPO are known as PR3-ANCA and MPO-ANCA.8 There is an association between ANCA and several major vasculitic syndromes: granulomatosis with polyangiitis (GPA), microscopic polyangiitis, eosinophilic granulomatosis with polyangiitis, and certain drug-induced vasculitis syndromes.8

GPA is a vasculitis of unknown origin that can damage organs by restricting blood flow and destroying normal tissue. Although any organ system may be involved, the disorder primarily affects the respiratory tract (sinuses, nose, trachea, and lungs) and the kidneys. Approximately 90% of patients with active GPA have ANCAs. In patients with limited disease presentations, typically limited to the respiratory tract, up to 40% may be ANCA-negative. Thus, although a positive ANCA test result is useful to support a suspected diagnosis, a negative ANCA test result does not exclude it. For this reason, the ANCA test is usually not used alone to diagnose GPA.

In patients with vasculitis, immunofluorescence after ethanol fixation reveals two characteristic patterns: cytoplasmic (cANCA) and perinuclear (pANCA). With cANCA, there is diffuse staining throughout the cytoplasm, which is usually caused by antibodies against PR3. The pANCA pattern is characterized by staining around the nucleus and perinuclear fluorescence. In patients with vasculitis, the antibody causing this pattern is generally directed against MPO.

Although detection and identification of ANCAs are most useful in diagnosing various vasculitides, ANCAs have been reported in connective tissue diseases (eg, RA, SLE, and myositis), chronic infections (eg, cystic fibrosis, endocarditis, and human immunodeficiency virus), and gastrointestinal diseases (eg, inflammatory bowel disease, sclerosing cholangitis, and autoimmune hepatitis). Some medications may induce vasculitis associated with positive ANCA (usually MPO-ANCA).9 The drugs most strongly associated with ANCA-associated vasculitis are propylthiouracil, methimazole, carbimazole, hydralazine, and minocycline. Penicillamine, allopurinol, procainamide, thiamazole, clozapine, phenytoin, rifampin, cefotaxime, isoniazid, and indomethacin are less commonly associated with the disorder.8

Quantitative Assay Results

Normal value: Negative

When used in the diagnosis of GPA, the specificity of PR3-ANCA is approximately 90%. The sensitivity of the test is about 90% when the disease is active and 40% when the disease is in remission. Thus, the sensitivity of PR3-ANCA is related to the extent, severity, and activity of the disease at the time of testing.

The use of obtaining serial PR3-ANCA tests in assessing disease activity is controversial. Some data suggest that a rise in titers predicts clinical exacerbations and justifies increasing immunosuppressive therapy. However, other studies have shown that disease flares cannot be predicted in a timely fashion by elevations in ANCA titers. Further, the immunosuppressive and cytotoxic therapies used are associated with substantial adverse effects. For these reasons, an elevation in ANCA should not be used as the sole justification for initiating immunosuppressive therapy. Rather, patients with rising ANCA titers should be monitored closely with therapy withheld unless there are clear clinical signs of active disease.8

Qualitative Assay Results

The association of cANCA and pANCA tests for various antigens and diseases are listed in Table 20-2. The presence of cANCAs denotes a spectrum of diseases ranging from idiopathic glomerulonephritis to extended GPA.10 In most cases of vasculitis, renal disorder, and granulomatous disease, patient sera are negative for cANCAs.

TABLE 20-2.
Rheumatic Disease Associations of Antinuclear Cytoplasmic Antibodies

ANCA

DISEASE

PERCENT ANCA POSITIVE (%)

cANCA

GPA

MPA

Pauci-immune crescentic glomerulonephritis

EGPA

70–90

20–30

10

10

pANCA

MPA

Pauci-immune crescentic glomerulonephritis

EGPA

GPA

65–90

80

30–40

25

EGPA = eosinophil granulomatosis with polyangiitis; MPA = microscopic polyangiitis.

Adapted from references 810.

The pANCA test has limited diagnostic value. A positive pANCA test result should be followed by antigen-specific assays such as anti-MPO. In ulcerative colitis, the specificity of the pANCA test has been reported to be as high as 94%. However, with only moderate sensitivity and inconsistent correlation between titers and disease activity, pANCA screening may be of little value. Although sensitivity can reach 85% in primary sclerosing cholangitis, the pANCA test lacks specificity in the differential diagnosis of autoimmune hepatic diseases. In RA, pANCA may be related to aggressive, erosive disease. The sensitivity of the test increases in RA complicated by vasculitis, but its specificity remains low.

Complement

The complement system consists of approximately 60 different plasma and membrane proteins that provide a defense mechanism against microbial invaders and serve as an adjunct or “complement” to humoral immunity. The system works by depositing complement components on pathologic targets and by the interaction of plasma proteins in a cascading sequence to mediate inflammatory effects such as opsonization of particles for phagocytosis, leukocyte activation, and assembly of the membrane attack complex (MAC).11 Six plasma control proteins and five integral membrane control proteins regulate this cascade. These proteins circulate normally in a precursor (inactive) form (eg, C3 and C4). When the initial protein of a given pathway is activated, it activates the next protein (eg, C3a and C4a) in a cascading fashion similar to that seen with coagulation factors.

Activation of this system can occur through any one of three proteolytic pathways:

  1. Classic pathway: This pathway is activated when IgM or IgG antibodies bind to antigens such as viruses or bacteria.

  2. Alternative pathway: This evolutionary surveillance system does not require the presence of specific antibodies.

  3. Lectin pathway: This pathway is activated similarly to the classic pathway, but instead of antibody binding, mannose-binding protein binds to sugar residues on the surface of pathogens.

Activation by any of the three pathways generates enzymes that cleave the third and fifth complement components (C3 and C5). A final common (or terminal) sequence culminates in the assembly of the MAC. Five proteins (C5 through C9) interact to form the MAC, which creates transmembrane channels or pores that displace lipid molecules and other elements, resulting in disruption of cell membranes and cell lysis.

Because the complement system is an important part of immune system regulation, complement deficiency predisposes an individual to infections and autoimmune syndromes. In disorders associated with autoantibodies and the formation of immune complexes, the complement system can contribute to tissue damage.

Serum complement levels reflect a balance between synthesis and catabolism. Hypocomplementemia occurs when the C3 or C4 concentration falls below its reference range. Most cases of hypocomplementemia are associated with hypercatabolism (complement depletion) due to activation of the immune system rather than decreased production of complement components (hyposynthesis). Most diseases associated with the formation of IgG- or IgM-containing circulating immune complexes can cause hypocomplementemia. Rheumatic diseases included in this category are SLE, RA with extraarticular disease, Sjögren syndrome, and systemic vasculitis. Nonrheumatic diseases associated with hypocomplementemia include antiphospholipid syndrome, subacute bacterial endocarditis, hepatitis B surface antigenemia, pneumococcal infection, gram-negative sepsis, viral infections (eg, measles), recurrent parasitic infections (eg, malaria), and mixed cryoglobulinemia.11,12

Because errors in interpretation of complement study results can occur, three important aspects should be considered when interpreting these results:

  1. Reference ranges are relatively wide. Therefore, new test results should be compared with previous test results rather than with a reference range. It is most useful to examine serial test results and correlate changes with a patient’s clinical picture.

  2. Normal results should be compared with previous results, if available. Inflammatory states may increase the rate of synthesis and elevate serum complement protein levels. For example, some patients with SLE have concentrations of specific complement components that are two to three times the upper limit of normal (ULN) when their disease is clinically inactive. When the disease activity increases to the point that increased catabolism of complement proteins occurs, levels may then fall into the reference range. It would be a misinterpretation to conclude that these “normal” concentrations represent an inactive complement system. Consequently, serial determinations of complement levels may be more informative than measurements at a single point.

  3. Complement responses do not correlate consistently with disease activity. In some patients, the increase and decrease of the complement system should not be used to assess disease activity.

Assessment of the complement system should include measurement of the total hemolytic complement activity by the complement hemolytic 50% (CH50) test and determination of the levels of C3 and C4.

Total Hemolytic Complement

Reference range: 150 to 250 International Units/mL

The total hemolytic complement (THC or CH50) measures the ability of a patient’s serum to lyse 50% of a standard suspension of sheep erythrocytes coated with rabbit antibody. All nine components of the classic pathway are required to produce a normal reaction. The CH50 screening test may be useful when a complement deficiency is suspected or a body fluid other than serum is involved. For patients with SLE and lupus nephritis, serial monitoring of CH50 may be useful for guiding effectiveness of drug therapy.

C3 and C4

Reference ranges: C3, 80 to 160 mg/dL or 0.8 to 1.6 g/L; C4, 16 to 48 mg/dL or 0.16 to 0.48 g/L

Because C3 is the most abundant complement protein, it was the first to be purified and measured by immunoassay. However, C4 concentrations appear to be more sensitive to smaller changes in complement activation and more specific for identifying complement activation by the classic pathway. Results of C3 and C4 testing are helpful in following patients who initially present with low levels and then undergo treatment, such as those with SLE.

Acute-Phase Reactants

The concentration of a heterogeneous group of plasma proteins, called acute-phase proteins or acute-phase reactants, increases in response to inflammatory stimuli such as tissue injury and infection. Concentrations of CRP, serum amyloid A protein, α1-acid glycoprotein, α1-antitrypsin, fibrinogen, haptoglobin, ferritin, and complement characteristically increase, whereas serum transferrin, albumin, and prealbumin (transthyretin) concentrations decrease. Their collective change is referred to as the acute-phase response.

In general, if the inflammatory stimulus is acute and of short duration, these proteins return to normal within days to weeks. However, if tissue injury or infection is persistent, acute-phase changes may also persist. Additionally, white blood cell (WBC) and platelet counts may be elevated significantly.

Rheumatic diseases are chronic and associated with varying severities of inflammation. The ESR and CRP are two tests that can be helpful in three ways: (1) estimating the extent or severity of inflammation, (2) monitoring disease activity over time, and (3) assessing prognosis.13 Unfortunately, both tests are nonspecific and cannot be used to confirm or exclude any particular diagnosis. However, the results may be helpful in RA, for example, for inclusion in an assessment of a patient’s overall disease activity (ie, disease activity score - 28) and subsequent treatment modifications.

Erythrocyte Sedimentation Rate

Reference range (Westergren method): men, 0 to 15 mm/hr; women, 0 to 20 mm/hr

For many years, the erythrocyte sedimentation rate (ESR) has been used widely as a reflection of the acute-phase response and inflammation. The test is performed by placing anticoagulated blood in a vertical tube and measuring the rate of fall of erythrocytes in millimeters/hour. In rheumatic diseases, the ESR is an indirect screen for elevated concentrations of acute-phase plasma proteins, especially fibrinogen.13 An elevated ESR occurs when elevated protein concentrations (especially fibrinogen) cause aggregation of erythrocytes, resulting in a faster fall of those cells in the tube.

Several factors unrelated to inflammation may result in an increased ESR, such as obesity and increasing age. The ESR also responds slowly to an inflammatory stimulus. Despite these limitations, the test remains in wide use because it is inexpensive and easy to perform, and a tremendous amount of data are available about its clinical significance in numerous diseases. The Westergren method of performing an ESR test is preferred over the Wintrobe method because of the relative ease of performing the former method in clinical or laboratory settings.

Correlation of serial Westergren ESR results with patient data may influence therapeutic decisions. Two rheumatic diseases, polymyalgia rheumatica and temporal arteritis (giant cell arteritis), are almost always associated with an elevated Westergren ESR. The ESR is usually >60 mm/hr but can be markedly elevated >100 mm/hr in these disorders. During initial therapy or treatment initiated after a disease flare, a significant decrease or a return to a normal ESR usually indicates that systemic inflammation has decreased substantially. In the absence of clinical symptoms, an increased ESR may indicate that more aggressive therapy is needed. Disease activity can then be monitored by ESR results. Of course, if symptoms are present, they should be treated.

C-Reactive Protein

Reference range: 0 to 0.5 mg/dL or 0 to 0.005 g/L

C-reactive protein (CRP) is a plasma protein of the acute-phase response. In response to a stimulus such as injury or infection, CRP can increase up to 1,000 times its baseline concentration. The precise physiologic function of CRP is unknown, but it is known to participate in activation of the classic complement pathway and interact with cells in the immune system.

Serum CRP levels can be quantitated accurately and inexpensively by immunoassay or laser nephelometry. Most healthy adults have concentrations of <0.3 mg/dL, although concentrations of 1 mg/dL are sometimes seen. Moderate increases range from 1 to 10 mg/dL, and marked increases are >10 mg/dL.13 Values >15 to 20 mg/dL are usually associated with bacterial infections. In general, concentrations >1 mg/dL reflect the presence of a significant inflammatory process. As with the Westergren ESR, serial measurements of CRP are the most valuable, especially in chronic inflammatory diseases.

Currently, the routine use of CRP for the assessment of rheumatic diseases is limited. As with the ESR, CRP concentrations generally increase and decrease with worsening and improving signs and symptoms, respectively. Nevertheless, CRP concentrations are not specific for any disease.

Using an assay method called high-sensitivity CRP (hs-CRP), several studies have shown a correlation between elevated levels and cardiovascular events, including myocardial infarction. The units of measurement for the hs-CRP (milligrams/liter) are different from those of the conventional CRP test (milligrams/deciliter). Because CRP levels fluctuate over time, when the hs-CRP is used for cardiovascular risk assessment, the test should be measured twice at least 2 weeks apart and the two values averaged.14

Human Leukocyte Antigen B27

Human leukocyte antigen B27 (HLA-B27) is an antigen on the surface of WBCs encoded by the B locus in the major histocompatibility complex on chromosome 6. The HLA-B27 test is qualitative and is either present or absent. Its presence is associated with autoimmune diseases known as seronegative spondyloarthropathies. An HLA-B27 test may be ordered when a patient has pain and inflammation in the spine, neck, chest, eyes, or joints and an autoimmune disorder associated with the presence of HLA-B27 is the suspected cause. The test may be obtained to confirm a suspected diagnosis of ankylosing spondylitis, Reiter syndrome, or anterior uveitis. However, a positive test result cannot distinguish among these diseases and cannot be used to predict progression, severity, prognosis, or degree of organ involvement. Some patients with these disorders may have a negative HLA-B27 test. Further, the test cannot definitively diagnose or exclude any rheumatic disease. It is frequently ordered in concert with other rheumatic tests (eg, RF, ESR, CRP), based on the clinical presentation.

A positive HLA-B27 in a person without symptoms or a family history of HLA-B27–associated disease is not clinically significant and in this case does not predict the likelihood of developing an autoimmune disease. However, the presence or absence of HLA antigens can be genetic. For example, it does not help predict the likelihood of developing an autoimmune disease. The presence or absence of HLA antigens is genetically determined. If a family member has an HLA-B27–related rheumatic disease, other family members who share the HLA-B27 antigen have a higher risk of developing a similar disease. Individuals who already know they are HLA-B27 positive may wish to seek genetic counseling to understand the hereditary impact on their family.

New genetic testing methods permit separation of HLA-B27 into subtypes. Approximately 15 subtypes have been identified, the most common of which are HLA-B*2705 and HLA-B*2702. The precise clinical significance of individual subtypes is an area of continuing investigation.15

Synovial Fluid Analysis

Synovial fluid is essentially an ultrafiltrate of plasma to which synovial lining cells add hyaluronate. This fluid lubricates and nourishes the avascular articular cartilage. Normally, synovial fluid is present in small amounts and is clear and acellular (<200 cells/mm3) with a high viscosity because of the hyaluronic acid concentration. Normal fluid does not clot because fibrinogen and clotting factors do not enter the joint space from the vascular space. Protein concentration is approximately one-third that of plasma, and glucose concentration is similar to that of plasma.

When performing arthrocentesis (joint aspiration), a needle is introduced into the joint space of a diarthrodial joint. With a syringe, all easily removed synovial fluid is drained from the joint space. Arthrocentesis is indicated as a diagnostic procedure when septic arthritis, hemarthrosis (bleeding into a joint space), or crystal-induced arthritis is suspected. Furthermore, arthrocentesis may be indicated in any clinical situation, rheumatic or nonrheumatic, if the cause of new or increased joint inflammation is unknown. Arthrocentesis is also performed to administer intraarticular corticosteroids.

When arthrocentesis is performed, the synovium may be inflamed, allowing fibrinogen, clotting factors, and other proteins to diffuse into the joint. Therefore, the collected synovial fluid should be placed in heparinized tubes to prevent clotting and allow determination of cell type and cell number. If diagnostic arthrocentesis is indicated, the aspirated joint fluid should be analyzed for volume, clarity, color, viscosity, cell count, culture, glucose, and protein. Synovial fluid is subsequently reported as normal, noninflammatory, inflammatory, or septic.16 Table 20-3 presents the characteristics of normal and three pathologic types of synovial fluid. The presence and type of crystals in the fluid should be determined. The presence of crystals identified by polarized light microscopy with red compensation can be diagnostic (Table 20-4) (Minicase 1).

TABLE 20-3.

Synovial Fluid Characteristics and Classification

CHARACTERISTIC

NORMAL

NONINFLAMMATORY

INFLAMMATORY

SEPTIC

Viscosity

High

High

Low

Variable

Color

Colorless to straw

Straw to yellow

Yellow

Variable

Clarity

Translucent

Translucent

Translucent–opaque

Opaque

WBC (/mm3)

<200

200–3,000

2,000–50,000

>50,000

PMN

<10%

<10%

Often >50%

>75%

Culture

Negative

Negative

Negative

Often positive

Glucose (a.m. fasting)

≅ blood

≅ blood

>25 mg/dL but lower than blood

<25 mg/dL (much lower than blood)

Protein (g/dL)

1–2

1–3

3–5

3–5

PMN = polymorphonuclear neutrophils; WBC = white blood cells.

TABLE 20-4.

Morphology of Synovial Fluid Crystals Associated with Joint Disease

CRYSTALS

SIZE (μm)

MORPHOLOGY

BIREFRINGENCEa

DISEASES

Monosodium urate

2–10

Needles, rods

Negative

Gout

CPPD

2–10

Rhomboids, rods

Positive (weak)

CPPD crystal deposition disease (pseudogout), OA

Calcium oxalate

2–10

Polymorphic, dipyramidal shapes

Positive

Renal failure

Cholesterol

10–80

Rectangles with notched corners; needles

Negative or positive

Chronic rheumatoid or osteoarthritic effusions

Depot corticosteroids

4–15

Irregular rods, rhomboids

Negative or positive

Iatrogenic postinjection flare

CPPD = calcium pyrophosphate dihydrate.

aThe property of birefringence is the ability of crystals to pass light in a particular plane. When viewed under polarized light, the crystals are brightly visible in one plane (birefringent) but are dark in a plane turned 90°. Birefringence observed under polarized light can be categorized as “positive” and “negative” based on the speed at which rays of light travel through the crystals in perpendicular planes (at right angles).

Nonrheumatic Tests

The three most commonly performed groups of nonrheumatic tests performed in rheumatology are the complete blood count (CBC), serum chemistry panel, and urinalysis. These tests are not specific for any rheumatic disorder, and abnormal results may occur in association with many rheumatic and nonrheumatic diseases. These tests are discussed from a more general perspective in other chapters.

Chronic inflammatory diseases such as RA and SLE are commonly associated with anemia. Microcytic anemia may occur as a result of drug therapy for rheumatic diseases (eg, gastroduodenal hemorrhage from nonsteroidal anti-inflammatory drugs [NSAIDs]). Autoimmune hemolytic anemia may be seen in SLE and other rheumatic diseases and is associated with a rapid onset that may be life threatening. The platelet count may be elevated in some disorders (thrombocytosis) and decreased in others (thrombocytopenia). Leukopenia may be associated with Felty syndrome and may also be caused by therapy with immunosuppressive agents used to treat rheumatic diseases. For example, thrombocytopenia and leukopenia may be seen in cases of SLE.

Antiphospholipid syndrome may be seen with SLE and other disorders and is associated with abnormalities in coagulation. Patients with antiphospholipid antibodies should be evaluated further to determine thrombosis risk.

Systemic lupus erythematosus may be associated with hepatic dysfunction, which can be assessed by determination of hepatic lab tests. Some drugs used in the treatment of rheumatic disease may also cause hepatic injury. Renal function tests (usually the serum creatinine [SCr] and blood urea nitrogen [BUN]) may provide evidence of renal involvement in patients with lupus nephritis. Proteinuria, hematuria, and pyuria may be seen in cases of SLE and with use of drugs to treat rheumatic disorders.

Assessment of Synovial Fluid

A 45-year-old woman presents to her physician with reports of pain and swelling in her hands and feet over the last 8 weeks. Physical examination reveals swollen and tender joints bilaterally and limited ability to close a fist or move her thumbs. She reports that she wakes up with stiff joints and it usually takes 60 to 90 minutes until she can use her hands effectively, sometimes longer. The synovial fluid from one swollen joint is aspirated. The aspirate, noted to be thin, cloudy, and yellow, is sent in heparinized tubes to the laboratory for Gram stain, bacterial culture, cell count, and chemistry panel. After receiving the pathology report, the physician reviews the preliminary laboratory results of the aspirate (refer to Table 20-3 for reference values):

  • 45,000 WBCs/mm3

  • 75% PMNs

  • 23 mg/dL glucose

  • 4.5 g/dL protein

  • Culture, negative

QUESTION: What is the likely diagnosis in this patient? What additional laboratory studies should be performed?

DISCUSSION: Based on physician observations and the pathology report, the aspirate appears to be inflammatory because the viscosity is low, the clarity is not translucent, and an infection is not present. Considered with the patient’s age, gender, joint presentation (bilateral, small joints), and stiffness that lasts for 60 minutes or more, the likely diagnosis is rheumatoid arthritis.

Additional testing should be considered to confirm a diagnosis of RA—specifically, ACPA, RF, and acute-phase reactants (ESR, CRP). Baseline CBC, chemistry panels, and assessment of renal and hepatic function should be conducted if recent results are not available. ACPA is especially helpful because it is the most specific for RA. In addition to confirming the diagnosis, a positive ACPA result, if present, is also indicative of more erosive disease and worse long-term prognosis. An early diagnosis allows aggressive treatment to be initiated with the goal of controlling systemic inflammation and slowing or stopping the erosions. RF alone is not specific enough to indicate a diagnosis of RA, but when combined with ACPA, the specificity of diagnosis, if both are positive, increases to 99.5%. The results of ESR and CRP are not specific to RA, but they provide information about the ongoing inflammation. The results are also helpful while the physician is evaluating the disease activity. The remaining test results are helpful for understanding the baseline status of the patient and determining treatment options that are safe for her to receive.

INTERPRETATION OF LABORATORY TESTS IN SELECT RHEUMATIC DISEASES

Rheumatoid Arthritis in Adults

In 2010, the American College of Rheumatology (ACR) and the European League Against Rheumatism (EULAR) published new classification criteria for RA.17 The criteria were developed to help define homogeneous treatment populations for research trials, not for clinical diagnosis. In practice, the clinician must establish the diagnosis for an individual patient using many more aspects (including perhaps some additional laboratory tests) based on the clinical presentation. Nevertheless, the formal classification criteria are useful as a general guide to making a clinical diagnosis. The 2010 criteria are aimed at diagnosing RA earlier in newly presenting patients so patients can be started on treatment sooner, with the goal of preventing erosive joint damage and improving long-term outcomes. Patients with erosive disease typical of RA and those with longstanding disease (including patients whose disease is inactive with or without treatment) should also be classified as having RA if they previously fulfilled the 2010 criteria.17

Two mandatory criteria must be met before the classification criteria can be applied to an individual patient. First, there must be evidence of definite clinical synovitis in at least one joint as determined by an expert assessor; the distal interphalangeal joints, first carpometacarpal joints, and first metatarsophalangeal joints are excluded from consideration because these joints are usually involved in osteoarthritis (OA). Second, the synovitis cannot be better explained by another disease such as SLE, psoriatic arthritis, or gout. When these two criteria are met, classification of definite RA is based on achieving a score of 6 or more out of 10 points in four domains:

  1. Joint distribution

    • 1 large joint (shoulder, elbow, hip, knee, ankle) = 0 points

    • 2 to 10 large joints = 1 point

    • 1 to 3 small joints (metacarpophalangeal joints, proximal interphalangeal joints, second to fifth metatarsophalangeal joints, thumb interphalangeal joints, wrists) = 2 points

    • 4 to 10 small joints = 3 points

    • – More than 10 joints (with at least 1 small joint) = 5 points

  2. Serology

    • Negative RF and ACPA = 0 points

    • Low-positive RF or ACPA = 2 points

    • High-positive RF or ACPA = 3 points

  3. Acute-phase reactants

    • Normal CRP and ESR = 0 points

    • Elevated CRP or ESR = 1 point

  4. Symptom duration (by patient report)

    • Less than 6 weeks = 0 points

    • 6 weeks or more = 1 point

Patients who do not achieve a score of 6 or higher should be regularly reassessed because they may meet the classification criteria cumulatively over time.

Anticitrullinated Protein Antibodies

Patients with a diagnosis of RA should be treated aggressively early in the disease course because in 90% of patients most damage from bone erosions occurs within the first 2 years. For this reason, a timely and accurate early diagnosis is critical. Many patients with early RA have mild, nonspecific symptoms; in these cases, the ability to detect a disease-specific antibody such as ACPA could be of crucial diagnostic and therapeutic importance.

The ACPA test has several useful characteristics as a marker for RA diagnosis and prognosis: (1) it is as sensitive as RF and more specific than RF for RA in patients with early as well as fully established disease; (2) it may be detectable in seemingly healthy persons years before the onset of clinical RA findings; (3) it may predict the future development of RA in patients with undifferentiated arthritis; and (4) it is a predictor for the eventual development of erosive disease. As stated previously, ACPAs can be detected in about 50% to 60% of patients with early RA, usually after having nonspecific symptoms for 3 to 6 months prior to seeing a physician.

In some patients with nonspecific arthritis, it is difficult to make a definitive diagnosis of RA because of the lack of disease-specific serum markers for other conditions in the differential diagnosis. In some situations, the presence of ACPAs may help differentiate RA from polymyalgia rheumatica or erosive forms of SLE.3

Several reports suggest that patients with early RA who are ACPA positive develop more erosive disease than those who are antibody-negative.3 Early identification of patients who are at risk for a more severe disease course could lead to more rapid and aggressive institution of disease-modifying therapies. However, clinical trials are needed to determine whether this diagnostic and therapeutic approach is indeed beneficial.

Rheumatoid Factor

In patients with RA, affected diarthrodial joints have an inflamed and proliferating synovium infiltrated with T lymphocytes and plasma cells. Plasma cells in the synovial fluid generate large amounts of IgG RF and abnormally low amounts of normal IgG. However, plasma cells in the bloodstream of patients with RA produce IgM RF predominantly.2

Approximately 75% to 80% of adults with RA have a positive RF titer, and most of those who are positive have titers of at least 1:320. A positive RF is not specific for the diagnosis of RA. Positive RF titers are also associated with some connective disease, such as SLE and Sjögren syndrome. RF levels may also be increased in some infections (eg, malaria, rubella, hepatitis C). Furthermore, up to 5% of the normal healthy population may be RF positive. Patients with RA generally have higher RF titers than individuals with other nonrheumatic conditions. In patients who have RA and a positive RF, the titer generally increases as disease activity (inflammation) increases. Consequently, as the serum RF titer increases, the specificity of the test for the diagnosis of RA also increases.2 Higher titers or serum concentrations suggest the presence of more severe disease than lower levels and are associated with a worse prognosis.

Rheumatoid factor is one of the two serologic tests (along with ACPA) included in the 2010 ACR/EULAR classification criteria for RA.17 Based on the reporting of RF levels in International Units/milliliter, a negative RF is considered to be less than or equal to the ULN for the laboratory and assay. A low-positive test is higher than the ULN but less than or equal to three times the ULN. A high-positive test is more than three times the ULN for the assay. When the RF is reported as only positive or negative by the laboratory, patients reported as having a positive RF should be scored as “low-positive” for RF scoring purposes.17

Although RFs are usually identified and quantified from serum samples, RA is a systemic, extravascular, autoimmune disease affecting the synovium. As a result, some RFs may be present in sites other than peripheral blood. IgG RFs are found in the synovial fluid of many patients with severe RA. IgA RF may be detected in the saliva of patients with RA or Sjögren syndrome. The presence of IgE RF is correlated with extraarticular findings of RA.2

Although most patients with RA are seropositive for RF, some patients have negative titers. However, some of these patients may have non-IgM RF, predominantly IgG RF. Also, some seronegative patients convert to seropositive on repeat testing. A small percentage of adult patients with RA (<10%) are considered to be truly seronegative. When compared with RF-positive patients, seronegative patients usually have milder arthritis and are less likely to develop extraarticular manifestations (eg, rheumatoid nodules, lung disease, and vasculitis).

Because current treatment guidelines call for aggressive early treatment of RA—before end-organ damage—clinicians must be aware of the relationship between disease onset and RF development. Unfortunately, the RF test is least likely to be positive at the onset of RA, when it might be of the most help. After RA has been diagnosed, RF titers are not routinely used to assess a patient’s current clinical status or modify a therapeutic regimen. A specific titer or a change in titers for an individual does not correlate reliably with disease activity.

In summary, RF is not sensitive or specific enough to use as the sole laboratory test to diagnose or manage RA. Although it is present in msot patients with RA, it is negative in some patients with the disease. RF may be useful as a prognostic indicator, because patients with RA with high RF titers generally have a more severe disease course.

Antinuclear Antibodies

Antinuclear antibodies are usually negative in patients with RA. The frequency of positive ANAs in patients with RA is highly variable. As determined by indirect immunofluorescence, this frequency varies from 10% to 70%, depending on the substrate used and the titer considered positive. In patients with a positive ANA, tests for dsDNA and Sm antibodies should be performed because these tests are highly specific for SLE.

Complement

The serum complement level is usually normal or elevated in RA. Complement elevations often occur as part of the acute-phase response. These increases parallel changes in other acute-phase proteins (eg, CRP). Elevations of total hemolytic activity (CH50), C3, and C4 are usually observed during active stages of most rheumatic diseases, including RA. The presence of circulating immune complexes in RA may lead to hypercatabolism of complement and acquired hypocomplementemia.

Acute Phase Reactants

As in other inflammatory diseases, the nonspecific ESR test is usually elevated in active RA. The degree of elevation is directly related to the severity of inflammation; the Westergren ESR can be 50 to 80 mm/hr or more in patients with severely active RA. It usually decreases or normalizes when systemic inflammation decreases during initial treatment or after treatment for a disease flare. However, there is a large variability in response to treatment among individuals. Subsequent increases in disease activity will be mirrored by corresponding increases in ESR.

C-reactive protein levels may be elevated (approximately 2 to 3 mg/dL) in adult patients with RA with moderate disease activity. However, there is substantial individual variability, and 5% to 10% of such patients have normal values. Some patients with severe disease activity have levels of 14 mg/dL or higher.

Because of their nonspecificity, the ESR and CRP are of little use in distinguishing between RA and other rheumatic diseases, such as OA or mild SLE. The tests also are elevated in patients with vasculitis associated with RA and reflect the generalized systemic inflammatory state. These tests are more appropriate for monitoring disease activity in RA. Elevations of ESR and CRP are individually associated with radiologic damage in RA as assessed by the number of joint erosions. Elevation of both ESR and CRP is a stronger predictor of radiologic progression than an elevation in CRP alone.18,19

Synovial Fluid Analysis

If a joint effusion is present and the diagnosis is uncertain, arthrocentesis and synovial fluid analysis are performed to exclude gout, pseudogout, or infectious arthritis.18 Synovial fluid analysis should include WBC count with differential, analysis for crystals, and Gram stain and culture. In early RA, the analysis typically reveals straw-colored, turbid fluid with fibrin fragments. A clot forms if the fluid is left standing at room temperature. There are usually 5,000 to 25,000 WBC/mm3, at least 85% of which are polymorphonuclear neutrophils (PMN). Complement C4 and C2 levels are usually slightly decreased, but the C3 level is generally normal. The glucose level is decreased, sometimes to <25 mg/dL. No crystals should be present, and cultures should be negative.

Nonrheumatic Tests

The CBC may reveal an anemia that is either normochromic-normocytic or hypochromic-microcytic. Microcytic anemia is due to iron deficiency that may result from gastrointestinal blood loss associated with drug use (eg, NSAIDs) or other causes. The WBC count may show a slight leukocytosis with a normal differential. Eosinophilia (>5% of the total WBC count) may be associated with RF-positive severe RA. Felty syndrome may be associated with granulocytopenia. Thrombocytosis may be present in clinically active RA as part of the acute-phase response. As the disease improves spontaneously or as a result of drug therapy, the platelet count returns toward normal.

Osteoarthritis

Osteoarthritis (OA) results from the complex interplay of numerous factors, such as joint integrity, genetics, mechanical forces, local inflammation, and biochemical processes.20,21 It is generally considered to be a noninflammatory arthritis; however, inflammation may be present. It is not considered an autoimmune disease. The synovium is normal, and the synovial fluid usually lacks inflammatory cells. Although affected joints are painful, they are frequently not inflamed. The primary use of laboratory tests when OA is suspected is to rule out other disorders in the differential diagnosis.

No clinical laboratory tests are specific for the diagnosis of OA. Laboratory tests that may be performed in patients suspected of having OA include ESR, RF titers, and evaluation of synovial fluid.21 The ESR (and CRP) is usually normal but may be slightly increased if inflammation is present. The RF test is negative, and serum chemistries, hematology tests, and urinalysis are normal. Research is underway to determine if certain biomarkers (ie, CRP), if elevated, correlate with disease activity or radiographic progression.

Synovial fluid analysis may be undertaken, especially in patients with severe, acute joint pain. Findings generally reveal either a noninflammatory process or mild inflammation (WBC <2,000 cells/mm3). Crystals are absent when the synovial fluid is examined using compensated polarized light microscopy (Minicase 2).

Juvenile Idiopathic Arthritis

The term juvenile idiopathic arthritis (JIA) encompasses a heterogeneous group of childhood arthritis conditions of unknown cause. Because JIA includes a variety of arthritis categories that differ from adult-onset RA, the term JIA has replaced the name juvenile rheumatoid arthritis.22 Juvenile idiopathic arthritis begins before 16 years of age and persists for at least 6 weeks; other known causes must be excluded before the diagnosis of JIA can be made. Juvenile idiopathic arthritis is divided into categories based on presenting clinical and laboratory findings: (1) systemic arthritis, (2) oligoarthritis, (3) polyarthritis (RF negative), (4) polyarthritis (RF positive), (5) psoriatic arthritis, (6) enthesitis-related arthritis, and (7) undifferentiated arthritis.22-25

The diagnosis of JIA is made primarily on clinical grounds. No single laboratory test or combination of tests can confirm the diagnosis. However, laboratory tests can be useful in providing evidence of inflammation, supporting the clinical diagnosis, and monitoring toxicity from therapy.

Evaluation of Arthritis

A 62-year-old postmenopausal woman presents to her family physician with reports of pain and stiffness in her knees for the last month. She notes the pain is worse in the morning when she wakes up and resolves about 20 minutes later. Her past medical history is significant for hypertension and dyslipidemia for approximately 5 years. She quit smoking 1 year ago following a 20 pack-per-year smoking history. She started a diet (low fat, low sodium) and exercise (walking 30 minutes daily) program about 2 months ago. On physical exam, no swelling or tenderness is found in her knees. Significant crepitus and decreased range of motion are noted. No additional joints appear to be affected. Current medications (stable for the past 6 months) include lisinopril, hydrochlorothiazide, atorvastatin, and one aspirin tablet a day. She is allergic to sulfa and intolerant to penicillin (stomach upset). She started taking ibuprofen about 1 month ago in response to the joint pain.

Laboratory results obtained at this visit:

  • RF titer 10 International Units/mL

  • ACPA 10 EU

  • ANA 1:40

  • ESR 15 mm/hr

  • CRP 0.3 mg/dL

QUESTIONS: What are two likely diagnoses, which one is most likely, and what data support that diagnosis?

DISCUSSION: The two most common causes of joint pain and stiffness include OA and RA. Upon initial presentation, either may be a possibility. The physical exam findings are key to identifying joints that are typically affected by RA versus OA. Morning stiffness is a common symptom of either condition; however, morning stiffness associated with OA typically lasts for less than 30 minutes, whereas stiffness associated with RA lasts an hour or more. The laboratory results are also helpful in distinguishing between the two conditions. First, OA does not involve systemic inflammation, so it is unlikely to cause elevated acute-phase reactants (ESR and CRP). Second, RF is negative; however, the presence of positive RF is not specific for a diagnosis of RA. Finally, ACPA is sensitive and specific for RA, and the result is negative. The clinical presentation (ie, large joints, no inflammation, joint stiffness that resolves in less than 30 minutes) and the normal/negative lab results indicate the patient is experiencing OA.

Rheumatoid Factor

Rheumatoid factor–positive polyarthritis constitutes 5% to 10% of JIA cases and is the childhood arthritis that is most similar to adult RA. It is defined as arthritis affecting five or more joints in the first 6 months of disease with a positive RF test on two occasions at least 3 months apart.22 RF-positive polyarthritis is six to 12 times more common in girls than boys. As in adult RA, the RF test usually detects IgM-anti-IgG. RF-negative polyarthritis constitutes 20% to 30% of new JIA cases. It also includes arthritis in five or more joints during the first 6 months, but the RF test is negative. Although the presence of RF overall is low in JIA, its presence indicates more aggressive disease.

Oligoarthritis is the most common form of JIA; it is four times more common in girls than boys and has a peak onset before the age of 6 years. Oligoarthritis affects four or fewer joints in the first 6 months; the RF test is usually negative. The RF is negative in systemic arthritis, psoriatic arthritis, enthesitis-related arthritis, and undifferentiated arthritis.22

Anticitrullinated Protein Antibodies

The APCA test alone is not helpful in diagnosing a subcategory of JIA. This is consistent with the fact that JIA is a heterogeneous group of disorders, most of which are different from adult RA. Similar to RA in adults, positive ACPA results in JIA have been associated with RF-positive disease and erosive arthritis.22 Approximately 60% to 70% of patients with RF-positive polyarthritis are ACPA positive. Approximately 50% to 80% of patients with RF-negative polyarthritis are ACPA positive.22

Antinuclear Antibodies

In oligoarthritis, 70% to 80% of children have positive (low to moderate titer) ANA tests, typically 1:40 to 1:320. ANA positivity is high in girls with an early onset of disease. Although ANA is not useful for monitoring or predicting a patient’s disease or symptoms, it is helpful to determine risk for developing uveitis. Patients diagnosed with oligo- or polyarticular arthritis at an early age who are ANA positive are at a higher risk of uveitis. This is especially important because a patient may be asymptomatic and thereby undiagnosed and untreated, potentially leading to permanent vision loss. The ANA test is positive in 40% of patients with RF-negative polyarthritis and positive in up to 55% of patients with RF-positive polyarthritis. The ANA is positive in about 15% to 20% of children with psoriatic arthritis. It may be positive in some patients with enthesitis-related arthritis. The test result is seldom positive (<10%) in children with systemic JIA.

Complement

As with adult-onset RA, serum complement components (especially C3) are usually elevated in systemic JIA.

Acute-Phase Reactants

In systemic arthritis, the ESR and CRP are typically high during an acute flare. In oligoarthritis, there is little systemic inflammation, and the ESR and CRP are usually normal. Some cases of oligoarthritis may be associated with mildly or moderately elevated ESR or CRP; however, elevated acute-phase reactants in this category should raise suspicion for other conditions, such as subclinical inflammatory bowel disease associated with arthropathy. Acute-phase reactants may be elevated in either RF-positive or RF-negative polyarthritis and in psoriatic arthritis. The ESR may be elevated in enthesitis-related arthritis, but this abnormality should raise suspicion for subclinical inflammatory bowel disease. Acute-phase reactants are typically mildly elevated in patients with the psoriatic arthritis subcategory of JIA.

Synovial Fluid Analysis

Arthrocentesis in JIA is typically consistent with inflammatory fluid. As in adult RA, synovial fluid glucose levels are low.

Nonrheumatic Tests

Children with systemic arthritis may have anemia, leukocytosis with neutrophilia, and thrombocytosis. The anemia is normochromic-normocytic (anemia of chronic disease); hemoglobin values may be in the range of 7 to 10 g/dL. WBC counts in the range of 20,000 to 30,000 cells/mm3 are not uncommon, and counts may exceed 60,000 to 80,000 cells/mm3. In severe cases, liver enzymes, ferritin, and coagulation screen also may be abnormal. Patients with enthesitis-related or psoriatic arthritis may have a mild anemia of chronic disease.

Systemic Lupus Erythematosus

Criteria for the classification of systemic lupus erythematosus (SLE) were updated by the EULAR and ACR in 2019. The criteria were updated to address shortcomings of previous criteria, including a need to improve sensitivity and specificity of the criteria and earlier detection of patients with early disease for inclusion in research trials. The wide variety of manifestations and unpredictable course often make SLE difficult to diagnose.

According to the new classification criteria, the first criterion that must be met is a positive ANA at a titer of ≥ 1:80. This is a significant change from previous criteria and the expert panel recommends further research into the very small group of patients that are ANA-negative and displaying symptoms similar to SLE. The new criteria include clinical domains (ie, hematologic, mucocutaneous, musculoskeletal, renal) and immunology domains (ie, antiphospholipid antibodies, SLE-specific antibodies). The classification criteria now follow a weighed system whereby clinical and immunologic features are given higher weighting to align with their significance. The weighting ranges from 2 to 10. A patient’s presentation is classified as SLE if 10 or more points are scored across all domains and at least one clinical criterion is present. When the criteria were applied to a cohort for validation purposes, the sensitivity slightly decreased to 96.1% and specificity increased to 93.4% when compared with previous criteria, 96.7% and 83.7%, respectively.26,27

Antinuclear Antibodies

Antinuclear antibody testing is usually performed initially if SLE is suspected because of its high sensitivity and ease of use. To be considered in the classification criteria, a patient must pass the entry criterion, which is a positive ANA (titer ≥1:80). The ANA test has low specificity for SLE; many other conditions are associated with a positive test result (eg, systemic sclerosis, polymyositis, dermatomyositis, RA, autoimmune thyroiditis or hepatitis, infections, malignancies, and many drugs). Some healthy persons also may have a positive ANA test result. Consequently, results of an ANA test are always interpreted in light of a patient’s clinical presentation.

Anti-dsDNA and anti-ssDNA antibodies

The anti–double-stranded DNA (anti-dsDNA) test is positive in 50% to 60% of patients with SLE at some point in the disease course, and the test is 95% specific for SLE. In contrast, testing for antibodies to single-stranded DNA (anti-ssDNA) has poor specificity for SLE but is more sensitive (90%). Although the anti-ssDNA antibody appears to be important in the immunopathogenesis of SLE, the test has little diagnostic utility because of poor specificity. There is evidence that anti-dsDNA and anti-ssDNA antibodies are important in the pathogenesis of lupus nephritis because they appear to correlate with its presence and severity. Titers of these antibodies tend to fall with successful treatment, frequently becoming undetectable during sustained remission.

Anti-Sm, Anti-Ro/SSA, and Anti-La/SSB Antibodies

The anti-Sm antibody is an immunoglobulin specific against Sm, a ribonucleoprotein found in the cell nucleus. The anti-Sm antibody test is positive in 20% to 30% of patients with SLE, and presence of these antibodies is pathognomonic for SLE.26 Anti-Ro/SSA and anti-La/SSB antibodies are present in 30% and 20% of patients with SLE, respectively, but they are not specific for the disease.

Complement

Total CH50 levels are decreased at some point in most patients with SLE. Complement levels decrease in SLE because of deposition of immune complexes in active disease (hypercatabolism). Complement depletion has been associated with increased disease severity, particularly renal disease. Analysis of various complement components has revealed low levels of C1, C4, C2, and C3. Serial determinations have demonstrated that decreased levels may precede clinical exacerbations.11 As acute episodes subside, levels return toward normal. Some experts consider it helpful to follow complement measurements in SLE patients receiving treatment, especially if C4 and C3 were low at the time of diagnosis.11

Acute-Phase Reactants

Serum ESR and CRP concentrations are elevated in many patients with active SLE, but many individuals have normal CRP levels. Those with acute serositis or chronic synovitis are most likely to have markedly elevated CRP levels. Patients with other findings of SLE, such as lupus nephritis, may have modest or no elevations.

Several studies have examined the hypothesis that elevations in CRP during the course of SLE result from superimposed infection rather than activation of SLE. In hospitalized patients, substantially elevated serum CRP levels occur most frequently in the setting of bacterial infection. Consequently, CRP elevations >6 to 8 mg/dL in patients with SLE (as well as other diseases) should signal the need to exclude the possibility of infection. Such CRP increases should not be considered proof of infection because CRP elevation can be related to active SLE in the absence of infection.

Nonrheumatic Tests

Antiphosopholipid antibodies (ie, anticardiolipin antibodies and the lupus anticoagulant) can occur as an idiopathic disorder and in patients with autoimmune and connective tissue diseases such as SLE.28,29 Anticardiolipin antibody and the lupus anticoagulant are closely related but are different antibodies. Consequently, an individual can have one antibody and not the other. The presence of these antibodies may increase the risk of future thrombotic events. This clinical situation is referred to as antiphospholipid syndrome (APS). When APS occurs in patients with no other diagnosis, it is referred to as primary APS. Patients who also have SLE or another rheumatic disease are said to have secondary APS.

Anemia is present in many patients with SLE. The CBC may reveal a normochromic-normocytic anemia (anemia of chronic disease) that is not associated with erythropoietin deficiency. Hemolytic anemia with a compensatory reticulocytosis also may occur due to antierythrocyte antibodies. Most patients also have a positive Coombs test result. Anemia in SLE also can result from blood loss, renal insufficiency, medications, infection, hypersplenism, and other reasons.30

Leukopenia is present in approximately 50% of patients but is usually mild. It results primarily from decreased numbers of lymphocytes, which may be caused by the disease or its treatment. If the patient is not being treated with corticosteroids or immunosuppressive agents, ongoing immunologic activity should be suspected. Neutropenia in SLE may occur from immune mechanisms, medications, bone marrow suppression, or hypersplenism.30 Mild thrombocytopenia (100,000 to 150,000/mm3) occurs in 25% to 50% of patients with SLE and is usually due to immune-mediated platelet destruction. Increased platelet consumption and impaired platelet production also may be contributing factors.30

Liver function tests may reveal increased hepatic aminotransferases (AST, ALT), lactate dehydrogenase, and alkaline phosphatase in patients with active SLE. These elevations usually decrease as the disease improves with treatment. Urinalysis is important to screen and monitor for lupus nephritis. The current recommendation for protein assessment is to use a 12- or 24-hour urine collection to calculate the protein/creatinine ratio. Hematuria and pyuria also may occur. If lupus nephritis is suspected, a renal biopsy should be conducted to confirm diagnosis and determine the level of disease activity.

Fibromyalgia

Fibromyalgia is a common syndrome associated with chronic widespread pain, fatigue, sleep disturbances, and other medical problems lasting for at least 3 months without another medical diagnostic explanation.31 It is a challenging syndrome to categorize and treat. It may be comorbid with other conditions, thereby making the differential diagnosis challenging.

Laboratory testing should be used prudently when evaluating patients with clinical features suggestive of fibromyalgia. A satisfactory patient assessment is usually obtained by a careful medical history and physical examination and perhaps performance of routine laboratory tests, such as CBC and serum chemistry, to rule out other disorders. Serologic tests such as ANA titers are not usually necessary unless there is strong evidence of an autoimmune disorder. Once a diagnosis is made, there is no evidence that repeatedly checking laboratory testing provides value.

If the results of laboratory testing suggest a diagnosis other than fibromyalgia, a more directed evaluation is required. Individuals who actually have fibromyalgia are sometimes misdiagnosed with autoimmune disorders. This may be due to the common complaints of arthralgias, myalgias, fatigue, morning joint stiffness, and a history of swelling of the hands and feet. Conversely, patients with existing autoimmune diseases may suffer from symptoms suggestive of fibromyalgia.

TESTS TO MONITOR DRUG THERAPY FOR SELECT RHEUMATIC DISORDERS

Pharmacotherapy for rheumatic diseases can cause significant adverse reactions that are reflected in laboratory test results.32-34 Abnormal test results may necessitate dose reduction, temporary discontinuation, or permanent withdrawal of the offending drug. The laboratory tests most commonly affected are the WBC count, platelet count, hepatic aminotransferases, total bilirubin, SCr, BUN, and urinalysis (Table 20-5). An important part of a patient care plan is regular evaluation of the associated laboratory tests to ensure safety of the medication regimen. In addition, patients should be counseled to report any signs or symptoms of adverse reactions so the healthcare provider can determine if they are related to the medication regimen.

TABLE 20-5.

Routine Laboratory Tests to Monitor Patients Receiving Select Drugs for Common Rheumatic Diseases

DRUG

DISEASE

LABORATORY TEST

ADVERSE DRUG REACTION

Abatacept

RA

Baseline tuberculin skin test

CBC with differential and platelet count

Monitor for infection

Infection, sepsis

Allopurinol

Gout and hyperuricemia

CBC with differential and platelet count

Serum uric acid

Hepatic aminotransferases

BUN, SCr

Myelosuppression

Gout flare (monitor uric acid)

Hepatotoxicity

Nephrotoxicity

Anakinra

RA

CBC with differential and platelet count

Monitor for infection

Neutropenia

Infection, sepsis

Belimumab

SLE

Monitor for infection

Infection, sepsis

Colchicine

Gout and hyperuricemia

CBC with differential and platelet count

Hepatic aminotransferases

BUN, SCr

Myelosuppression

Elevated aminotransferases

Corticosteroids

RA, OA, SLE, gout flares

CBC with differential and platelet count

Serum sodium, potassium, bicarbonate

Serum calcium

Blood glucose

Fasting lipid panel

Urinalysis

Anemia due to peptic ulceration and blood loss

Electrolyte disturbances

Osteoporosis

Hyperglycemia

Dyslipidemia

Glycosuria

Febuxostat

Gout and hyperuricemia

Serum uric acid

Hepatic aminotransferases

Gout flare (monitor uric acid)

Elevated aminotransferases

Hydroxychloroquine

RA

CBC with differential and platelet count

Urinalysis

Thrombocytopenia

Proteinuria

JAK inhibitors

(baricitinib, tofacitinib, upadacitinib)

RA

Baseline tuberculin skin test

CBC with differential and platelet count

Monitor for infection

Fasting lipid panel

Infection, sepsis

Hepatic enzyme elevations

Dyslipidemia

Leflunomide

RA

CBC with differential and platelet count

Hepatic aminotransferases, bilirubin

Pancytopenia

Elevated aminotransferases

Hepatic necrosis

Lesinurad

Gout and hyperuricemia

Serum uric acid

BUN, SCr

Gout flare (monitor uric acid)

Methotrexate

RA

CBC with differential and platelet count

Hepatic aminotransferases, bilirubin, serum albumin

Monitor for infection

Leukopenia

Pancytopenia

Hepatotoxicity

Infection, sepsis

Mycophenolate mofetil

SLE

CBC with differential and platelet count

Monitor for infection

Neutropenia, red cell aplasia

Opportunistic infections, sepsis

NSAIDs, including aspirin

RA, OA, SLE, gout flares

CBC with differential and platelet count

Hepatic aminotransferases

BUN, SCr

Sodium, potassium

Urinalysis

Anemia (due to gastroduodenal ulceration and blood loss)

Hepatotoxicity

Nephrotoxicity

Electrolyte disturbances

Proteinuria, hematuria, pyuria

Pegloticase

Gout and hyperuricemia

Serum uric acid

Gout flare (monitor uric acid)

Probenecid

Gout and hyperuricemia

Serum uric acid

BUN, SCr

Gout flare (monitor uric acid)

Rituximab

RA

CBC with differential and platelet count

Monitor for infection

Infection, sepsis

Sulfasalazine

RA

CBC with differential and platelet count

Hepatic aminotransferases

Leukopenia

Hepatotoxicity

TNF antagonists

(adalimumab, certolizumab, etanercept, golimumab, infliximab)

RA

Baseline tuberculin skin test

CBC with differential and platelet count

Monitor for infection

Activation of tuberculosis

Infection, sepsis

Tocilizumab

RA

CBC with differential and platelet count

Monitor for infection

Hepatic transaminases

Fasting lipid panel

Infection, sepsis

Hepatic enzyme elevations

Dyslipidemia

A common approach in treatment of rheumatic diseases is “treat to target.” Once a patient is diagnosed with a rheumatic condition and a treatment regimen is determined, a timeframe for follow-up (eg, 1 month) should be established. This timeframe should also consider the time to see evidence of efficacy from the medication regimen. At the point of follow-up, the current disease activity should be evaluated. If the patient has moderate or high disease activity, the medication regimen should be adjusted to bring the disease activity under control. The target for most conditions is remission or, at a minimum, low disease activity. The medication regimen should be continually reevaluated and adjusted if the patient has not yet achieved the target of treatment.

For example, if a patient with RA is started on a regimen of methotrexate and etanercept, prior to initiating therapy, the following lab tests should be conducted: CBC with differential and platelet count, hepatic transferases, bilirubin, and serum albumin. A tuberculin skin test should be placed. Assuming all tests come back normal, the healthcare provider should follow up with the patient in 1 month to assess safety (repeat lab tests) and efficacy (assess disease activity) to determine if the treatment regimen of methotrexate and etanercept is still appropriate or if adjustment should be made. The patient should be counseled upon initiation of the medications to monitor for signs and symptoms of infection and contact the healthcare provider if they occur.

TESTS TO GUIDE MANAGEMENT OF GOUT AND HYPERURICEMIA

The serum uric acid and urine uric acid concentrations are the two most commonly used tests to diagnose gout and assess the effectiveness of its treatment. The BUN and SCr also should be monitored as appropriate.

Serum Uric Acid

Reference range: 4 to 8.5 mg/dL (237 to 506 μmol/L) for males >17 years old; 2.7 to 7.3 mg/dL (161 to 434 μmol/L) for females >17 years old

Uric acid is the metabolic end-product of the purine bases of DNA. In humans, uric acid is not metabolized further and is eliminated unchanged by renal excretion. It is completely filtered at the renal glomerulus and is almost completely reabsorbed. Most excreted uric acid (80% to 86%) is the result of active tubular secretion at the distal end of the proximal convoluted tubule.35-37

As urine becomes more alkaline, more uric acid is excreted because the percentage of ionized uric acid molecules increases. Conversely, reabsorption of uric acid within the proximal tubule is enhanced and uric acid excretion is suppressed as urine becomes more acidic.

In plasma at normal body temperature, the physicochemical saturation concentration for urate is 7 mg/dL. However, plasma can become supersaturated, with the concentration exceeding 12 mg/dL. In nongouty subjects with normal renal function, urine uric acid excretion abruptly increases when the serum uric acid concentration approaches or exceeds 11 mg/dL. At this concentration, urine uric acid excretion usually exceeds 1,000 mg/24 hr.

Hyperuricemia

When serum uric acid exceeds the upper limit of the reference range, the biochemical diagnosis of hyperuricemia can be made. Hyperuricemia can result from an overproduction of purines and reduced renal clearance of uric acid. When specific factors affecting the normal disposition of uric acid cannot be identified, the problem is diagnosed as primary hyperuricemia. When specific factors can be identified (eg, another disease or drug therapy), the problem is referred to as secondary hyperuricemia.

As the serum urate concentration increases above the upper limit of the reference range, the risk of developing clinical signs and symptoms of gouty arthritis, renal stones, uric acid nephropathy, and subcutaneous tophaceous deposits increases. However, many hyperuricemic patients are asymptomatic. If a patient is hyperuricemic, it is important to determine if there are potential causes of false laboratory test elevation and contributing extrinsic factors. In general, clinical studies have not shown that impaired renal function is caused by chronic hyperuricemia (unless there are other renal risk factors and excluding acute uric acid nephropathy resulting from tumor lysis syndrome). However, long-term, high serum uric acid levels (eg, ≥13 mg/dL in men or 10 mg/dL in women) may predispose individuals to renal dysfunction. This level of hyperuricemia is uncommon, and a conclusive link to renal insufficiency has not been established. Recent studies suggest reducing serum urate levels may slow progression of renal failure, risk of myocardial infarction, and improve insulin resistance. Further exploration is needed to fully appreciate the effects of hyperuricemia independent of a gout diagnosis.

Exogenous causes

Medications are the most common exogenous causes of hyperuricemia. The two primary mechanisms whereby drugs increase serum uric acid concentrations are (1) decreased renal excretion resulting from drug-induced renal dysfunction or competition with uric acid for secretion within the kidney tubules and (2) rapid destruction of large numbers of cells from antineoplastic therapy for leukemias and lymphomas.

The reduction in glomerular filtration rate accompanying renal impairment decreases the filtered load of uric acid and causes hyperuricemia. Several drugs cause hyperuricemia by renal mechanisms that may include interference with renal clearance of uric acid. These agents include low-dose aspirin, pyrazinamide, nicotinic acid, ethambutol, ethanol, cyclosporine, acetazolamide, hydralazine, ethacrynic acid, furosemide, and thiazide diuretics. Diuretic-induced volume depletion results in enhanced tubular reabsorption of uric acid and a decreased filtered load of uric acid. Salicylates, including aspirin, taken in low doses (1 to 2 g/day) may decrease urate renal excretion. Moderate doses (2 to 3 g/day) usually do not alter urate excretion. Large doses (>3 g/day) generally increase urate renal excretion, thereby lowering serum urate concentrations.

Many cancer chemotherapeutic agents (eg, methotrexate, nitrogen mustards, vincristine, 6-mercaptopurine, and azathioprine) increase the turnover rate of nucleic acids and the production of uric acid. Drug-induced hyperuricemia after cancer chemotherapy, especially high-dose regimens, can lead to acute renal failure. Allopurinol is routinely administered prophylactically to decrease uric acid formation. In other clinical situations, drug-induced hyperuricemia may not be clinically significant.

The decision to continue or discontinue a drug that may be causing hyperuricemia depends on three factors: (1) the risk of precipitating gouty symptoms, based on the patient’s past history and current clinical status, (2) the feasibility of substituting another drug that is less likely to affect uric acid disposition, and (3) the plausibility of temporarily or permanently discontinuing the drug. If the regimen of the causative drug must remain unchanged, pharmacologic treatment of hyperuricemia may be instituted.

Diet is another exogenous cause of hyperuricemia. High-protein, weight-reduction programs can greatly increase the amount of ingested purines and subsequent uric acid production. If the average daily diet contains a high proportion of meats, the excess nucleoprotein intake can lead to increased uric acid production. Fasting or starvation also can cause hyperuricemia because of increased muscle catabolism. Furthermore, lead poisoning from paint, batteries, or “moonshine,” in addition to recent alcohol ingestion, obesity, diabetes mellitus, and hypertriglyceridemia, are associated with increases in serum uric acid concentration (Minicase 3).

Endogenous causes

Endogenous causes of hyperuricemia include diseases, abnormal physiologic conditions that may or may not be disease related, and genetic abnormalities. Diseases include (1) renal diseases (eg, renal failure), (2) disorders associated with increased destruction of nucleoproteins (eg, leukemia, lymphoma, polycythemia, hemolytic anemia, sickle cell anemia, toxemia of pregnancy, and psoriasis), and (3) endocrine abnormalities (eg, hypothyroidism, hypoparathyroidism, pseudohypoparathyroidism, nephrogenic diabetes insipidus, and Addison disease).

Predisposing abnormal physiologic conditions include shock, hypoxia, lactic acidosis, diabetic ketoacidosis, alcoholic ketosis, and strenuous muscular exercise. In addition, men and women are at risk for developing asymptomatic hyperuricemia at puberty and menopause, respectively. Genetic abnormalities include Lesch-Nyhan syndrome, gout with partial absence of the enzyme hypoxanthine guanine phosphoribosyltransferase, increased phosphoribosyl pyrophosphate P-ribose-PP synthetase, and glycogen storage disease type I.

Hypouricemia

Hypouricemia is not important pathophysiologically, but it may be associated with low-protein diets, renal tubular defects, xanthine oxidase deficiency, and drugs (eg, high-dose aspirin, allopurinol, probenecid, and megadose vitamin C).

Assays and Interferences with Serum Uric Acid Measurements

In the laboratory, the concentration of uric acid is measured by either the phosphotungstate colorimetric method or the more specific uricase method. With the colorimetric method, ascorbic acid, caffeine, theophylline, levodopa, propylthiouracil, and methyldopa can all falsely elevate uric acid concentrations. With the uricase method, purines and total bilirubin >10 mg/dL can cause a false depression of uric acid concentrations. False elevations may occur if ascorbic acid concentrations exceed 5 mg/dL or if plasma hemoglobin exceeds 300 mg/dL (in hemolysis).

Urine Uric Acid Concentration

Reference range: 250 to 750 mg/24 hr (1.48 to 4.46 mmol/24 hr)

In hyperuricemic individuals who excrete an abnormal amount of uric acid in the urine (hyperuricaciduria), the risk of uric acid and calcium oxalate nephrolithiasis increases. However, the prevalence of stone formation is only twice that observed in the normouricemic population. When a stone does form, it rarely produces serious complications. Furthermore, treatment can reverse stone disease related to hyperuricemia and hyperuricaciduria.

Hyperuricemia and Gout

A 45-year-old obese man began a daily exercise program 2 weeks ago in an attempt to lose 50 lb. In addition, he has begun a high-protein liquid diet because he knows that “fatty foods are not healthy.” He sees his family physician for his first complete physical examination in approximately 7 years. He tells his physician he has recently started walking briskly for 1 hour three times a week and is watching his diet carefully. The only abnormal finding on physical examination is a BP of 150/95 mm Hg. After drawing blood for a CBC with differential and a full chemistry panel and obtaining a urine sample for urinalysis, the physician prescribes hydrochlorothiazide 25 mg once daily for hypertension. Three days later, he is notified that his laboratory results, including serum uric acid, are normal.

Two weeks later, he returns on crutches to see his physician. He explains that he injured his right foot 3 days prior when taking his daily walk before sunrise, and he accidentally stubbed his right foot on a rock. He also says he woke up 2 days prior with a fever and felt as if his right great toe was “in a vise while an ice-cold knife was being pushed into the joint.”

Examination of his right foot reveals abrasions on all five toes. The skin of the great toe appears shiny, the toe is swollen and warm to the touch, and he is in obvious pain. Whitish fluid oozes from a small wound on the dorsal aspect of the great toe. After anesthetizing the joint, the physician aspirates several drops of the whitish fluid. The physician then performs a Gram stain and examines the fluid on a slide, finding needle-shaped crystals but no bacteria. He also orders a serum uric acid level.

QUESTION: What is this patient’s likely diagnosis and prognosis?

DISCUSSION: He probably has experienced his first acute gout attack. Although his previous serum uric acid concentration was described as normal, one endogenous and two exogenous factors may have precipitated this attack. Hypertension is frequently associated with hyperuricemia. Also, the sudden change to a high-protein diet greatly increased his ingestion of purines, which are metabolized to uric acid. Finally, he also was started on hydrochlorothiazide, which is an inhibitor of the renal clearance of uric acid. The abrupt change in physical exertion probably did not contribute to the attack because it was low in intensity.

Although he attributes the condition to his traumatic toe-stubbing event, his physician notes that none of his other abraded toes appear to be “infected.” Examination of the synovial fluid using a polarizing-light microscope reveals monosodium urate crystals without bacteria (Table 20-4). An elevated serum uric acid level would be consistent with a diagnosis of gout, but some patients can have acute gout attacks with a serum urate level that is within the reference range.

With appropriate treatment for acute gout, discontinuation of hydrochlorothiazide, and adequate follow-up monitoring, the patient’s symptoms should improve substantially within 24 to 48 hours. Some patients never have a second gout attack, whereas others experience frequent and severe episodes. If the serum uric acid concentration that was ordered is reported as highly elevated (>10 mg/dL), initiating therapy to reduce hyperuricemia may be considered after resolution of the acute attack. According to the 2020 ACR guidelines for treating hyperuricemia, patients should be evaluated on a case-by-case basis to determine causes for elevated uric acid and need for urate-lowering therapy.38 Any patient with frequent attacks (defined as two or more per year), tophi, radiographic damage due to gout, chronic kidney disease stage 3 or greater, or a history of urolithiasis should receive urate-lowering therapy.38 The results of a 24-hour urine collection would be useful in determining whether the patient is an overproducer or underexcretor of uric acid and help to guide therapy with either allopurinol or probenecid, respectively.

Pathologically, uric acid nephropathy—a form of acute renal failure—is a direct result of uric acid precipitation in the lumen of collecting ducts and ureters. Uric acid nephropathy most commonly occurs in two clinical situations: (1) patients with marked overproduction of uric acid secondary to chemotherapy-induced tumor lysis (leukemia or lymphoma) and (2) patients with gout and profound hyperuricaciduria. Uric acid nephropathy also has developed after strenuous exercise or convulsions.

In hyperuricemia unrelated to increased uric acid production, quantification of urine uric acid excreted in 24 hours can help to direct prophylaxis or treatment. Patients at higher risk of developing renal calculi or uric acid nephropathy (patients with gout or malignancies) excrete ≥1,100 mg of uric acid per 24 hours. Prophylaxis may be recommended for these patients; allopurinol should be used instead of uricosuric agents (eg, probenecid) to minimize the risk of nephrolithiasis. Prophylactic therapy may be started at the onset of gout-like symptoms.

SUMMARY

Diagnosing and managing rheumatic diseases rely heavily on a thorough medical history and physical exam. Clinicians can use laboratory tests to help confirm or rule out specific diagnoses. Every laboratory test ordered must be carefully evaluated to determine the next steps in an individual patient’s care.

When used alone, no single test is diagnostic for any particular disease. However, positive RF, ACPA, and ANA tests are commonly observed in patients with RA and SLE, respectively. The cANCA antibody is highly specific for the disease spectrum of GPA, and anti-MPO antibodies are highly specific for systemic vasculitis and idiopathic crescentic glomerulonephritis. The most complete screen of complement activation includes measurements of C3, C4, and CH50. The Westergren ESR and CRP tests are nonspecific markers of systemic inflammation and must be interpreted in light of the clinical presentation and other laboratory tests.

Patients with hyperuricemia are usually asymptomatic. After treatment of an episode of acute gout, the decision to initiate antihyperuricemic therapy depends on the frequency and severity of acute attacks. Allopurinol or febuxostat therapy is recommended for patients at risk for forming renal calculi.

LEARNING POINTS

1. How are laboratory tests used in patients with RA and OA?

ANSWER: A carefully collected history and physical can help to distinguish between RA and OA in a patient who reports joint pain. Identifying which joints are affected and inquiring about morning joint stiffness and the duration of time necessary to resume normal function of the affected joint provide important information. Laboratory tests can be conducted once the history and physical are completed to rule in or rule out diagnoses. ACPA and RF, if positive, can rule in a diagnosis of RA with 99.5% specificity. ESR and CRP, if positive, indicate the presence of systemic inflammation, though they are not specific to RA. If ACPA, RF, ESR, and CRP are negative or normal, a diagnosis of OA is more likely.

2. How important are the sensitivity and specificity of laboratory tests for diagnosing or assessing rheumatic diseases?

ANSWER: With many rheumatic diseases, significant information can be gathered through a patient’s medical history and physical exam to narrow down the potential causes of the patient’s signs and symptoms. Then and only then should lab tests be considered to rule in or rule out specific diseases. Lab tests should not be ordered arbitrarily. Understanding the sensitivity and specificity of the tests available is critical to understanding their association with diagnosis. For example, if a new test to diagnose RA is only 75% sensitive, this means that 25% of patients who actually have the disease will show a negative result. If a new test to diagnose RA is only 75% specific, 25% of people tested who have a positive result do not actually have RA. For some laboratory tests, poor specificity is due to substances not associated with the disease that cross-react with the target compound.

3. What issues should be considered before ordering a laboratory test for a patient with a rheumatic disorder?

ANSWER: Laboratory testing can be expensive and may be inconvenient to perform. Consequently, several questions should be posed before ordering another test.

  1. Are the results of other tests already available that provide the same information?

  2. If a test was performed previously, are there important reasons to repeat the test now?

  3. Has enough time elapsed since the previous test to make new results meaningful?

  4. Will the results of this test change the diagnosis, prognosis, or therapeutic interventions I might make? In other words, will knowing this result change what I do?

  5. Are the benefits to the patient worth the possible discomfort, inconvenience, and extra cost?

The results of laboratory tests should always be interpreted in light of the clinical picture (ie, the patient’s signs and symptoms).

REFERENCES