THE VALUE OF ISOTYPE DETERMINATION OF SERUM ANTIBODIES AGAINST CHLAMYDIA FOR THE DIAGNOSIS OF CHLAMYDIA REACTIVE ARTHRITIS

In clinical rheumatology, the diagnosis of Chlamydia reactivearthritis is difficult because an incomplete form of the diseasecan closely resemble an undifferentiated seronegative mono/oligoarthritis.We investigated whether measuring specific isotypes of anti-Chlamydiaantibodies in serum can improve the diagnosis, by comparingsuch antibody concentrations in the serum of patients with well-defineddisease, i.e. Chlamydia trachomatis sexually acquired reactivearthritis (CT-SARA), with other arthritides. Antibody levelswere determined by enzyme-linked immunosorbent assay (ELISA).When considering two different isotypes and their combination,the best sensitivity (63%) was obtained for IgM and/or IgA resultswith a specificity of 81%. The patients with CT-SARA and SARAhad the highest levels of antibodies of all isotypes tested.It is concluded that, in our experimental conditions, only veryhigh values of specific isotypes could indicate a diagnosisof Chlamydia reactive arthritis. KEY WORDS: Sexually acquired reactive arthritis, Chlamydia, Antibodies, Enzyme-linked immunoassay     

THE HUMORAL IMMUNE RESPONSE TO CHLAMYDIA TRACHOMATIS IN PATIENTS WITH ACUTE REACTIVE ARTHRITIS

Sera from 25 patients with clinical signs of reactive arthritiswere analysed for antibodies against Chlamydia trachomatis byimmunoblotting. Purified elementary bodies, purified Chlamydiaouter membrane complexes, and purified recombinant subcomponentswere used as antigens. Antibodies against C. trachomatis cysteinerich outer membrane protein 2 (Omp2) and lipopolysaccharide(LPS) were detected in 10 patients. Thus 40% of the patientspresented antibodies specific for C.trachomatis. There was nocorrelation between acute reactive arthritis and antibodiesto heat-shock proteins GroEL, GroES and DnaK. KEY WORDS: Chlamydia trachomatis, DnaK, GroEL, Omp2, Lipopolysaccharide, Humoral immune response     

SALMONELLA REACTIVE ARTHRITIS: SERUM AND SECRETORY ANTIBODIES IN EIGHT PATIENTS IDENTIFIED AFTER A LARGE OUTBREAK

Serial measurements of serum and secretory antibodies to Salmonellatyphimurium were made by ELISA in eight patients with suspectedreactive arthritis identified after a large outbreak of Salmonellagastroenteritis. All three patients from whom Salmonella hadbeen isolated developed significant serum IgG, IgA and IgM antibodyresponses. Only one of the three possessed HLA-B27. A furtherthree patients, two with HLA-B27, had raised antibodies, althoughnone had experienced gastroenteritis. Salmonella infection wasnot confirmed in the remaining two patients. The three B27-negativepatients with confirmed reactive arthritis had HLA-B locus antigenswhich serologically cross-react with B27. One of six patientswith confirmed reactive arthritis was under the age of 25 yearswhereas 256 of 418 (61%) patients with uncomplicated enteritiswere under this age. The development of reactive arthritis mayfollow subclinical Salmonella infection and is influenced bygenetic and age-related factors. KEY WORDS: Arthritis, Salmonella, Antibodies     

ONCOGENE EXPRESSION IN SYNOVIAL FLUID CELLS IN REACTIVE AND EARLY RHEUMATOID ARTHRITIS: A BRIEF REPORT

Since it has been implied that cellular oncogenes might havea role in the pathogenesis of rheumatoid arthritis (RA), wehave examined the expression of c-myc, c-myb, c-fos, c-jun andc-Ha-ras oncogenes in the cells from synovial fluid (SF) andperipheral blood (PB) of patients with reactive arthritis (RcA)and early RA. Oncogene expression was studied using RNA hybridizationswith ³²P-labelled probes. From the SF, mononuclear and granulocytecell fractions were used separately. Significant differencesbetween ReA and RA were observed only for c-myb in PB mononuclearcells and c-jun in SF granulocytes. Regarding the expressionof c-myc, c-fos and c-Ha-ras oncogenes, no difference betweenReA and RA was observed. Comparison to normal controls was madeusing PB mononuclear cells; only the expression of c-fos tendedto be slightly increased in RA, without statistical significance,however. We conclude that oncogene activation in the synovialinflammation is not a phenomenon specific for RA. KEY WORDS: Oncogenes, Rheumatoid arthritis, Reactive arthritis     

Amplification of plasmid and chromosome chlamydia dna in synovial fluid of patients with reactive arthritis and undifferentiated seronegative oligoarthropathies

Objective. To investigate the hypothesis that whole bacteria might be found in the joints of patients with Chlamydia-associated reactive arthritis. Methods. The presence of 2 plasmid- and 2 chromosome-specific sequences of Chlamydia DNA was investigated by amplification with the polymerase chain reaction, in synovial fluid (SF) samples from 71 patients with various arthropathies. Results. Chlamydia DNA was found in SF samples from 22 patients. Conclusion. Whole chlamydiae are likely present in the SF of patients with Chlamydia-associated reactive arthritis.     

A SEARCH FOR CHLAMYDIA TRACHOMATIS IN SYNOVIAL FLUIDS FROM PATIENTS WITH REACTIVE ARTHRITIS USING THE POLYMERASE CHAIN REACTION AND ANTIGEN DETECTION METHODS

A polymerase chain reaction (PCR) technique to detect Chlamydiatrachomatis DNA was used to examine synovial specimens frompatients with reactive arthritis. We were able to detect C.trachomatis DNA in synovial specimens which had been seededwith intact elementary bodies or chlamydial DNA. However, wewere unable to detect chlamydial DNA in unseeded synovial specimensfrom 10 patients with sexually acquired reactive arthritis,17 patients with reactive arthritis and 11 control patientswith other arthropathies. In addition, using a monoclonal antibodytechnique, we were unable to detect chlamydial antigen in anyof the synovial cell deposits examined. We conclude that C.trachomatis DNA was not present in the joints of these patientsat the time of synovial fluid collection, and suggest that eitherDNA degradation occurred rapidly after viable chlamydiae hadentered the joint or that chlamydial DNA was not present atany stage of the reactive response. KEY WORDS: Joint disease, Sexually acquired reactive arthritis, Oligoarthritis, DNA hybridization     

EXPRESSION OF bcl-2 IN RHEUMATOID ARTHRITIS

Since defective apoptosis has been suggested to play a rolein the development of autoimmune diseases, we have investigatedthe expression of the proto-oncogene bcl-2 in patients withrheumatoid arthritis (RA). The expression of bcl-2 was studiedin peripheral blood (PB) and synovial fluid (SF) lymphocytesand synovial tissues (ST) from patients with RA using immunohistochemistry,flow cytometry and nucleic acid hybridization. Patients withreactive arthritis (ReA) or osteoarthritis (OA) and healthyindividuals were used as controls. The expression of bcl-2 proteinin PB lymphocytes and the expression of bcl-2 mRNA in PB mononuclearcells (PBMC) was similar in healthy controls and patients withRA. However, bcl-2 protein expression was significantly reducedin SF lymphocytes when compared to PB lymphocytes. Similar resultswere observed with lymphocytes from patients with ReA, and irrespectiveof whether total lymphocytes, T cells or different T-cell subsetswere studied. In the synovial sections, the expression of bcl-2was restricted to lymphocytes, and bcl-2+ cells were observedin the majority of samples from patients with RA, OA and ReA.These data indicate that the expression of bcl-2 is not increasedin the lymphocytes or ST derived from patients with RA. Instead,decreased expression of bcl-2 protein in SF lymphocytes comparedto PB lymphocytes was demonstrated. We suggest that bcl-2 doesnot play a significant role in the pathogenesis of RA. KEY WORDS: bcl-2, Rheumatoid athritis, Lymphocytes     

DETECTION OF MYCOBACTERIUM TUBERCULOSIS ANTIGEN IN SYNOVIAL FLUID OF PATIENTS WITH RHEUMATOID ARTHRITIS

By use of antimycobacterial saline extract antibodies, Mycobacteriumtuberculosis (MT) antigens have been detected in synovial fluid(SF) of patients with rheumatoid arthritis (RA) by a highlyspecific and sensitive double-antibody sandwich enzyme-linkedimmunosorbent assay (ELISA). Absorbance for 15 gout, 14 osteoarthritis(OA) patients, 12 patients with spondylarthropathies (SA) andeight patients with other inflammatory disorders (OID) rangedfrom 0.001 to 0.025 with the mean values of 0.0086 ±0.0078, 0.0077 ± 0.0051, 0.0069 ± 0.0059 and 0.0113± 0.0059, respectively. Among 65 SF of patients withRA examined, 34 were found to be negative for MT antigen withabsorbance ranging from 0.002 to 0.024 and a mean value of 0.0114± 0.0070. For 31 (47.7%) MT antigen-positive specimensof RA, optical density ranged from 0.052 to 2.446 with a meanvalue of 0.5564 ± 0.7354. Significant statistical difference(P < 0.05) was found when MT antigen positives were comparedwith MT antigen negatives, gout, OA, SA and OID groups. Ourresults indicate that MT may be relevant to the pathogenesisof RA. KEY WORDS: MT antigen and RA, Synovial fluid, Antimycobacterial saline extract antibody, ELISA     

Avidity of anti-yersinia antibodies in yersiniosis patients with and without yersinia-triggered reactive arthritis

Avidity of IgM, IgG, and IgA class anti-Yersinia antibodies was compared in sera of 22 patients with yersiniosis and subsequent reactive arthritis versus sera of 22 patients without postinfection complications. An enzyme-linked immunosorbent assay was used for antibody determination. Less than 2 months after onset of the infection, the patients with arthritis had fewer high-avidity IgM antibodies against the bacterial lipopolysaccharide (P = 0.035) and more high-avidity IgA antibodies against bacterial cell extract (P = 0.039) than did the patients without arthritis. This difference increased with time.     

Alteration of chlamydia trachomatis biologic behavior in synovial membranes suppression of surface antigen production in reactive arthritis and reiter's syndrome

Objective. To investigate the biologic state of Chlamydia and its surface antigen expression in the synovial membranes of patients with Chlamydia-associated reactive arthritis/Reiter's syndrome (ReA/RS). Methods. Expression of chlamydial lipopoly-saccharide (LPS), major outer membrane protein (MOMP), and elementary body (EB) antigens was studied by gold labeling immunoelectron microscopy on 6 synovial membrane and 2 synovial fluid (SF) pellet samples from 6 patients with Chlamydia-associated arthritis. The study findings were compared with 24-hour cultures of HeLa cells infected with Chlamydia trachomatis EB. Results. Persistent C trachomatis infection was found in all 6 synovial membrane samples from patients who had either early or chronic arthritis. The infection persisted despite antibiotic treatment, including a 1-month course of doxycycline therapy. Most persistent organisms were atypical reticulate bodies (RBs) found in both fibroblasts and macrophages. Specific, but weak, immunogold staining for all 3 antibodies was found on both intracellular RBs and extracellular EBs. In the SF samples, Chlamydia surface antigens were detected only in phagosomes containing degraded electron-dense materials. Conclusion. The synovial membrane biopsies conducted in this study of Chlamydia-associated ReA/RS revealed atypical RBs with diminished MOMP and LPS expression. Such altered organisms may escape immune surveillance and contribute to disease chronicity; moreover, these organisms may be difficult to detect and treat in some ReA/RS patients.     

ARTHRITIS IN SCLERODERMA

Thirty-four patients (24 females and 10 males) selected from300 consecutive patients with established systemic sclerosis(SSc), with a current or past history of articular symptoms,were clinically documented and further studied using thermographyand bone scan to define the pattern of arthritis. Clinical evidenceof synovitis was observed in 30 (88%) and joint inflammationwas detected in 31 (91%) by the above-mentioned imaging techniques.A distinctive subset of 10 patients with deforming arthritiswas characterized in which seven (70%) patients fulfilled criteriafor both rheumatoid arthritis and SSC; three of these satisfiedthe criteria for diagnosis of CREST, but none met the criteriaof mixed connective tissue disease. These patients, as a group,when compared with the rest showed limited skin involvement(skin score of 19 ± 11 vs 33 ± 14; P < 0.05)and were positive for rheumatoid factor (80 vs 13%; P < 0.05)and anticentromere antibodies (37 vs 4%; P < 0.05). KEY WORDS: Scleroderma, Arthritis, Thermography, Bone scan     

REACTIVE ARTHRITIS: UROGENTTAL SWAB CULTURE IS THE ONLY USEFUL DIAGNOSTIC METHOD FOR THE DETECTION OF THE ARTHRITOGENIC INFECTION IN EXTRA-ARTICULARLY ASYMPTOMATIC PATIENTS WITH UNDIFFERENTIATED OLIGOARTHRITIS

Reactive arthritis (ReA) is a scronegative oligoarthritis triggeredby a preceding extra-articular infection. While evidence ofa microbial infection is mandatory for establishing the diagnosisof ReA, the sensitivity of bacteriological and serological testshas not been determined in patients without symptoms of infection.In a retrospective study, we evaluated the usefulness of urogenitalswab cultures, serology and stool culture to identify infectionsin 234 patients with undifferentiated oligoarthritis. One hundredand forty-four patients complaining about joint pain who hadno sign or history of inflammatory arthritis served as controls.Urogenital swab cultures showed a microbial infection in 44%of the patients with oligoarthritis (15% Chlamydia, 14% Mycoplasma,28% Ureaplasma), whereas in the control group only 26% had apositive result (4% Chlamydia, 7% Mycoplasma, 21% Ureaplasma)(P < 0.001). A Chlamydia IgG-antibody titre     

Yersinia-specific antibodies in serum and synovial fluid in patients with Yersinia triggered reactive arthritis.

OBJECTIVES--To further evaluate the role of bacterial antigens in triggering inflammation in the joint in patients with reactive arthritis by studying local antibody synthesis in the joint. METHODS--Yersinia-specific antibodies in paired serum and synovial fluid samples from 29 patients with yersinia triggered reactive arthritis were studied using an enzyme linked immunosorbent assay (ELISA), an inhibition ELISA with six monoclonal antibodies against lipopolysaccharide or released proteins of yersinia and immunoblotting. Antibodies of IgM, IgG and IgA classes, as well as antibodies of IgA subclasses and those containing secretory component were measured against the lipopolysaccharide and the sodium dodecyl sulphate extract of whole Yersinia enterocolitica O:3 bacteria. RESULTS--It was shown that yersinia-specific antibodies, as well as antibodies against other microbial antigens (rubella, measles, Bordetella pertussis, tetanus toxoid and Candida albicans) in synovial fluid mirror those in serum by concentration, by specificity and by distribution in classes and subclasses. CONCLUSION--These results do not suggest any strong local antibody production, but indicate that the majority of yersinia antibodies in the synovial fluid are derived from the circulation.     

ANTI-PROTEUS ANTIBODIES AND PROTEUS ORGANISMS IN RHEUMATOID ARTHRITIS: A CLINICAL STUDY

We have studied anti-Proteus antibodies (APA), isolation ofProteus, and their relation to various measures of RA diseaseactivity. Seventy RA patients with a CRP>10 mg/l had higherAPA titres than 17 RA patients with CRP     

THE EFFECT OF NON-STEROIDAL ANTI-INFLAMMATORY DRUGS ON FAECAL FLORA AND BACTERIAL ANTIBODY LEVELS IN RHEUMATOID ARTHRITIS

The faecal flora and bacterial antibody levels of 22 patientswith active rheumatoid arthritis (RA) were compared with thoseof 26 patients with osteoarthritis (OA) undergoing comparabletreatment with non-steroidal anti-inflammatory drugs (NSAIDs),and a further 22 patients with OA who were not receiving NSAIDs.Faecal counts of Clostridium perfringens were significantlyhigher in the RA patient group and in those OA patients receivingNSAIDs, compared with those OA patients not taking NSAIDs (P=0.032,P=0.0004 respectively). Total aerobic and anaerobic counts were,however, identical in all three groups. Levels of serum IgA antibody to the alpha toxin of Cl. perfringenswere higher in the RA group and in the OA group taking NSAIDsthan in OA patients not taking NSAIDs (P=0.011, P=0.055). SerumIgG antibody to alpha toxin was higher in the RA group thanin OA patients both on and off NSAIDs (P=0.019, P=0.0072) andalso a group of normal controls (P=0.032). These results suggest that the increased faecal counts of Cl.perfringens together with the associated increased antibodylevels seen in this and previous studies are more likely toresult from NSAID therapy used to treat the disease than froma disease specific changk in bowel flora. KEY WORDS: Rheumatoid arthritis, Cl. perfringens, NSAIDs     

Correlation between <Emphasis Type="Italic">Yersinia enterocolitica</Emphasis> and type I collagen reactivity in patients with arthropathies

We investigated the association with Yersinia infection in patients with arthropathies in our region. To assess the reactivity to articular antigens, the correlation of anti-Yersinia with anti-type I and type II collagen antibodies was studied. Sera from 124 patients with musculoskeletal symptoms, and 47 synovial fluids (SF) from patients with rheumatoid arthritis (RA), spondyloarthopathies (SpA) or osteoarthritis (OA) were examined. Immunoglobulins against Yersinia enterocolitica, type I and type II collagens were determined by enzyme-linked immunosorbent assay. Immunoglobulin (Ig) A to Yersinia lipopolysaccharide (LPS) was present in 13/124 sera (10%) and 3/47 SF (6%). By Western blot, IgA to Yersinia outer proteins (Yops) was found in 14/124 sera (11%) and 2/47 SF (4%). Yersinia DNA from SF was not amplified by polymerase chain reaction. We found a significant correlation with anti-collagen type I but not type II antibodies. These results suggest different reactivity to articular collagen in patients with Yersinia antibodies.     

RHEUMATOID FACTOR ASSOCIATED WITH A SECRETORY COMPONENT IN RHEUMATOID ARTHRITIS

The authors' objective was to study the serum secretory immunoglobulinA (S-IgA) concentration and the presence of rheumatoid factor(RF) complexed with a secretory component (SC) in rheumatoidarthritis (RA). Sixty-three RA patients were studied. Therewere 49 healthy subjects in the control group. The S-IgA concentrationand the presence of IgA isotype RF were determined by ELISAin the serum. The presence of SC complexed to RF (SC–RF)was studied by a sandwich-type enzyme-linked immunosorbent assaywith an antibody against the SC used to capture S-immunoglobulin,and associated anti-globulin activity was revealed with a peroxidase-conjugatedhuman IgG Fc fragment. We observed a significant increase inS-IgA in RA (mean 76.8 µg/ml ± 152.9 S.D.), ascompared to controls (mean 13.6 µg/ml± 11.9 S.D.)(P <0.01). Forty-one per cent of RA patients presented aS-IgA concentration above the upper threshold, but we did notobserve any association with disease activity. S-IgA concentrationwas correlated with the presence of IgA-RF. Twenty-seven RApatients had a positive SC–RF versus one in the controlgroup (P<0.01). The presence of SC–RF was associatedwith an increased S-IgA concentration (P <0.0001), and thepresence of RF-IgA (P <0.002). However, no association withdisease activity was noted. Our study showed that serum S-IgAwas increased in RA, and that part of the RF were complexedwith SC. These results suggest contribution of mucosal lymphocytesin the pathogenesis of RA. KEY WORDS: Secretory immunoglobulin, Rheumatoid factor, Rheumatoid arthritis, Mucosa-associated lymphoid tissue     

Anti-tissue transglutaminase antibodies in inflammatory and degenerative arthropathies

Recent studies identified tissue transglutaminase (tTG) as the antigen eliciting antiendomysial antibodies (EMA) in celiac disease (CD). Anti-tTG antibodies have therefore been proposed as a serological test for CD. Nevertheless, IgA anti-tTG but not EMA have also been found in inflammatory bowel disease patients, suggesting that these antibodies are linked to a tissue lesion rather than to an auto-immune component of CD. To confirm this hypothesis, we evaluated the presence of IgA anti-tTG in patients with inflammatory and degenerative diseases, in whom tissue lesions presented far away from the intestinal mucosa. The study was carried out on the serum and synovial fluid (SF) of 68 patients with rheumatoid arthritis (RA=33), psoriatic arthritis (PsA=26) and osteoarthritis (OA=9). In RA, PsA and OA sera, IgA anti-tTG were positive in 33%, 42% and 11% of patients, respectively. Serum anti-tTG levels were significantly higher in RA (p<0.0001), PsA (p<0.0001) and OA (p<0.02) with respect to healthy controls. SF anti-tTG levels were significantly higher in PsA (p<0.018) than in OA. A good correlation between serum and synovial fluid anti-tTG levels was found in all arthropathies This study suggests that tTG is not the only antigen of EMA and, furthermore, that IgA anti-tTG antibodies represent a general lesion-associated event. Moreover, the significant correlation between serum and synovial fluid anti-tTG levels allow us to hypothesise that these antibodies could be synthesized in the site of arthritic lesions.     

Increased levels of autoantibodies against copper-oxidized low density lipoprotein,malondialdehyde-modified low density lipoprotein and cardiolipin in patients with rheumatoid arthritis

Objectives. To analyse the association of autoantibodiesagainst cardiolipin (CL) and oxidized low density lipoproteins[copper-oxidized low density lipoprotein (oxLDL), malondialdehyde-modifiedLDL (MDA-LDL)] with rheumatoid arthritis (RA) and cardiovascularcomplications. Methods. One hundred and twenty-one patients with RA wereconsecutively included. Autoantibodies were determined by ELISA.Healthy individuals from the same region were used as controls. Results. Levels of IgG, IgM and IgA antibodies against MDA-LDLand CL, as well as IgG and IgA antibodies against oxLDL wereincreased in the patients (P<0.01). The prevalence of IgG,IgM and IgA antibodies against CL was higher than in the normalpopulation (74, 82 and 14%, respectively). The prevalenceof IgG and IgA antibodies against oxLDL was also significantlyincreased (35 and 25%, respectively) and so was the prevalenceof IgG and IgM antibodies against MDA-LDL (17 and 26%, respectively)compared with controls. The levels of IgM and IgA antibodiesagainst aCL and IgM against MDA-LDL were increased in patientswith extra-articular manifestations. Patients who developedmyocardial infarction had a higher prevalence of IgG antibodiesagainst MDA-LDL (P=0.04). There were substantial correlationsbetween the levels of antibodies against oxLDL, MDA-LDL andCL. Conclusions. RA patients had increased levels and prevalenceof autoantibodies against CL, oxLDL and MDA-LDL, with associationsto severity of disease and cardiovascular complications. KEY WORDS: Autoantibody, Rheumatoid arthritis, Cardiolipin, oxLDL, MDA-LDL, Cardiovascular disease. Correspondence to: A. K. Lefvert, Immunological Research Unit,CMM L8:03, Karolinska Hospital, 171 76 Stockholm, Sweden.     

ANTIBODIES TO FOUR GRAM-NEGATIVE BACTERIA IN RHEUMATOID ARTHRITIS WHICH SHARE SEQUENCES WITH THE RHEUMATOID ARTHRITIS SUSCEPTIBILITY MOTIF

The bacteria Proteus, Serratia, Escherichia and Pseudomonaspossess sequences resembling the rheumatoid arthritis susceptibilitysequence EQRRAA, but antibodies were elevated only to Proteusin 66 RA patients (P<0.001) when compared to 61 active ankylosingspondylitis patients and 60 controls. KEY WORDS: Rheumatoid arthritis, Proteus mirabilis, Gram-negative bacteria