Serum Epstein-Barr virus DNA, detected by droplet digital PCR, correlates with disease activity in patients with rheumatoid arthritis.

OBJECTIVES: To study the prevalence of asymptomatic activation of Epstein-Barr virus (EBV) in patients with rheumatoid arthritis (RA) and to analyse the correlation of serum EBV DNA with the disease activity. METHODS: The level of EBV DNA was determined by droplet digital PCR assay from the serum of 46 DMARD naive early RA (ERA) and 22 chronic RA (CRA)-patients at study onset. Follow-up samples from 31 ERA and 16 CRA patients were obtained after starting or modifying the anti-rheumatic treatment. EBV DNA was also measured from 33 healthy controls and 9 patients with adult onset Still's disease (AOSD). Disease activity was assessed by the disease activity score (DAS28). RESULTS: At baseline, EBV DNA was detected in the serum of 7 of the 46 ERA patients all of whom had moderate or high disease activity. In the follow-up samples, 11 of 31 patients were EBV DNA positive. At baseline EBV positive patients had significantly higher disease activity (p=0.036) and the concentration of EBV DNA correlated significantly with DAS28 (rs=0.333, p=0.024). EBV DNA was detected in 3 of 22 CRA patients at study onset and in 8 of 16 in the follow-up samples. At follow-up EBV positive patients had significantly higher DAS28 (p=0.027) and the concentration of EBV DNA correlated significantly with DAS28 (rs=0.724, p=0.002). Only one of the healthy controls and none of the AOSD patients were positive for EBV DNA. CONCLUSIONS: Active RA is associated with a lytic EBV infection which may have a role in the pathogenesis of RA.

A survey on the medication adherence to methotrexate among rheumatoid arthritis patients treated with self-administered biologic drugs.

OBJECTIVES: Methotrexate (MTX) is the most widely used co-therapy among rheumatoid arthritis (RA) patients using biological disease-modifying anti-rheumatic drugs (bDMARDs). However, adherence to MTX treatment remains a concern with estimates of adherence ranging from 59 to 63%. The objective of this study was to assess the self-reported use and adherence to MTX among RA patients treated with self-administered bDMARDs. METHODS: An electronic questionnaire survey was conducted in 68 community pharmacies in Finland. To be included in the present study patients had to be at least 18 years old, be currently using a self-administered bDMARD and be diagnosed with RA. The results are presented as medians with their respective interquartile ranges (IQR) or percentages. RESULTS: Of the 158 pharmacy customers asked to participate, 135 (85%) consented to complete the questionnaire. The included respondents were predominantly female (72%) with a median age of 55 (IQR 44-65) and rheumatic activity of 3 out of 10 (IQR 2-6.5). The majority (91%) of the included respondents were using TNF-inhibitors and 27% of all patients were on biologic monotherapy. MTX was currently used by 45% of the respondents while 50% were past users. Of the current MTX users, 6.8% identified themselves moderately non-adherent to the treatment. MTX-related adverse events were important factors associated with nonadherence and discontinuation of the treatment. CONCLUSIONS: Only 45% of the respondents were currently using MTX co-therapy, but the ones who did were adherent to their treatment. Self-reported adherence may however be subject to social desirability bias and recall bias.

Long-term real-life experience with rituximab in adult Finnish patients with rheumatoid arthritis refractory or with contraindication to anti-tumor necrosis factor drugs.

OBJECTIVE: The objective of this study was to evaluate the long-term safety and efficacy of repeated rituximab (RTX) infusions in the treatment of rheumatoid arthritis in daily clinical practice in Finland. METHODS: Data were collected from the medical records of a total of 151 patients with rheumatoid arthritis treated with RTX and followed up for at least 12 months after the treatment onset. Change in the 28-joint Disease Activity Score (DAS28), European League Against Rheumatism response criteria and proportions of patients reaching disease remission (DAS28 < 2.6) or low disease activity (DAS28 < 3.2) were used to assess the clinical response. RESULTS: Of the 151 patients 128 received 2 courses, 76 received 3 courses, and 42 received 4 courses of RTX. The mean time to retreatment for the first 4 courses varied between 11 and 13 months. Median DAS28 decreased from 5.4 (0.5-8.6) to 3.3 (0.6-6.6) after the first course. After the second treatment course, the DAS28 was 3.1 (range, 0.1-6.5). The median precourse baseline DAS28 before the second and third courses were 4.6 (range, 1.7-7.8) and 4.24 (range, 1.7-7.2), respectively. The number of previously failed tumor necrosis factor inhibitors did not predict response to RTX in this patient cohort with extensive use of previous disease-modifying antirheumatic drugs (median = 6). CONCLUSIONS: The treatment as-needed regimen used in this study cohort led to delayed RTX retreatment and disease flare in a significant proportion of patients. A regular retreatment every 6 months, at least, after the first 2 treatment courses in patients who are not in remission could allow better control of disease activity.

Diabetic myonecrosis complicated by emphysematous pyomyositis and abscess caused by Escherichia coli: a case report.

BACKGROUND: Necrotizing myopathies and muscle necrosis can be caused by immune-mediated mechanisms, drugs, ischemia, and infections, and differential diagnosis may be challenging. CASE PRESENTATION: We describe a case of diabetic myonecrosis complicated by pyomyositis and abscess caused by Escherichia coli. A white woman in her late forties was admitted to the hospital with a 1.5 week history of bilateral swelling, weakness, and mild pain of the lower extremities and inability to walk. She had a history of type 1 diabetes complicated by diabetic retinopathy, neuropathy, nephropathy, and end-stage renal disease. C-reactive protein was 203 mg/l, while creatinine kinase was only mildly elevated to 700 IU/l. Magnetic resonance imaging of her lower limb muscles showed extensive edema, and muscle biopsy was suggestive of necrotizing myopathy with mild inflammation. No myositis-associated or myositis-specific antibodies were detected. Initially, she was suspected to have seronegative immune-mediated necrotizing myopathy, but later her condition was considered to be explained better by diabetic myonecrosis with multifocal involvement. Her symptoms alleviated without any immunosuppressive treatment. After a month, she developed new-onset and more severe symptoms in her right posterior thigh. She was diagnosed with emphysematous urinary tract infection and emphysematous myositis and abscess of the right hamstring muscle. Bacterial cultures of drained pus from abscess and urine were positive for Escherichia coli. In addition to abscess drainage, she received two 3-4-week courses of intravenous antibiotics. In the discussion, we compare the symptoms and findings typically found in pyomyositis, immune-mediated necrotizing myopathy, and diabetic myonecrosis (spontaneous ischemic necrosis of skeletal muscle among people with diabetes). All of these diseases may cause muscle weakness and pain, muscle edema in imaging, and muscle necrosis. However, many differences exist in their clinical presentation, imaging, histology, and extramuscular symptoms, which can be useful in determining diagnosis. As pyomyositis often occurs in muscles with pre-existing pathologies, the ischemic muscle has likely served as a favorable breeding ground for the E. coli in our case. CONCLUSIONS: Identifying the etiology of necrotizing myopathy is a diagnostic challenge and often requires a multidisciplinary assessment of internists, pathologists, and radiologists. Moreover, the presence of two rare conditions concomitantly is possible in cases with atypical features.

Cytotoxic response persists in subjects treated for tuberculosis decades ago.

BACKGROUND: Data exploring the potential use of effector molecules produced by cytotoxic T lymphocytes (CTLs) in the immunodiagnostics of tuberculosis (TB) are scarce. The present study focused a) to gain an insight into the discriminatory power of CTLs in patients with acute pulmonary or extra-pulmonary TB, or latent tuberculosis infection (LTBI); and b) to evaluate the influence of various anti-TB therapeutic schemes on the immunological profiles of residual CTLs. METHODS: Immunological signatures of antigen-specific CTLs were explored in patients with active pulmonary and extra-pulmonary TB, LTBI and in those treated for TB decades ago by using ELISPOT, intracellular flow cytometry and extracellular CD107a detection. RESULTS: No difference was seen between active TB, LTBI or any of those treated for TB in the ELISPOT analysis of antigen-specific Granzyme B (GrB), Perforin (Prf) and interferon-gamma (IFN-γ) producing lymphocytes, the FACS analysis of the intracellular expression of IFN-γ, or the surface expression of CD107a degranulation factor of both CD8+ and CD4+ antigen-specific T cell subsets. The effector memory (TEM) phenotype proved predominant in the surface marker profiling both in active TB and LTBI. The proportion of the CD107a degranulation factor proved higher in the central memory (TCM) than in the other cell subsets in all the study groups. Interestingly, functionally and phenotypically similar CTLs profiles were observed in active TB, LTBI and in all the three groups treated for TB. CONCLUSION: The phenotypic and functional profiling of CTLs has a limited potential in the immunodiagnostics of active TB. Antigen-specific CTLs persist in patients treated for TB decades ago regardless of the efficacy of implemented and completed anti-TB therapy.

Combined Expression of IFN-γ, IL-17, and IL-4 mRNA by Recall PBMCs Moderately Discriminates Active Tuberculosis from Latent Mycobacterium tuberculosis Infection in Patients with Miscellaneous Inflammatory Underlying Conditions.

New biomarkers are needed for discriminating active tuberculosis (TB) from latent TB infection (LTBI), especially in vulnerable groups representing the major diagnostic challenge. This pilot study was carried out to explore the diagnostic potential of selected genes, IFN-γ, IL-17, IL-4, and FoxP3, associated with TB immunity and immunopathology. IFN-γ, IL-17, IL-4, and FoxP3 mRNA expression levels were measured by quantitative reverse transcription PCR (RT-qPCR) from antigen-stimulated peripheral blood mononuclear cells of patients with active TB (n = 25); patients with miscellaneous inflammatory disorders and concomitant LTBI (n = 20), rheumatoid arthritis (RA) being the most predominant in the group (n = 11); and in healthy Bacillus Calmette-Guérin (BCG) vaccinees (n = 8). While the levels of FoxP3 mRNA did not differ between the tested groups, the cumulative expression levels of purified protein derivative-stimulated IFN-γ, IL-17, and IL-4 mRNAs were found to distinguish active TB from the whole group of LTBI with 48% sensitivity and 85% specificity. When restricting the LTBI group to RA cases only, the sensitivity was 56% and specificity 100%. When interpreting the result as positive in at least one of the mRNAs IFN-γ, IL-17, or IL-4, sensitivity of 64% and specificities of 75% (heterogeneous group of LTBI) or 100% (LTBI with RA) were achieved. Moderate discrimination of active TB from LTBI with miscellaneous inflammatory underlying conditions by using combined quantitative expression of IFN-γ, IL-17, and IL-4 mRNA seems not to be of high diagnostic potential.

Epstein-Barr virus in peripheral blood is associated with response to rituximab therapy in rheumatoid arthritis patients.

Autoreactive B cells infected by Epstein-Barr virus (EBV) are suspected to be involved in the etiology of various human chronic autoimmune diseases. This motivated us to study the relationship between peripheral blood EBV load at baseline and treatment response to B cell-depleting therapy in rheumatoid arthritis (RA) patients. Thirty-five RA patients who started treatment with rituximab (RTX) in a routine clinical setting were assessed for baseline disease activity using disease activity score using 28 joint counts (DAS28) (erythrocyte sedimentation rate [ESR]). Treatment response was evaluated 3-7 months after RTX. EBV load in baseline whole blood (WB) samples was determined using quantitative PCR. EBV DNA was detected in 16/35 (46 %) of the WB samples. In these 16 EBV-positive patients, the median viral load was 3.15 (2.68-4.00) log copies/ml. Good/moderate European League Against Rheumatism (EULAR) response was observed in 16/16 of the EBV DNA-positive vs 13/19 EBV DNA-negative patients, p = 0.022. Significant response (DAS28 change >1.2) was observed in 14/16 of the EBV DNA-positive vs 10/19 EBV DNA-negative patients, p = 0.035. The decline in DAS28 after RTX was 2.10 (1.03-4. 78) in the EBV DNA-positive vs 1.47 (-0.7-4.70) in the EBV DNA-negative patients, p = 0.13. EBV load at baseline significantly correlated with change in DAS28 after RTX (τB = -0.261, p = 0.042). Our results suggest that the presence of EBV genome in WB could serve as a predictive marker to RTX therapy in RA.

Rates of serious infections and malignancies among patients with rheumatoid arthritis receiving either tumor necrosis factor inhibitor or rituximab therapy.

OBJECTIVE: Because of the role of tumor necrosis factor (TNF) in host defense, it was hypothesized that its inhibition might lead to an increased risk of malignancies and infections. The objective of our study was to assess the incidence of serious infections leading to hospitalization and malignancies among patients with rheumatoid arthritis (RA) receiving either TNF inhibitor or rituximab (RTX) therapy. METHODS: The study population was identified from the National Register for Biologic Treatment in Finland and the hospital records of Central Finland Central Hospital for conventional disease-modifying antirheumatic drug (cDMARD) users. Data on infections and malignancies were acquired from national healthcare registers. A Poisson model was used to calculate the adjusted incidence rate ratios (aIRR) and was composed of age, sex, time from diagnosis, year of the beginning of the followup, rheumatoid factor status, Disease Activity Score at 28 joints, Health Assessment Questionnaire, prior malignancy, prior serious infection, prior biologic use, and time-updated use of methotrexate, sulfasalazine, hydroxychloroquine, and oral corticosteroids as confounders. RESULTS: In total, during the followup of 10,994 patient-years, 92 malignancies and 341 serious infections were included in the analyses. The aIRR of infections compared to cDMARD users were 1.2 (95% CI 0.63-2.3), 0.84 (95% CI 0.53-1.3), 0.98 (95% CI 0.60-1.6), and 1.1 (95% CI 0.59-1.9) for the patients treated with infliximab (IFX), etanercept, adalimumab, and RTX, respectively. The crude rates of malignancies were highest among the users of cDMARD and RTX, and lowest among patients treated with IFX with no differences in aIRR. CONCLUSION: Our results provide some reassurance of the safety of biologic treatments in the treatment of RA.

Effect of cyclical intermittent etidronate therapy on circulating osteoprotegerin levels in patients with rheumatoid arthritis.

OBJECTIVE: To evaluate the role of serum osteoprotegerin (OPG) as a biochemical marker for disease activity assessment and drug monitoring in patients with rheumatoid arthritis (RA) treated with cyclical etidronate. DESIGN: Forty patients (35 women and 5 men) with RA of <5 years duration were randomized to receive intermittent cyclical etidronate therapy in conjunction with anti-rheumatic therapy or anti-rheumatic therapy alone (without etidronate) in a 2-year, open-label protocol. METHODS: Radiographs of hands and feet and serum samples for the determination of OPG, amino terminal propeptide (PINP), cross-linked C-telopeptide (ICTP) and amino terminal telopeptid of type I collagen were obtained at baseline and at 24 months. RESULTS: Etidronate treatment had no effect on circulating OPG levels, although the significant decline in PINP and ICTP (P=0.001 and P=0.04 respectively) reflected the efficacy of the anti-resorptive therapy. At baseline and at study termination, serum OPG correlated significantly with age (r=0.45; P=0.003 and r=0.56; P=0.0002 respectively). OPG was not related to biochemical markers of bone metabolism, indices of disease activity or radiographic disease progression. At baseline, the mean serum OPG was higher in patients receiving 5-10 mg/day prednisone (82.8+/-4.0 pg/ml, n=16) compared with those receiving <5 mg/day or with no prednisone (69.7+/-4.7 pg/ml, n=23) (P=0.05). CONCLUSIONS: Our results suggested that serum OPG measurement, perhaps because of the complexity of the regulation of the OPG, may be difficult to utilize in the evaluation of anti-resorptive therapy. Moreover, low dose corticosteroid-associated osteoporosis is probably not mediated by inhibition of OPG.

Modification of clearview tuberculosis (TB) enzyme-linked immunosorbent assay for TB patients not infected with HIV.

Diagnosis of active tuberculosis by detection of urinary lipoarabinomannan (uLAM) from Mycobacterium tuberculosis is an attractive approach. Concentrating urine 100-fold allowed quantitation of uLAM at levels equal to picograms/ml of nonconcentrated urine. The approach of concentrating urine 100-fold improved the clinical sensitivity of the Clearview TB enzyme-linked immunosorbent assay (ELISA) from 7% to 57% yet impaired its specificity from 97% to 89%. (This study has been registered at University Hospital of Turku under registration no. 47/180/2009, Helsinki University Central Hospital under 149/2010, University Hospital of Kuopio under 105/2010, and China Medical University Hospital, Taichung, under DMR-99-IRB-075-2.).

Outcomes of switching anti-TNF drugs in rheumatoid arthritis--a study based on observational data from the Finnish Register of Biological Treatment (ROB-FIN).

The aim of this study was to assess, based on observational data from the Finnish Register of Biological Treatment, the outcomes of switching an initial tumor necrosis factor (TNF) blocker to another in the treatment of rheumatoid arthritis (RA). RA patients, who started biological therapy with a TNF blocker between May 1999 and April 2009 and who switched to another TNF blocker, were studied (n=479). The outcomes were assessed according to the reason for and type of the switch. Outcome assessments included American College of Rheumatology 50 responder index (ACR50) response at 3 months after the switch, treatment duration of the second TNF blocker, and swollen joint counts, CRP and DAS28 score at the 3 months, best and last observations of the first and second TNF blocker, respectively. In those who switched due to lack of effectiveness (LOE), the disease activity parameters fell significantly from baseline upon use of infliximab or adalimumab, but had increased prior to the switch. Switching to another TNF blocker (etanercept or adalimumab) restored the response initially achieved with the first TNF blocker. The disease activity parameters fell significantly from baseline upon use of etanercept, and were maintained but not further improved after switching to adalimumab. TNF blocker switching seemed to be most beneficial in secondary LOE (defined as loss of ACR50 response). In those who switched due to adverse events (AE) or other reasons, a similar degree of response as had been achieved with the first agent was also achieved and maintained with the second agent. The results suggest that a second TNF blocker can restore the response in cases of secondary LOE and maintain it after switching due to an AE.

Validation of an automated intact N-terminal propeptide of type I procollagen (PINP) assay.

OBJECTIVES: N-terminal propeptide of type I procollagen assay (PINP) reflects the rate of type I collagen synthesis DESIGN AND METHODS: Different sera were fractioned by gel filtration and analyzed with intact and total PINP assays. The sizes of the antigens were determined by western blotting. The thermal stability was tested at +37°C, +4°C and room temperature (RT). RESULTS: Automated intact PINP assay hardly measured monomeric form. In haemodialysis patients intact and total PINP assays gave significantly different results. The monomeric PINP antigen in serum was larger than the trimeric PINP antigen. PINP were thermally stable at least 7 days at +4°C and at RT but the results of both assays were decreased similarly at +37°C. CONCLUSIONS: The IDS-iSYS intact PINP assay is precise and sensitive. It seems that monomeric form is not derived from the thermal instability of the trimers but acts as a confounding factor.

Biological therapy for psoriatic arthritis in clinical practice: outcomes up to 2 years.

OBJECTIVE: To evaluate the performance of biological drugs in psoriatic arthritis (PsA) in a routine care setting, using the Finnish national register of biological treatment (ROB-FIN). METHODS: Patients with PsA who started therapy with infliximab or etanercept between June 2000 and February 2006 (n = 127) were followed for up to 24 months. Response was evaluated using American College of Rheumatology response criteria including individual measures. RESULTS: Significantly diminished values for swollen and tender joints, patient's global and pain assessments, doctor's global assessment of disease activity, erythrocyte sedimentation rate, C-reactive protein, and Health Assessment Questionnaire score were observed within 3 months after commencement of both infliximab and etanercept. Values remained significantly lower throughout the 24 months of followup. ACR20 response at 3 months was 79% (n = 22/28) for infliximab and 76% (n = 34/45) for etanercept. The first biological drug was discontinued in 16% due to lack of effectiveness and in 6% due to adverse events. CONCLUSION: Anti-tumor necrosis factor-α therapy, often combined with conventional disease-modifying antirheumatic drugs, appeared to have limited toxicity and persistent effectiveness for up to 2 years in a cohort of Finnish patients with severe peripheral PsA.

A case of Poncet disease diagnosed with interferon-gamma-release assays.

BACKGROUND: A 55-year-old, HLA-B27-positive Finnish woman presented with migratory, sterile polyarthritis. INVESTIGATIONS: Physical examination, chest radiography, serologic testing, microscopy, M. tuberculosis-specific interferon gamma enzyme-linked immunospot (ELISPOT) assay, smear and culture of synovial fluid for acid-fast bacilli, and PCR. DIAGNOSIS: The patient's assayed blood and synovial fluid lymphocytes were reactive, and the numbers of M. tuberculosis-specific T cells, as determined by ELISPOT, were twofold to sixfold higher in synovial fluid than in blood. Cultures for acid-fast bacilli remained negative, while PCR specific for DNA of the M. tuberculosis complex was positive in synovial fluid cells. These results suggested that the patient had either active or latent M. tuberculosis infection. This finding, coupled with the presence of polyarthritis, led to a diagnosis of Poncet disease. MANAGEMENT: Standard antituberculous therapy consisting of isoniazid, rifampicin and pyrazinamide was given for 2 months, followed by isoniazid and rifampicin for 4 months. Inflamed joints were treated with methylprednisolone injections. The polyarthritis resolved within 4 months of initiating antituberculous therapy.

Erosive arthritis in a patient with pycnodysostosis: an experiment of nature.

OBJECTIVE: The excellent poster painter Henri de Toulouse-Lautrec is the most famous patient with cathepsin K-deficient pycnodysostosis. Cathepsin K is believed to play a major role in osteoclast-driven bone resorption. In this study we explored the role of cathepsin K in bone resorption in a patient with a cathepsin K mutation causing pycnodysostosis in whom psoriatic arthritis also developed. We hypothesized that the patient would develop only inflammatory synovitis but would not develop bone erosions or other osteolytic changes. METHODS: Monocytes from the patient with pycnodysostosis and normal control monocytes were isolated and stimulated to fuse and form multinuclear osteoclast-like cells, which were identified by evaluating messenger RNA expression of osteoclast markers. The ability to resorb bone was assessed by determining the extent of pit formation and levels of collagen degradation products generated by cathepsin K (C-terminal crosslinking telopeptide of type I collagen [CTX]) and matrix metalloproteinases (pyridinoline crosslinked C-terminal telopeptide of type I collagen). These experiments were also done in normal control cells after incubation with the cathepsin K inhibitor E64 during bone resorption. RESULTS: In contrast to our a priori hypothesis, the patient developed a mutilating disease with extensive bony erosions associated with lysis of some of the distal phalanges of her hands and feet. After stimulation of monocytes from this patient, the cells formed multinuclear tartrate-resistant acid phosphatase-positive and calcitonin receptor-positive multikaryons, which, however, totally lacked cathepsin K. These multinuclear cells were able to resorb bone but, in contrast to normal control osteoclasts, did not produce CTX. The resorption pattern was abnormal in that, unlike normal control osteoclasts, both osteoclasts from the patient and E64-inhibited osteoclasts did not leave extensive osteoclast trails, but were relatively sessile. CONCLUSION: In this "experiment of nature" we observed that cathepsin K is not necessary for bone degradation. These findings may be pertinent to our understanding of the functions of cathepsin K inhibitors, which are currently being developed as drugs to treat metabolic bone diseases.

Two year randomized controlled trial of etidronate in rheumatoid arthritis: changes in serum aminoterminal telopeptides correlate with radiographic progression of disease.

OBJECTIVE: To investigate the effect of intermittent cyclical etidronate treatment on radiographic progression, bone collagen markers, and clinical disease activity in patients with rheumatoid arthritis (RA). METHODS: Forty patients with RA of less than 5 years' duration were randomized to receive intermittent cyclical etidronate therapy in conjunction with antirheumatic therapy or antirheumatic therapy alone (without etidronate) in a 2 year open-label protocol. Radiographs of hands and feet and serum samples for determination of aminoterminal propeptide (PINP), crosslinked C-telopeptide (ICTP), and aminoterminal telopeptides (NTx) of type I collagen were obtained at baseline and at 24 months. RESULTS: There was significant and similar worsening of the radiologic scores in both treatment groups. Both PINP, a marker of bone formation, and ICTP, an indicator of collagen degradation, declined in the etidronate group compared to the control group (p = 0.001 and p = 0.042, respectively). The groups did not differ for the change in serum NTx, a specific systemic marker of osteoclastic bone resorption. However, the change in serum NTx correlated significantly with the increase in erosion score in the total study population and in the control group (r = 0.41, p = 0.01 and r = 0.48, p = 0.034, respectively). CONCLUSION: Etidronate therapy did not prevent radiologic progression in patients with RA, but the decline in serum PINP and ICTP concentrations suggests a favorable effect on general bone metabolism. Correlation between the change in serum NTx and worsening of the erosion score provides biochemical evidence that osteoclast is the principal cell type responsible for focal bone resorption in RA.