[Clinical presentation of tuberculosis in routine practice]. / Klinische Präsentationen der Tuberkulose im Alltag.

Since the beginning of the biologics era tuberculosis is known to be a potential life-threatening complication during treatment of patients with rheumatic diseases. National and international societies have developed recommendations for tuberculosis screening and treatment of patients at risk for development of tuberculosis. Owing to the relative rareness of overt tuberculosis in patients with rheumatic diseases, the experience of individual rheumatologists with this complication is limited. Therefore, we have analyzed the tuberculosis cases from 2006-2011 in our rheumatology referral center (treating more than 1,500 inpatient and 8,000 outpatient cases every year) to obtain a real-life picture more than 10 years after initiation of the first application of biologics outside of controlled clinical trials. We identified 4 cases that illustrate the difficulties of diagnosis and treatment.

Differential transcriptome analysis of intraarticular lesional vs intact cartilage reveals new candidate genes in osteoarthritis pathophysiology.

OBJECTIVE: To elucidate disease-specific molecular changes in osteoarthritis (OA) by analyzing the differential gene expression profile of damaged vs intact cartilage areas within the same joint of patients with OA of the knee using a combination of a novel RNA extraction technique and whole-genome oligonucleotide arrays. METHODS: The transcriptome of macroscopically affected vs intact articular cartilage as determined by visual assessment was analyzed using an optimized mill-based total RNA isolation directly from the tissue and high density synthetic oligonucleotide arrays. Articular cartilage samples were obtained from patients with OA of the knee. Expression of differentially regulated genes was validated by real-time quantitative polymerase chain reaction and immunohistochemistry. RESULTS: The amount of RNA obtained by the optimized extraction procedure was at least 1 microg per 500 mg of cartilage and fulfilled the common quality requirements. After hybridization onto HG-U133 Plus 2.0 GeneChips (Affymetrix), 28.6-51.7% of the probe sets on the microarray showed a detectable signal above the signal threshold in the individual samples. A subset of 411 transcripts, which appeared to be differentially expressed, was obtained when applying predefined filtering criteria. Of these, six genes were found to be up-regulated in the affected cartilage of all patients, including insulin-like growth factor binding protein 3 (IGFBP-3), wnt-1-inducible signaling protein 1 (WISP-1), aquaporin 1 (AQP-1), delta/notch-like EGF-repeat containing transmembrane (DNER), decay accelerating factor (DAF), complement factor I (IF). CONCLUSION: The optimized methodical approach reported here not only allows to determine area-specific gene expression profiles of intraindividually different low-RNA containing OA cartilage specimens. In addition, this study also revealed novel genes not yet reported to play a role in the pathophysiology of joint destruction in OA.

Evaluation of latent tuberculosis infection in patients with inflammatory arthropathies before treatment with TNF-alpha blocking drugs using a novel flow-cytometric interferon-gamma release assay.

OBJECTIVE: To compare the efficacy of the conventional skin test and a novel flow cytometric whole blood assay in the diagnosis of latent tuberculosis infection (LTBI) in patients with rheumatological diseases evaluated for treatment with TNF-alpha-blocking agents. METHODS: Prospective study of 97 consecutively enrolled patients, who were assessed for the presence of LTBI through clinical history, Mendel-Mantoux skin testing and chest X-ray. In addition, T-cell reactivity towards tuberculin (PPD, purified protein derivative) and the Mycobacterium tuberculosis-specific proteins ESAT-6 and CFP-10 was determined ex vivo using a flow cytometric whole blood assay. RESULTS: After standard screening, 15% of patients receiving TNF-alpha-blocking therapy were pretreated with isoniazide (INH), another 5% of patients did not receive TNF-alpha-blocking therapy because of LTBI. PPD-reactivity in the skin was observed in 14% of patients compared with 39% with the whole blood test. Analysis of the M. tuberculosis-specific response to ESAT-6 and CFP-10 revealed positive results in 16% of patients. Using a decision tree incorporating history, chest X-ray and either skin-test or ESAT-6/CFP-10 results, 18 or 22% of the patients, respectively, were classified as latently infected with M. tuberculosis. Four patients treated with INH because of a positive skin reaction did not show reactivity to ESAT-6/CFP-10 in the whole blood assays. Another six patients not pretreated with INH because of negative skin tests would have received INH, had the results of the whole blood assay been taken into account. CONCLUSION: The Mendel-Mantoux skin test has a low sensitivity and specificity for the diagnosis of LTBI in this cohort of patients, potentially resulting in both over- and under-treatment with prophylactic INH when compared with the flow cytometric analysis of whole blood T-cell reactivity to proteins specific to M. tuberculosis. Use of T-cell based in vitro tests may help to refine diagnostic testing for LTBI.

Generation of classification criteria for chronic fatigue syndrome using an artificial neural network and traditional criteria set.

OBJECTIVE: The definition of chronic fatigue syndrome (CFS) is still disputed and no validated classification criteria have been published. Artificial neural networks (ANN) are computer-based models that can help to evaluate complex correlations. We examined the utility of ANN and other conventional methods in generating classification criteria for CFS compared to other diseases with prominent fatigue, systemic lupus erythematosus (SLE) and fibromyalgia syndrome (FMA). PATIENTS AND METHODS: Ninety-nine case patients with CFS, 41 patients with SLE and 58 with FMA were recruited from a generalist outpatient population. Clinical symptoms were documented with help of a predefined questionnaire. The patients were randomly divided into two groups. One group (n = 158) served to derive classification criteria sets by two-fold cross-validation, using a) unweighted application of criteria, b) regression coefficients, c) regression tree analysis, and d) artificial neural networks in parallel. These criteria were validated with the second group (n = 40). RESULTS: Classification criteria developed by ANN were found to have a sensitivity of 95% and a specificity of 85%. ANN achieved a higher accuracy than any of the other methods. CONCLUSION: We present validated criteria for the classification of CFS versus SLE and FMA, comparing different classification approaches. The most accurate criteria were derived with the help of ANN. We therefore recommend the use of ANN for the classification of syndromes with complex interrelated symptoms like CFS.

[Pharmacotherapeutic aspects of femoral head necrosis]. / Pharmakotherapeutische Aspekte der Femurkopfnekrose.

Avascular necrosis (AVN) of the hip is one of the unsolved problems in orthopedics, especially with regard to drug therapy. Only a few studies exist using a long-term approach with vasodilating agents such as prostaglandins, anticoagulants, hormones, as well as lipid lowering and bone protective drugs such as bisphosphonates. However, using these medications several studies have demonstrated a significant reduction in pain, in the destruction of the femoral head as well as in overall disability. This resulted in a reduced need for joint replacement in patients with AVN and to a substantial improvement in quality of life. Of note, drug therapy should be initiated in the early phases of AVN as later stages appear to be less responsive to medication.

Increased titres of anti-nuclear antibodies do not predict the development of associated disease in the absence of initial suggestive signs and symptoms.

OBJECTIVE: To determine whether patients with elevated anti-nuclear antibodies (ANA), absent extractable nuclear antigen (ENA) reactivity, and no definite associated disease develop an ANA-associated disease (AAD). METHODS: Patients with ANA titres of at least 1:320 and no ENA reactivity were identified by searching the database of our laboratory serving a tertiary care university hospital between 1998 and 2002. Medical records of this index time point were reviewed to exclude patients with active AAD at screening. Case patients were contacted by questionnaire between 2004 and 2005 and invited for a clinical visit to ascertain the individual disease status. RESULTS: Seventy-six patients were evaluated after a median follow-up of 32 months. An AAD was diagnosed in eight patients: connective tissue disease (CTD) in three, autoimmune hepatitis in two, rheumatoid arthritis in one, encephalomyelitis disseminate in one, and lymphoma in one. The only predictive factor associated with the development of AAD was the suspicion of an autoimmune disease by the treating physician at the initial evaluation. In the absence of initial suspicion for an autoimmune disease, only two out of 54 patients developed AAD, whereas six out of 22 patients with initial disease suspicion developed a defined AAD. CONCLUSION: In the absence of a clinical suspicion, elevated ANA titres have a low positive predictive value of 4% for developing AAD for the upcoming 3 years.

Antibiotic treatment of Lyme borreliosis: what is the evidence?

Antibiotic treatment of all disease manifestations after infection with Borrelia sensu lato spp aims at resolving the presenting disease manifestation and preventing late stage disease. The goals are resolution of the preventing manifestation and prevention of the spread of bacteria to prevent late disease like arthritis. Different borrelial species prevail in Europe. The natural disease course of European borreliosis is not well defined and the effect of antibiotic treatment is unclear.

Stringent endocrinological testing reveals subnormal growth hormone secretion in some patients with fibromyalgia syndrome but rarely severe growth hormone deficiency.

OBJECTIVE: Several reports suggest that growth hormone (GH) deficiency may be a pathogenic factor in fibromyalgia syndrome (FM). This hypothesis has never been adequately examined. METHODS: We measured serum GH concentration after insulin induced hypoglycemia in subjects with FM. GH secretion in subjects with a maximal GH increase < 10 ng/ml after hypoglycemia was assessed by additional arginine stimulation. RESULTS: In one of 56 subjects tested, GH remained below 3 ng/ml in both tests, satisfying the criteria for adult GH deficiency. Thirty-two subjects (67%) had a maximal GH > 10 ng/ml. We retrospectively found an inverse correlation between low density lipoprotein levels and maximal GH concentration in a subgroup of patients. CONCLUSION: Severe GH deficiency is not a significant pathogenic factor in most patients with FM. We observed an impaired reactivity of the somatotropic axis in one-third of patients with FM, in keeping with a functional alteration of the hypothalamus.

[Excessive hyperferritinemia as an indication of a reactive hemophagocytosis syndrome]. / Exzessive Hyperferritinämie als Hinweis auf ein reaktives Hämophagozytose-Syndrom.

HISTORY AND ADMISSION FINDINGS: A 17-year-old girl with a history of a polyarthritis of unknown etiology was admitted because of acute fever and general weakness. There were palpable cervical lymph nodes and her body temperature was 39.5 degrees C. INVESTIGATIONS: GOT was raised to 282 U/1, GPT to 266 U/l lactate dehydrogenase to 1275 U/I and bilirubin to 0.6 mg/dl. The Quick value was 67%, albumin 28 mg/dl. White cell count was decreased to 1700/microl, with 43% granulocytes, 39% lymphocytes, 17% monocytes. Platelet count was 64,000/microl. Ultrasound revealed splenomegaly. Ferritin was markedly raised to 11,860 ng/ml (normal up to 150 ng/ml). An epstein-barr-virus infection was found. THERAPY AND CLINICAL COURSE: Suspecting a reactive hemophagocytosis syndrome, she was treated with prednisolone (2 mg/kg). The diagnosis was confirmed by a bone marrow aspirate. The patient's condition and laboratory values improved rapidly. CONCLUSION: Markedly increased ferritin levels in a clinically septic patient with an underlying rheumatic disease indicates a hemophagocytotic syndrome. High dosage steroid should be started before there is biopsy confirmation of the disease.

Cartilage oligomeric matrix protein (COMP) and collagen IX are sensitive markers for the differentiation state of articular primary chondrocytes.

Primary chondrocytes dedifferentiate in serial monolayer with respect to their morphological and biosynthetic phenotype. They change from a round to a flattened fibroblast-like shape, and collagen I is secreted instead of the cartilage-specific collagen II. We analysed in detail the time course of dedifferentiation of mature bovine articular chondrocytes in monolayer for up to 32 weeks. Assessment of RNA expression by reverse transcription-PCR led to the identification of two novel phenotypical markers, the cartilage oligomeric matrix protein (COMP) and collagen IX, which are down-regulated faster than the widely accepted marker, collagen II. The different kinetics of COMP and collagen expression suggest differential regulation at the level of transcription. Immunostaining and metabolic labelling experiments confirmed the switch in the collagen expression pattern and the rapid down-regulation of de novo synthesis of COMP and collagen IX. Culture of chondrocytes in a three-dimensional matrix is known to stabilize the chondrocytic phenotype. We maintained cells for up to 28 weeks in an alginate bead system, which prevented dedifferentiation and led to a stabilization of collagen and COMP expression. Immunohistochemical analysis of the alginate beads revealed a similar distribution of matrix proteins to that found in vivo. Chondrocytes were transferred after a variable length of monolayer culture into the alginate matrix and the potential for redifferentiation was investigated. The re-expression of COMP and collagen IX was differentially regulated. The expression of COMP was re-induced within days after transfer into the three-dimensional matrix, while the expression of collagen IX was irreversibly down-regulated. In summary, these results demonstrate that the potential for redifferentiation decreases with increasing length of monolayer culture and show that the alginate bead system represents an attractive in vitro model to study the chondrocyte de- and re-differentiation processes, as well as extracellular matrix assembly.

Comparison of long-term transgene expression after non-viral and adenoviral gene transfer into primary articular chondrocytes.

Different gene transfer approaches to achieve long-term transgene expression in cultured primary bovine chondrocytes were compared using enhanced green fluorescent protein (EGFP) as a reporter. Transduction with a high-capacity adenoviral vector was 82% efficient when analysed by fluorescence microscopy, while up to 42% of plasmid-transfected cells were EGFP positive with FuGene as a transfection reagent. Rapid dominant marker selection of plasmid-transfected cells was achieved in monolayer culture. With either method of gene transfer, a high proportion of the chondrocytes remained transgene positive during prolonged alginate culture. Transgene transcription in single cells was quantified with a confocal laser scanning microscope. Detection of EGFP expression was more sensitive with this method, identifying more transgene-expressing cells than conventional fluorescence microscopy. Long-term EGFP expression was higher in adenovirally transduced chondrocytes embedded in alginate as compared to plasmid-transfected cells cultured in monolayer or in alginate. Both the adenoviral and the plasmid-based approach appear suited for studies of the molecular and cellular mechanisms by which mutations in cartilage matrix proteins cause disease.