Nete Hornung

Soluble CD163: a marker molecule for monocyte/macrophage activity in disease.

By immunoprecipitation we have identified a soluble plasma form of CD163 (sCD163), the IL-6 inducible macrophage-receptor for clearing haptoglobinhaemoglobin complexes. A sandwich ELISA for measuring sCD163 was established and used to determine the sCD163 levels in normal subjects and patients with inflammatory and myeloproliferative diseases. In normal subjects, the concentration of sCD163 was high (median 1.9 mg/l) with low intraindividual variation. Highly increased levels were seen in patients with sepsis, myeloid leukaemia and in patients with Gaucher disease characterized by accumulation of tissue macrophages. Although the physiological role of sCD163 remains unknown, our present data suggest that sCD163 might prove to be a valuable marker molecule in infectious and myeloproliferative diseases.

Plasma Adiponectin in Patients with Active, Early, and Chronic Rheumatoid Arthritis Who Are Steroid- and Disease -Modifying Antirheumatic Drug-Naive Compared with Patients with Osteoarthritis and Controls

Objective. Rheumatoid arthritis (RA) is a systemic chronic inflammatory joint disease, whereas osteoarthritis (OA) is a local joint disease with low-level inflammatory activity. The pathogenic role of the adipocytokine adiponectin is largely unknown in these diseases. We hypothesized (1) that plasma adiponectin concentrations differ in healthy controls and patients with early disease-modifying antirheumatic drug (DMARD)-naive RA, chronic RA, and OA; (2) that changes in adiponectin are observed during methotrexate (MTX) treatment of chronic RA; and (3) that adiponectin correlates to disease activity measures in RA. Methods. Plasma adiponectin was analyzed with a validated in-house immunoassay. We measured adiponectin in healthy controls (n = 45) and patients with early DMARD-naive RA (n = 40), chronic RA (n = 74), and OA (n = 35). In a subgroup of patients with chronic RA (n = 31), the longitudinal effect of MTX treatment on adiponectin (Week 0 vs Week 28) was investigated. Results. Adiponectin differed significantly between healthy controls (mean 4.8 ± SD 2.7 mg/l) and the 3 groups, with 8.9 ± 4.8 mg/l in early RA, 11.6 ± 5.6 mg/l in chronic RA, and 14.1 ± 6.4 mg/l in OA. Longitudinally, MTX treatment increased adiponectin significantly from 9.7 ± 4.5 mg/l at Week 0 to 11.0 ± 4.5 mg/l at Week 28 in chronic RA. No correlations to disease activity measures were found. Conclusion. Both early DMARD-naive and chronic RA were associated with higher plasma adiponectin compared to healthy controls, but lower plasma adiponectin than OA. Adiponectin increased 13% during MTX treatment. In patients with RA and OA body mass index, age, sex, and disease activity measures failed to explain the findings.

Fecal calprotectin in healthy children

Abstract Calprotectin is a protein found in the cytosol of inflammatory cells and is a marker of the presence and the degree of inflammation in the bowel system. Calprotectin in feces has great diagnostic value in the matter of inflammatory bowel disease (IBD). In feces, the protein is stable up to seven days, and since the protein can easily be measured with an ELISA, the use of fecal calprotectin (FC) means no invasive measures. For adults and children over 4 years, a cut-off level of 50 mg/kg has been well established for diagnostic purposes. Because previous studies have proven that children under the age of four in general have higher FC values than older children and adults, there is a need for a cut-off level for this age group. In order to establish that, the normal values for FC in children from 0–4 years were investigated. Some 75 stool samples from healthy children were collected and the levels of FC were analyzed. The results were compared to 157 pediatric cases where FC analysis had been performed for diagnostic purposes. As a result, three cut-off levels were established based on the 97.5% percentiles of FC in different age groups: 538 mg/kg (1 < 6 months), 214 mg/kg (6 months < 3 years) and 75 mg/kg (3 < 4 years).

Differential effect of methotrexate on the increased CCR2 density on circulating CD4 T lymphocytes and monocytes in active chronic rheumatoid arthritis, with a down regulation only on monocytes in responders

Objectives: To evaluate the effect of orally administered methotrexate (MTX) on the density of CC chemokine receptor 2 (CCR2) and CXC chemokine receptor 3 (CXCR3) on circulating monocytes, and the coexpression of CXCR3 and CCR2 on CD4 T lymphocytes in patients with active chronic rheumatoid arthritis. Methods: All 34 patients with rheumatoid arthritis fulfilled the 1987 American Rheumatism Association criteria and were followed for 16 weeks after starting MTX. Peripheral blood mononuclear cells were analysed for CCR2 and CXCR3 density by three-colour flow cytometry before initiation of MTX and at week 12. Results: 22 (65%) patients were non-responders, 12 (35%) patients responded to MTX by American College of Rheumatology (ACR)20% criteria, and 8 (24%) of these patients responded by ACR50%. In patients with active rheumatoid arthritis before starting MTX, CCR2 density on circulating monocytes, CD4+ CXCR3+ and CD4+ CXCR3− T lymphocytes was increased compared with controls. During 12 weeks of MTX treatment, the CCR2 density on monocytes decreased significantly in the ACR50% group but not in the ACR20% and non-responder groups. The increased CCR2 density on CD4+ CXCR3+ and CD4+ CXCR3− T lymphocytes was unaffected by the reduction in disease activity measured in relation to MTX treatment. The percentage of both monocytes and CD4+ CXCR3+ and CD4+ CXCR3− T lymphocytes among the peripheral circulating mononuclear cells did not change during MTX treatment. Conclusions: Active chronic rheumatoid arthritis is characterised by enhanced CCR2 density on circulating monocytes and CD4+ CXCR3+ and CD4+ CXCR3− T lymphocytes. During MTX treatment, a decrease in CCR2 density on monocytes in the ACR50% responder group was associated with decreased disease activity. The increased CCR2 density on CD4+ CXCR3+ and CD4+ CXCR3− T lymphocytes was uninfluenced by MTX and disease activity.

In Active Chronic Rheumatoid Arthritis, Dipeptidyl Peptidase IV Density is Increased on Monocytes and CD4 + T lymphocytes

The effect of low‐dose methotrexate (MTX) treatment on the CD26 density on circulating monocytes and CD4+ T lymphocytes or levels of soluble CD26 (sCD26) has not yet been described in rheumatoid arthritis (RA). While CD26 in T lymphocytes is involved in the activation and proliferation of T lymphocytes, little is known of the role of CD26 in monocytes as it has only recently been localized to monocytes. We analysed the CD26 density by flow cytometry and levels of sCD26 in plasma before initiation of MTX treatment and 12u2003weeks later. This was done on 34 RA patients fulfilling the 1987 American College of Rheumatology (ACR) criteria followed for 16u2003weeks after starting MTX treatment. CD26 density on monocytes was increased in RA patients compared with healthy controls before MTX treatment (Pu2003<u20030.01). After 12u2003weeks of MTX treatment, the CD26 density on monocytes decreased significantly in the ACR‐50% group (Pu2003=u20030.03), but not in the ACR‐20% and the non‐responder group (Pu2003=u20030.15 and 0.87). The increased CD26 density on CD4+ T lymphocytes (Pu2003<u20030.01) was unaffected by the reduction in disease activity in relation to MTX treatment. The percentage of monocytes and CD4+ T lymphocytes among peripheral blood circulating mononuclear cells did not change during MTX treatment. No effect of MTX treatment was observed on the plasma levels of sCD26. Active chronic RA is characterized by enhanced CD26 density on circulating monocytes and CD4+ T lymphocytes. MTX treatment decreased CD26 density on monocytes in the ACR‐50% responder group and was associated with decreased disease activity. The enhanced CD26 density on CD4+ T lymphocytes was uninfluenced by MTX treatment.

A new quantitative RT-PCR assay for thymidylate synthase mRNA in blood leukocytes applied to cancer patients and healthy controls.

Thymidylate synthase (TS) is the target enzyme for 5-fluorouracil (5-FU). TS mRNA and protein levels in colorectal tumours are among the most important determinants for tumour response to 5-FU. TS mRNA levels in blood leukocytes may give information on pharmacokinetic and pharmacodynamic actions of 5-FU on TS as it has previously been shown that inhibition of TS levels by 5-FU in bone marrow leukocytes resembles the degree of TS inhibition in colorectal tumours. The aim of this study was to develop a quantitative high-throughput RT-PCR assay for TS mRNA expression in blood leukocytes (CURT-PCR). Furthermore the TS mRNA levels in blood of patients with colorectal cancer and healthy controls was compared. TS mRNA levels in 17 patients with colorectal cancer did not differ from 20 matched controls whereas a group of 14 younger controls had significantly lower TS mRNA expression than patients and matched controls. In order to investigate the sensitivity of the assay towards cellular reactions such as proliferative stimuli, isolated blood leukocytes were stimulated with phytohemagglutinin both in mitogenic and non-mitogenic concentrations and an induction of TS mRNA expression was measured in both cases. TS activity and cellular proliferation also increased but only at mitogenic concentrations, suggesting that TS mRNA expression is an early leukocyte activation marker. This new CURT-PCR assay may allow improved studies of functional kinetics of drugs with impact upon TS. Further studies are required to establish the possible clinical benefit of TS mRNA measurements in blood leukocytes.

Calprotectin in patients with chronic rheumatoid arthritis correlates with disease activity and responsiveness to methotrexate

Abstract Objective: Calprotectin (myeloid-related protein 8/14) is elevated in inflammatory diseases and a correlation of serum calprotectin and disease activity in rheumatoid arthritis (RA) has been shown. In this study, we investigated plasma calprotectin as a disease marker in patients with chronic RA treated with methotrexate (MTX) monotherapy and compared plasma calprotectin with C-reactive protein (CRP) in this matter. Methods: Seventy-six patients with chronic RA were included in this open prospective study and of these 40 were included prior to initiation of MTX therapy. The patients were followed with laboratory and clinical parameters for 52–56u2009weeks. Plasma calprotectin was analyzed at the start of study and at various intervals. Radiographic evaluation was performed at baseline and after 17.2u2009months and progression in joint destruction was measured with Larsen score. The response to MTX was evaluated according to the American College of Rheumatology criteria. Results: Patients starting MTX treatment had significantly higher levels of plasma calprotectin compared to patients well established on MTX therapy (pu2009=u2009.008). Among the 40 patients naive to MTX, 25 responded to MTX therapy and serum calprotectin decreased significantly in these patients (pu2009=u2009.0007). The radiographic damage showed no relation to calprotectin. Conclusions: Plasma calprotectin is associated with disease activity in patients with chronic RA and is more strongly correlated to MTX response compared to CRP. The role of calprotectin as a disease marker is promising and the advantages compared to CRP needs to be further investigated.

Elevated calprotectin in patients with atrial fibrillation with and without heart failure

Abstract Calprotectin is an inflammatory marker, which has been found elevated in patients suffering from cardiac conditions, e.g. myocardial infarction, unstable angina and chronic heart failure. Inflammation has further been linked to atrial fibrillation (AF). However, the association between calprotectin and AF is unknown. We aimed to compare calprotectin levels in patients suffering from AF with healthy adults. In addition, AF patients with and without heart failure were compared. Calprotectin was measured in patients undergoing elective direct current cardioversion for AF. Calprotectin was determined before, 4u2009hours and 3 months after cardioversion. Healthy blood donors were used to verify the reference interval for calprotectin. In total, 104 prospectively enrolled patients were included. The median serum calprotectin level for AF patients was 1.6u2009μg/mL before cardioversion. Calprotectin levels increased significantly 4u2009h (1.9u2009μg/mL) and 3u2009months (2.2u2009μg/mL) after cardioversion. Blood donors’ median serum calprotectin (1.3u2009μg/mL) was significantly lower than AF patients. AF patients with heart failure had significantly higher calprotectin at baseline compared with AF patients without a history of heart failure (2.0u2009μg/mL vs. 1.5u2009μg/mL). The difference was not significant at 4u2009h (2.0u2009μg/mL vs. 1.7u2009μg/mL) or 3u2009months (2.5u2009μg/mL vs. 2.2u2009μg/mL). In conclusion, the calprotectin levels in patients with AF were significantly higher than healthy blood donors and were further increased after cardioversion. AF patients with heart failure had significantly higher levels of calprotectin than AF patients without heart failure.