Chemokine and chemokine receptor expression in paired peripheral blood mononuclear cells and synovial tissue of patients with rheumatoid arthritis, osteoarthritis, and reactive arthritis.

BACKGROUND: Chemokine receptors and chemokines have a crucial role in leucocyte recruitment into inflamed tissue. OBJECTIVE: To examine the expression of an extensive number of chemokines and receptors in a unique bank of paired samples of synovial tissue (ST) and peripheral blood (PB) from patients with different forms of arthritis to assist in identifying suitable targets for therapeutic intervention. METHODS: Synovial biopsy specimens were obtained from 23 patients with rheumatoid arthritis (RA), 16 with osteoarthritis, and 8 with reactive arthritis. ST chemokine (CCL2/MCP-1, CCL5/RANTES, CCL7/MCP-3, CCL8/MCP-2, CCL14/HCC-1, CCL15/HCC-2, CCL16/HCC-4), chemokine receptor (CCR1, CCR2b, CCR5, CXCR4), and CD13 expression was analysed by immunohistochemistry and two colour immunofluorescence. Chemokine receptor expression (CCR1, CCR3, CCR5, CCR6, CCR7) on PB cells was studied by flow cytometry. Non-parametric tests were used for statistical analysis. RESULTS: Abundant expression of CCR1, CXCR4, and CCR5 was found in all forms of arthritis, with a specific increase of CCL5 and CCL15 in RA. CCL7, CCL8, CCL14, CCL15, and CCL16 were detected for the first time in ST. The results for PB analysis were comparable among different arthritides. Interestingly, compared with healthy controls, significantly lower expression of CCR1 (p<0.005) and CCR5 (p<0.05) by PB monocytes in the patient groups was seen. DISCUSSION: A variety of chemokines and receptors might have an important role in several inflammatory joint disorders. Although other receptors are involved as well, migration of CCR1(+) and CCR5(+) cells towards the synovial compartment may play a part in the effector phase of various forms of arthritis.

Synovial tissue macrophages: a sensitive biomarker for response to treatment in patients with rheumatoid arthritis.

BACKGROUND: Previous work identified synovial sublining macrophage numbers as a potential biomarker for clinical efficacy in rheumatoid arthritis. OBJECTIVE: To investigate the association between changes in infiltration of synovial macrophages and clinical improvement after antirheumatic treatment. METHODS: 88 patients who participated in various clinical trials were studied. All patients underwent serial arthroscopy before initiation of treatment and after different time intervals. Immunohistochemical and digital image analysis were performed according to standardised procedures to detect changes in CD68+ synovial sublining macrophages in relationship to changes in the 28 joint count Disease Activity Score (DAS28). Statistical analysis was performed using one way analysis of variance, the independent samples t test, linear regression, and the standardised response mean (SRM). RESULTS: For good, moderate, and non-responders, according to the DAS28 response criteria, there was a significant difference in the change in sublining macrophages (mean (SEM) cells/mm(2) -643 (124), -270 (64), and -95 (60), respectively; p<0.0003). There was a significant correlation between the change in the number of macrophages and the change in DAS28 (Pearson correlation 0.874, p<0.01). The change in sublining macrophages explained 76% of the variation in the change in DAS28 (p<0.02). The sensitivity to change of the biomarker was high in patients treated actively (SRM >0.8), whereas the ability to detect changes in placebo treated patients was weak (SRM <0.3). CONCLUSION: The results suggest that changes in synovial sublining macrophages can be used to predict possible efficacy of antirheumatic treatment.

[Blockade of chemokines in chronic inflammatory diseases]. / Blokkade van chemokinen bij chronische ontstekingsziekten.

Chemokines form a relatively new family of chemotactic cytokines that seem to play a central role in the migration and activation of leukocytes in a wide variety of immune-mediated disorders. Specific therapy targeted against these proteins may well become an effective treatment in many of these disorders. The efficacy and safety of chemokine blockade is currently being investigated in clinical trials on patients with HIV infection, multiple sclerosis, rheumatoid arthritis and other chronic inflammatory disorders. The first results have been reported in patients with rheumatoid arthritis and these seem to confirm the potency of this treatment.

Chemokines in joint disease: the key to inflammation?

Targeting chemokines and/or chemokine receptors appears to be an intriguing new approach to treating chronic inflammatory disorders like rheumatoid arthritis, inflammatory bowel diseases, multiple sclerosis, and transplant rejections. The involvement of chemokines and chemokine receptors in inflammatory joint diseases, the in vitro and in vivo characteristics of the chemokine family in inflammatory joint disease, and initial clinical data on chemokine blockade in patients with rheumatoid arthritis suggest that targeting the chemokine and chemokine receptor family might provide a new, promising antirheumatic strategy.

T cells, fibroblast-like synoviocytes, and granzyme B+ cytotoxic cells are associated with joint damage in patients with recent onset rheumatoid arthritis.

OBJECTIVE: To determine immunohistological markers in synovial tissue of patients with early rheumatoid arthritis (RA) which are associated with unfavourable disease outcome. METHODS: Synovial tissue was obtained from 36 patients with RA within 1 year after the initial symptoms and before starting disease modifying antirheumatic drug treatment. Clinical, laboratory, and radiological assessments (Larsen score) were performed at the time of the biopsy and at the end of follow up (mean 58 months, range 38-72). Immunohistological analysis was performed to detect T cells, B cells, plasma cells, fibroblast-like synoviocytes (FLS), macrophages, and granzyme B+ cytotoxic cells. The sections were evaluated by digital image analysis. RESULTS: Patients were divided into two groups based upon the radiological progression per year of follow up: group I with mild progression (n = 20; Larsen <2 points/year); group II with more severe progression (n = 16; Larsen > or =2 points/year). Regression analysis with a univariate model showed that the numbers of granzyme B+ cytotoxic cells (relative risk (RR) = 12, p = 0.003), T cells (RR = 11, p = 0.013), and FLS (RR = 10, p = 0.020) discriminated between groups I and II. A multivariate model demonstrated that the numbers of T cells (RR = 1.2, p = 0.015) and FLS (RR = 1.4, p = 0.013) were independent discriminators between groups I and II. CONCLUSION: The numbers of granzyme B+ cytotoxic cells, T cells, and FLS in synovial tissue of patients with RA are related to the severity of joint damage. The data suggest a pathogenetic role for these cells in the process of joint damage.

Chemokine blockade and chronic inflammatory disease: proof of concept in patients with rheumatoid arthritis.

BACKGROUND: Chemokines and their receptors are considered important contributors in cell migration and inflammation in chronic inflammatory disorders. Chemokines affecting monocytes/macrophages are considered potential therapeutic targets, but no studies of the effects of blocking the chemokine repertoire in humans with a chronic inflammatory disease have been reported. OBJECTIVE: To carry out a double blind, placebo controlled, phase Ib clinical trial with a specific, oral CCR1 antagonist. METHODS: 16 patients with active rheumatoid arthritis (RA) were randomised 3:1 to active:placebo treatment for 14 days. Synovial biopsy specimens were obtained on days 1 and 15. Immunohistochemistry was used to detect the presence of various cell types before and after treatment and the results measured by digital image analysis. Results before and after treatment were compared by paired t test, and a two sample t test was used to compare the changes from baseline in the two groups. RESULTS: All patients completed the study. A significant reduction in the number of macrophages (p=0.016), intimal macrophages (p=0.026), and CCR1+cells (p=0.049) in patients treated with the chemokine antagonist compared with the placebo group occurred in the synovium. Significant decreases in overall cellularity, intimal lining layer cellularity, CD4+ T cells, and CD8+ T cells also occurred in treated patients. Cells lacking CCR1 were not affected. Trends towards clinical improvement were seen in the treated patients but not in the placebo group. Severe side effects were not reported. CONCLUSION: Specific chemokine receptor blockade can result in relevant biological effects in patients with active RA.

The development of clinical signs of rheumatoid synovial inflammation is associated with increased synthesis of the chemokine CXCL8 (interleukin-8).

Paired synovial tissue samples were obtained from both clinically uninvolved (CU) and clinically involved (CI) knee joints of eight rheumatoid arthritis (RA) patients. In addition, biopsies were taken from five control subjects. We observed the expression of the chemokines CXCL8, CXCL9, CXCL10, CCL2 and CCL4 in CI and CU joints of RA patients. In particular, CXCL8 protein levels were specifically increased in CI joints compared with CU joints, which was confirmed by immunohistochemistry and in situ hybridization.

Quantification of the cell infiltrate in synovial tissue by digital image analysis.

OBJECTIVE: The experience with digital image analysis (DIA) in the assessment of synovial inflammation is limited. In this study we compared DIA with two currently applied methods for the evaluation of the synovium. METHODS: Synovial tissue (ST) specimens were obtained by arthroscopy from knee joints of rheumatoid arthritis (RA) patients with variable disease expression and in control subjects. CD68+ macrophages and CD3+ T-cells were detected by immunohistochemical staining using two different labelling techniques. The labelled ST sections were quantified using three different analysis techniques: manual cell counting (MC), semi-quantitative analysis (SQA) and DIA. RESULTS: We observed strongly positive correlations between the three methods when examining T-cell infiltration: between MC and SQA: sigma=0.91 (P<0. 0001), between MC and DIA: sigma=0.95 (P<0.0001), and between SQA and DIA: sigma=0.83 (P<0.0001). Similarly, for the analysis of synovial intimal macrophages, positive correlations were noted between MC and SQA (sigma=0.62; P=0.002), MC and DIA (sigma=0.56; P<0.01), and SQA and DIA (sigma=0.79; P<0.0001). Finally, the analysis of synovial sublining macrophages revealed positive correlations between MC and SQA (sigma=0.95; P<0.0001), MC and DIA (sigma=0.64; P=0.001) and SQA and DIA (sigma=0.69; P<0.0001). CONCLUSION: These three different methods generated similar results. DIA offers the opportunity of a reliable and time-efficient analysis of the synovial infiltrate.

Immunohistological analysis of synovial tissue for differential diagnosis in early arthritis.

OBJECTIVE: An early diagnosis in patients presenting with arthritis is important to provide information about prognosis and to initiate treatment. The objective of this study was to determine which markers applied in immunohistological analysis of synovial tissue (ST) specimens could be used to differentiate rheumatoid arthritis (RA) from other forms of arthritis. METHODS: Synovial biopsies were obtained by blind needle techniques from 95 patients with early arthritis. After follow-up of at least 2 yr to verify the diagnosis, the patients could be classified as follows: RA (n=36), undifferentiated arthritis (UA; n=21), osteoarthritis (OA; n=17), reactive arthritis (ReA; n=10), ankylosing spondylitis (AS; n=3), psoriatic arthritis (PsA; n=2) and crystal-induced arthritis (CA; n=6). ST sections were analysed by immunohistochemistry using monoclonal antibodies against CD3, CD4, CD8, CD22 (B cells), CD38 (plasma cells), CD68 (macrophages) and CD55 (fibroblast-like synoviocytes). RESULTS: Logistic regression analysis revealed that the higher scores for the numbers of CD38+ plasma cells and CD22+ B cells in RA were the best discriminating markers comparing RA to non-RA patients (CD38: P=0.0001; CD22: P<0.05). Polychotomous regression analysis comparing three diagnostic categories (1: RA; 2: UA, ReA, AS and PsA; 3: OA and CA) also identified the score for the number of CD38+ plasma cells (P<0.0001) as well as the numbers of CD68+ macrophages in the synovial sublining (P=0.05) as discriminating markers. CONCLUSION: The results suggest that immunohistochemical analysis of ST specimens from early arthritis patients can be used to differentiate RA from non-RA patients. The numbers of plasma cells, B cells and macrophages are especially increased in ST of patients with RA. Future studies in early arthritis patients with clinical features which do not allow an immediate confident diagnosis may clarify the role of this test system in differential diagnosis.