[Show abstract][Hide abstract] ABSTRACT: Patients presenting with RA before the age of 45 years (younger onset) are known to have more aggressive disease compared with patients presenting after the age of 65 years (older onset). Coordinated expansion of circulating CD5+ B cell and TCR γδ+ T cell levels has been reported in patients with RA. This study assesses the peripheral blood levels of these two cell types in RA patients with younger and older onset of disease. CD5+ B cell levels were significantly elevated in the younger onset RA group (26±6 γδ+4±5%) compared with the older onset RA group (14±2 γδ+1±2%; P
[Show abstract][Hide abstract] ABSTRACT: CXCL13 and CCL21 have been functionally implicated in lymphoid tissue organization both in the upstream phases of lymphoid tissue embryogenesis and in ectopic lymphoid neogenesis in transgenic mice. Here, we analyzed the relationship between CXCL13 and CCL21 production and lymphoid tissue organization in rheumatoid synovitis as a model of a naturally occurring ectopic lymphoneogenesis. Through systematic analysis of mRNA and protein expression, we defined the microanatomical relationship between CXCL13 and CCL21 in progressive aggregational and structural phases of synovial inflammatory infiltrate. We provide the first direct in situ evidence that production of CXCL13 and CCL21 (rather than simply protein binding) is associated with inflammatory lymphoid tissue formation and development with the demonstration, in organized aggregates, of a secondary lymphoid organ-like compartmentalization and vascular association. Notably, the presence of CXCL13 and CCL21 (protein and mRNA) was also demonstrated in non-organized clusters and minor aggregational stages, providing evidence that their induction can take place independently and possibly upstream of T-B compartmentalization, CD21(+) follicular dendritic cell network differentiation and germinal center formation. Our data support the concept that, under inflammatory conditions, CXCL13 and CCL21 participate in lymphoid tissue microanatomical organization, attempting to recapitulate, in an aberrant lymphoid neogenetic process, their homeostatic and morphogenetic physiologic functions.
No preview · Article · Jun 2005 · European Journal of Immunology
[Show abstract][Hide abstract] ABSTRACT: Adhesion mechanisms play a major role in the recruitment of peripheral blood lymphocytes (PBL) which characteristically infiltrate rheumatoid arthritis (RA) synovium and other chronically inflamed tissues. Through a sequential series of complex integrated adhesion and signalling events, 'multistep model of migration', specific subsets of PBL are recruited into inflamed tissues. In this process both leucocyte receptors and microvascular endothelial (MVE) counter-receptors play a critical role. The MVE in particular, during an inflammatory state, is the target of various inflammatory mediators that cause the up-regulation of several cell adhesion molecules (CAM). One of the most important factors known to be a powerful inducer of MVE CAM is TNF-alpha. Conversely, blocking TNF-alpha causes a down-modulation of CAM expression. To test directly the capacity of TNF-alpha to induce cell migration into RA synovium we adapted a model in which synovial grafts were implanted into SCID mice subcutaneously. Using this model we demonstrate that: (i) transplants remain viable and become vascularized and fed by mouse subdermal vessels; (ii) the mouse vasculature connects to the transplant vasculature which maintains the ability to express human CAM; (iii) intragraft injections of TNF-alpha up-regulate the expression of human CAM, following the down-regulation which occurred 4 weeks post-transplantation; and (iv) the up-regulation of graft CAM is associated with increased human PBL migration into the transplants. This study provides direct evidence in vivo of the capacity of TNF-alpha to induce cell migration. In addition, it provides the experimental background for the optimal use of this model.
[Show abstract][Hide abstract] ABSTRACT: Constitutive differences between individuals in cytokine production may determine the variation in the course of inflammatory arthritis.
The association between interleukin 10 (IL-10) production and joint destruction was studied by comparing IL-10 mRNA content in synovial biopsies from seven patients with destructive joint disease and six patients with non-destructive joint disease. The IL-10 mRNA content was 0.4 +/- 0.6 arbitrary units in erosive joints compared with 2.3 +/- 1.2 arbitrary units in non-erosive joints (P: < 0.03, Mann-Whitney U:-test). As this difference suggested that IL-10 production was associated with joint destruction, we tested whether the IL-10 locus determined the extent of joint damage.
Innate differences in IL-10 production are locus-dependent. In line with these data, we showed that innate differences in IL-10 protein production were also present as differences in IL-10 mRNA levels. We tested if polymorphisms in the promoter of IL-10 were associated with the extent of joint damage.
In a cohort study of female rheumatoid arthritis patients followed for 12 yr, the extent of joint destruction differed significantly between patients with different IL-10 genotypes. In patients with the -1082AA genotype who were studied prospectively, the mean increase in radiographic damage score (modified Sharp score of X-rays of hands and feet) during the first 6 yr was 9 +/- 9 per yr vs 19 +/- 16 per yr for patients with the genotype -1082GG (P: < 0.02). In line with these data, cultures of endotoxin-stimulated whole blood from 158 donors showed that the presence of the allele associated with less joint destruction correlated with slightly higher IL-10 production.
Both the immunogenetic and the synovial biopsies suggest that a variation in IL-10 production is associated with joint destruction.
[Show abstract][Hide abstract] ABSTRACT: CD1 is a novel class of molecules which present non-protein antigens to T cells. The objective of this study was to evaluate the expression of CD1 in the skin and synovium of patients with psoriatic arthritis (PsA) in comparison with rheumatoid arthritis (RA) and osteoarthritis (OA).
Paired lesional skin (SK) and synovial membrane (SM) from four PsA patients, paired SK and SM from four RA patients, SM from eight RA and eight OA patients, and normal SK from four volunteers were studied using standard immunohistochemistry.
In all PsA and RA skin samples CD1-positive cells were abundantly detected both in the dermis and in the epidermis. However, in the 24 SM examined CD1-positive cells were rarely found. In one patient only with RA, a few CD1a-positive cells were found in the SM. CD1b was scarcely expressed in the lining layer (LL) of five SM and in very few cells in the sublining layer (SL) of 11 SM. CD1c was rarely expressed in the LL of six SM and in very few cells in the SL of 13 SM.
The paucity of CD1 in the PsA and RA synovium suggests that different subsets of antigen-presenting cells are involved in the pathogenesis of dermatitis and synovitis, respectively.
[Show abstract][Hide abstract] ABSTRACT: Previous studies have used various techniques for microscopic analysis of rheumatoid synovium, ranging from rapid analysis of limited areas of tissue to detailed quantification of extensive areas. The sensitivity and reproducibility of these methods have not been tested. This study sought to determine the minimum area of rheumatoid synovium needed to allow accurate microscopic analysis of synovial inflammation. Multiple synovial tissue samples were obtained from patients with rheumatoid arthritis at knee arthroplasty (n = 10), knee arthroscopy (n = 10) and by blind needle biopsy (n = 23). Lining layer thickness, sublining T-cell infiltration and vascularity were measured in all high-power fields (hpf) throughout every sample obtained from each patient. These complete measured results were compared with estimated results from limited numbers of hpf from each patient. It was observed that lining layer thickness estimated from as few as five readings from 3 samples/patient correlated significantly with the measured results obtained from as many as 85 readings/patient [Tau (T) = 0.70-0.94 for the three groups, all P < or = 0.005). Estimated measures of T-cell infiltration and vascularity derived from only 17 randomly selected hpf from 3 samples/patient (equivalent to 1 mm2) correlated significantly with the measured results obtained from up to 150 hpf/patient (T = 0.65-0.94, all P < or = 0.002). Quantitative analysis of inflammation in synovial tissue samples is both accurate and practical when restricted to an evaluation of a limited number of microscopic fields. It is proposed that lining layer thickness may be confidently quantified from five randomly selected readings from three tissue samples, and that sublining T-cell infiltration and vascularity may be quantified from 17 randomly selected hpf from the same samples.
Full-text · Article · Jul 1998 · British journal of rheumatology
[Show abstract][Hide abstract] ABSTRACT: To determine whether interleukin-1 (IL-1) and interleukin-1 receptor antagonist (IL-1ra) are produced by different macrophage
subsets, we applied immunoperoxidase and double-labelling immunofluorescence techniques to 10 rheumatoid arthritis (RA) and
10 osteoarthritis (OA) synovial membranes. In RA, greater numbers of early 27E10+ macrophages were found in the sublining
layer while mature 25F9+ macrophages were more abundant in the lining layers. The majority of IL-1alpha+ cells were also IL-1ra+
(79 +/- 12% sublining layer, 98 +/- 2% lining layer). In OA sublining layer, a higher percentage of cells double stained for
25F9 and IL-1ra was detected compared to those double stained for 25F9 and IL-1alpha (P < 0.004). In OA, 25F9+ macrophages
demonstrated a lower percentage of IL-1alpha+ in the lining and sublining layers compared to RA (P < 0.02 and P < 0.004, respectively).
It may be concluded that once monocytes have migrated into the RA joint, they undergo phenotypic and functional changes from
an early profile (27E10+, CD14+, low percentage of IL-1+ and IL-1ra+ cells) to a mature profile (CD14+/-, 25F9+, RM3/1+, high
percentage of IL-1+ and IL-1ra+ cells).
Preview · Article · Sep 1997 · British journal of rheumatology
[Show abstract][Hide abstract] ABSTRACT: The objective was to elucidate the immunological abnormalities underlying polymyositis (PM) and dermatomyositis (DM). The phenotype of peripheral blood mononuclear cell (PBMNC) subsets and cell surface expression of activation (CD25 and HLA-DR) and adhesion (LFA-1) molecules was studied in 12 patients with PM and in 10 patients with DM. PBMNC subsets and expression of T-cell activation molecules were evaluated by cytofluorography. Double immunofluorescence and indirect immunoperoxidase techniques were applied to muscle biopsies to define T-cell phenotype and LFA-1/ICAM-1 expression. In PM, the absolute number of circulating cytotoxic (CD8+CD28+) T cells was selectively reduced. T cells showed increased expression of activation molecules, CD25 and HLA-DR, and increased adhesion capacity as the absolute numbers of CD3+CD25+, CD8+HLA-DR+, CD3+LFA-1+('bright') and CD8+ CD8+LFA-1+('bright') cells were higher than in healthy donors and DM patients. In PM muscle biopsies, T cells were mainly CD3+CD8+ and LFA-1+; additionally, endothelial cells and myofibres surrounded by T cells showed positive staining for ICAM-1. In DM, there was a general lymphopenia that led to a decreased absolute number of all T-lymphocyte subsets. It is proposed that in PM, in contrast to DM, LFA-1/ICAM-1 interactions enable activated CD8+ T cells to migrate selectively into the inflamed muscle and to adhere to myofibres, leading to tissue injury.
Full-text · Article · Oct 1996 · British journal of rheumatology
[Show abstract][Hide abstract] ABSTRACT: Patients presenting with RA before the age of 45 years (younger onset) are known to have more aggressive disease compared with patients presenting after the age of 65 years (older onset). Coordinated expansion of circulating CD5+ B cell and TCR gamma delta+ T cell levels has been reported in patients with RA. This study assesses the peripheral blood levels of these two cell types in RA patients with younger and older onset of disease. CD5+ B cell levels were significantly elevated in the younger onset RA group (26.6+/-4.5%) compared with the older onset RA group (14.2+/-1.2%; P<0.01). TCR gamma delta+ T cell levels were also significantly raised in the young patients (4.0+/-0.9%) compared with elderly patients (1.6+/-0.2%; P<0.01). T cell levels (CD3+) were similar in both groups (young 66.4+/-3.3%; old 74.3+/-3.4% (mean+/-s.e.m.); NS). Total B cell levels (CD19+) were also similar in these groups (7.7+/-0.7% versus 8.9+/-1.8%; NS). A significant positive correlation was observed between the CD5+ B and TCR gamma delta+ T cell types in the patients (r=0.72, P<0.05). Compared with age-matched normal controls, the younger onset patients had similar CD5+ B cell and TCR gamma delta+ T cell levels to the elderly controls (CD5+ B cells 30.2+/-3.0%; TCR gamma delta+ T cells 3.0+/-0.8%). Conversely, older onset RA patients had CD5+ B cell levels similar to the young controls (12.3+/-1.9%). Spontaneous in vitro synthesis of immunoglobulins (IgM, IgA and IgG) and rheumatoid factors (IgM and IgA isotypes) were not significantly different in both patient groups. The coordinate expansion of circulating CD5+ B cells and gamma delta+ T cells seen in patients with RA presenting before 45 years of age and not after 65 years of age may suggest a potential role for these cells in more aggressive disease states.
[Show abstract][Hide abstract] ABSTRACT: To investigate whether T cell migration into different sites of inflammation (skin and synovium) within the same individual is principally regulated by tissue-specific homing or by more general mechanisms related to inflammation.
Expression of cutaneous lymphocyte antigen (CLA) and its ligand, E-selectin, was analyzed by immunohistochemistry and immunofluorescence using paired skin and synovial membrane (SM) samples from patients with psoriatic arthritis (PsA). To investigate disease specificity, delayed-type hypersensitivity (DTH) skin lesions, induced by tuberculin purified protein derivative, and SM from patients with rheumatoid arthritis (RA), were studied as controls. To directly examine cell migration in in vivo, the proportion of CLA+ T lymphocytes migrating into suction-induced skin blisters was assessed by flow cytometry. Using the same technique, levels of paired peripheral blood and synovial fluid (SF) T cells were also analyzed.
CLA+ T cells preferentially accumulated in the skin, but not in the joint, of patients with PsA. Similarly, CLA+ T lymphocytes predominated in the DTH skin lesions of RA patients, but were very rare in the SM of RA patients, and were scarcely represented in the SF of patients with several chronic inflammatory arthropathies. In addition, CLA+ T lymphocytes preferentially migrated into epidermal skin blisters. This preferential pattern of CLA+ T cell accumulation was not related to the selective expression of E-selectin, since this was similar in the skin and SM of both PsA and RA patients.
The distinct pattern of T cell infiltration into sites of inflammation within the skin and synovium is regulated by both organ-specific homing and general inflammation-related mechanisms.
No preview · Article · Feb 1996 · Arthritis & Rheumatology
[Show abstract][Hide abstract] ABSTRACT: Rheumatoid arthritis (RA) is a chronic inflammatory disease characterised by a CD4 T cell infiltrate in the synovial membrane (SM). Two subpopulations of T cells, differentiated on the basis of the cytokine pattern produced have been identified. Thl typified by IL-2 and IFN-g are critical to cell-mediated immunity and Th2 typified by IL-4 and IL-10 are important in humoral immunity. The aim of this study was to measure the steady state RNA levels of these Thl and Th2 cytokines in die peripheral blood (PB), synovial fluid (SF) and SM compartments of 14 RA patients with long standing disease. For this purpose, a semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR) method was applied using GAPDH as a housekeeping gene. Variable amounts of cytokine mRNAs were detected in the PB, SF and SM. IL-4 was lower than the other three cytokines in the SF (p<0.02) and SM (P <0.01). Furthermore IL-4 was detected in 50% of SF and SM compared to 93% of PB. IL-10 SF mRNA levels correlated with the SM levels (r=0.7, p<0.04). The results show at IL-2, IFN-g, IL-4 and IL-10 mRNA are detected in the PB, SF and SM of RA patients with levels of IL-4 being the lowest. Furthermore, there is no clear-cut Thl or Th2 mRNA cytokine pattern.
[Show abstract][Hide abstract] ABSTRACT: To study the effect of chimeric anti-CD4 monoclonal antibody (MAb) therapy on synovial inflammation, in order to interpret the clinical experience with anti-CD4 treatment.
The immunohistologic features of synovial biopsy specimens before and 4 weeks after anti-CD4 MAb (cM-T412) therapy were studied in patients with rheumatoid arthritis. The patients received intravenous doses of either placebo (n = 1) or 10 mg (n = 4), 25 mg (n = 2), or 50 mg (n = 1) of cM-T412 daily for 5 consecutive days.
Although the patients did not experience clinical improvement, significant decreases in the number of circulating CD4+ cells, the degree of synovial inflammatory infiltration, and the mean scores for expression of adhesion molecules were found in the 7 patients 4 weeks after receiving cM-T412. The scores for infiltration with CD4+ and other inflammatory cells were particularly reduced following treatment with either 25 mg or 50 mg cM-T412. Cytokines, such as interleukin-1 beta and tumor necrosis factor alpha, could still be detected in the synovial tissue after treatment.
The decline in the numbers of inflammatory cells and adhesion molecules in synovial tissue after CD4+ cell depletion supports the view that CD4+ T cells orchestrate local cellular infiltration. The lack of clinical effect of anti-CD4 therapy might be explained by an insufficient decrease in the number of synovial CD4+ cells and by the persistence of cytokines. Determination of whether more adequate dosing would lead to a clinical improvement must await further study.
No preview · Article · Oct 1995 · Arthritis & Rheumatology
[Show abstract][Hide abstract] ABSTRACT: To investigate the role of cytokines and cell adhesion molecules in the pathogenesis of Sjögren's syndrome (SS).
Using an indirect immunoperoxidase technique we assessed the expression of the cytokines interleukin-1 alpha (IL-1 alpha), interleukin-1 beta (IL-1 beta), interleukin-8 (IL-8), transforming growth factor beta (TGF beta) and granulocyte macrophage colony stimulating factor (GM-CSF), of the adhesion molecules intercellular adhesion molecule-1 (ICAM-1), lymphocyte function associated antigen-1 (LFA-1), the activated molecular form of LFA-1 (NKI-L16), CD2, and LFA-3, and of a panel of cellular markers in the minor salivary glands.
In SS and chronic sialoadenitis (CS), the ductal epithelial cells and acini expressed all the cytokines examined. The percentage of glandular mononuclear cells which stained positive for cytokines did not differ significantly between SS and CS. NKI-L16 was detected on 33.6 (SD 10.1)% and 15.3 (4.3)% of LFA-1 cells in SS and CS, respectively (p < 0.002).
SS and CS did not differ in the pattern of cytokines examined. The characteristic cell clustering seen in the salivary glands in SS may be caused by the upregulation of NKI-L16.
Preview · Article · Mar 1995 · Annals of the Rheumatic Diseases
[Show abstract][Hide abstract] ABSTRACT: Cytokines, released from mononuclear cells (MNC) are mediators of joint destruction in rheumatoid arthritis (RA). The mechanisms of action of gold salts used in the treatment of RA are unknown. The aim of this study was to investigate cytokine expression and intensity of MNC infiltrate in the RA synovial membrane (SM) following treatment with sodium aurothiomalate (SAT).
Sequential blind needle biopsies were obtained at entry into the study and at two and 12 weeks after the start of SAT therapy in 10 patients with active RA. SMs were stained with a panel of monoclonal antibodies to assess cytokine expression (IL-1 alpha, IL-1 beta, TNF-alpha, IL-6, and GM-CSF).
There was a significant decrease in IL-1 alpha, IL-1 beta, IL-6 and TNF-alpha expression 12 weeks after treatment (p < 0.004, p < 0.002, p < 0.009 and p < 0.004 respectively). This was noted in the lining layer, the perivascular aggregates and the connective tissue areas. Detailed examination of the MNC infiltrate showed a significant reduction in inflammatory monocytes (MONO) in the lining layer at two weeks (p < 0.03). A decrease in the number of CD68+ macrophages (MAC) was noted in the perivascular and connective tissue areas at 12 weeks. No significant changes were observed in the number of T and B cells and blood vessels.
The results suggest that gold may suppress RA disease activity by diminishing MONO and MAC numbers and consequently monokine production in the SM.
Full-text · Article · May 1994 · Annals of the Rheumatic Diseases
[Show abstract][Hide abstract] ABSTRACT: To analyse the mononuclear cell populations in synovial membrane biopsies obtained before treatment from patients with rheumatoid arthritis (RA) and to correlate the findings with the degree of joint damage occurring over one year.
Multiple needle biopsy specimens were obtained from inflamed knee joints on entry to the study. The tissue samples were examined using immunohistochemical techniques. The degree of joint damage was estimated using the Larsen radiological index.
Twelve patients were studied. It was observed that there was a significant correlation between the number of synovial tissue macrophages and the degree of joint erosion over one year (r = 0.66; p = 0.04). The synovial lining layer contained large numbers of macrophages and the cellularity of the lining layer correlated significantly with the number of macrophages infiltrating the sublining areas (r = 0.65; p = 0.01). Finally, the cellularity of the lining layer correlated with the synovial fluid levels of interleukin-6 (r = 0.66; p = 0.04). The radiological course did not correlate with infiltrating T or B lymphocyte populations, but did correlate with other previously identified indicators of the clinical course, including a high index of disease activity and IgA rheumatoid factors levels.
This study suggests that synovial tissue macrophages play a critical role in the pathogenesis of joint erosion in RA.
Preview · Article · Feb 1994 · Annals of the Rheumatic Diseases
[Show abstract][Hide abstract] ABSTRACT: To compare, by immunohistochemistry, the cellular and cytokine profile in rheumatoid arthritis (RA) and osteoarthritis (OA) synovial membranes (SMs). Synovium was obtained at knee arthroplasty from 10 patients with RA and 10 with OA.
Synovial membranes were stained with a panel of monoclonal antibodies (MAb) to assess cytokine expression (IL-1 alpha, IL-1 beta, IL-6, GM-CSF, TNF-alpha and EGF) and the intensity of the mononuclear cellular infiltrate (MNC).
Significantly greater percentages of IL-1 alpha, IL-1 beta, IL-6, TNF-alpha, GM-CSF and EGF cells were detected in all areas of the rheumatoid SMs when compared with osteoarthritic SMs. Five RA but only one OA SM demonstrated focal lymphoid aggregates. Lining layer thickening was noted in RA SMs only. The intensity of the MNC and number of blood vessels were greater in the RA group.
The results suggest that the differences in cytokine production by RA and OA SMs are quantitative but that the greater thickness of the synovial lining layer and higher vascularity may be specific to RA.
Full-text · Article · Jan 1994 · Annals of the Rheumatic Diseases
[Show abstract][Hide abstract] ABSTRACT: Avidin-biotin (AB) systems are commonly employed to investigate salivary gland sections either in immunohistochemistry or in immunofluorescence techniques. We noted non-specific staining in the ductal epithelium of minor salivary gland (SG) sections from 5 Sjögren's syndrome (SS), 5 chronic sialoadenitis (CS), and one normal parotid gland (NP) when incubated with AB peroxidase complex. Inhibition of endogenous peroxidase did not prevent the staining while saturation of supposed biotin-like molecules in the tissue with added avidin led to the loss of this non-specific staining. These results demonstrate the presence of endogenous avidin binding activity (EABA) in the epithelial cells of salivary gland ducts. We suggest that the avidin and streptavidin systems should not be used for immunohistological examination of the ductal epithelium of salivary glands.
No preview · Article · Jan 1994 · Clinical and experimental rheumatology
[Show abstract][Hide abstract] ABSTRACT: To assess parent-child agreement of child disability and pain.
Twenty children with juvenile chronic arthritis aged between 8 and 16 years and their mothers were assessed using a recently developed measure of child disability and pain.
Results demonstrated a high level of agreement between children and their mothers with respect to disability. However, there was no correlation between the children's and their mothers' assessment of pain.
This suggests that parents of children with chronic arthritis give accurate information regarding disability, but not regarding pain.
No preview · Article · Oct 1993 · The Journal of Rheumatology