bluuebunny/crossref_metadata_2025 huggingface dataset:
The effect of methotrexate on mouse bone cells in culture
Authors: May Kimberly P. , Mercill Don , McDermott Michael T. , West Sterling G.
Published: February 2007

Objective. We have recently shown that administration of long‐term, low‐dose methotrexate (MTX) causes severe osteopenia in female rats. This osteopenia is characterized both by decreased osteoblast function without a decrease in osteoblast numbers, and by increased bone resorption that is believed to represent a physiologic remodeling response by osteoclasts. The present study investigates the effects of varying doses of MTX on mouse bone cells in culture.

Methods. Cells were obtained by sequential digestion of neonatal mouse calvariae, and cultured with fetal calf serum (10% for osteoblast‐like cells and 2% for osteoclast‐like cells). After 1 week, MTX was added to each culture in concentrations of 0.6 μM, 0.4 μM, 0.2 μM, 0.1 μM, 1 nM, and 0.5 nM. All experiments were done on 24 wells for each MTX concentration and for the controls. The effect on osteoblastic cells was assessed, at 7 days, by cell counts and by measurement of lysate alkaline phosphatase and supernatant osteocalcin levels, and, at 21 days, by analysis of the calcified matrix production, which was cultured with ascorbic acid and β‐glycerophosphate. For osteoclastic cells, cell count and lysate acid phosphatase levels were determined.

Results. Levels of osteoblastic cells and lysate alkaline phosphatase were not changed by any of the concentrations of MTX. Matrix calcification and supernatant osteocalcin levels were diminished by MTX in a dose‐responsive manner. Osteoclast‐like cell numbers and acid phosphatase levels were not significantly affected by MTX.

Conclusion. These results suggest that diminished mouse osteoblastic cell function occurs with very low mean concentrations of MTX, in a dose‐responsive manner. The mechanism seems to be inability of the cell to synthesize and calcify matrix, possibly through defective osteocalcin production. Thus, low‐dose MTX may have an important impact on bone density by slowing osteoblastic matrix production.
The effect of leukotriene synthesis inhibitors in models of acute and chronic inflammation
Authors: Nickerson‐Nutter Cheryl L. , Medvedeff Eugene D.
Published: February 2007

Objective. To assess the efficacy of leukotriene synthesis inhibitors, alone and in combination with a nonsteroidal antiinflammatory drug, as potential treatments for rheumatoid arthritis (RA), using the mouse air pouch model and the collagen‐induced arthritis (CIA) model.

Methods. Two selective leukotriene synthesis inhibitors, Bay x 1005 and Bay y 1015, were compared with zileuton in terms of their ability to decrease exudate volume, cell infiltration, and leukotriene B4 (LTB4) production in response to zymosan injection in the mouse air pouch model. The mouse CIA model was used to assess the effect of leukotriene synthesis inhibitors in a model of chronic inflammation. Bay y 1015 and Bay x 1005, and the cyclooxygenase inhibitor naproxen, were evaluated individually and in combination, for their antiarthritic potency in the mouse CIA model.

Results. The results indicate that neither zileuton, Bay x 1005, nor Bay y 1015 inhibited exudate production. All 3 compounds decreased LTB4 levels in the air pouch, with Bay y 1015 being the most effective. Cell infiltration was significantly decreased with Bay x 1005, but the degree of this decrease did not appear to correlate with LTB4 levels. No inhibition of arthritis was observed with any compound administered alone. In contrast, a significant inhibition of CIA was observed in animals that received both naproxen and either Bay y 1015 or Bay x 1005.

Conclusion. Inhibitors of both cyclooxygenase and leukotriene synthesis in combination may be a more effective treatment of RA than either class of inhibitors alone.
TSG‐6 expression in human articular chondrocytes: Possible implications in joint inflammation and cartilage degradation
Authors: Maier Rainer , Wisniewski Hans‐Georg , Vilcek Jan , Lotz Martin
Published: February 2007

Objective. The hyaluronan‐binding protein TSG‐6 (tumor necrosis factor‐stimulated gene 6) forms a stable complex with the serine protease inhibitor, inter‐α‐inhibitor, potentiates the inhibition of plasmin activity, and has antiinflammatory effects in vivo. This study examines the expression of TSG‐6 in human articular chondrocytes and cartilage.

Methods. Human articular chondrocytes and cartilage explants were stimulated with cytokines, growth factors, and other agents. TSG‐6 expression was analyzed by imaging‐assisted Northern and Western blotting.

Results. TSG‐6 messenger RNA (mRNA) expression was upregulated by cytokines and growth factors, predominantly interleukin‐1β (IL‐1β), tumor necrosis factor α (TNFα), platelet‐derived growth factor AA (PDGF‐AA), and transforming growth factor β1 (TGFβ1). TSG‐6 mRNA induction by TGFβ1 was delayed as compared with IL‐1β. Treatment of the cells with the glucocorticoid dexamethasone neither induced TSG‐6 mRNA nor did it affect IL‐1β‐induced transcript levels. TSG‐6 mRNA induction may involve several signal transduction pathways. The strong transcriptional stimulation by phorbol myristate acetate suggests protein kinase C (PKC)‐mediated signaling. In contrast, PKA‐ and Cadependent signals are only marginally involved as messengers leading to increased TSG‐6 levels after IL‐1β and TNFα treatment. In chondrocyte and cartilage organ cultures, both free TSG‐6 (35 kd) and the complex with inter‐α‐inhibitor (120 kd) were present and upregulated by IL‐1β, TNFα, or TGFβ1.

Conclusion. Chondrocytes are a source of TSG‐6 which may play a role in cartilage remodeling and joint inflammation.
Stimulation of TIMP‐1 production by oncostatin m in human articular cartilage
Authors: Nemoto Osamu , Yamada Harumoto , Mukaida Masahiro , Shimmei Masayuki
Published: February 2007

Objective. To investigate the effects of the interleukin‐6 (IL‐6) family cytokines, such as IL‐6, IL‐11, leukemia inhibitory factor (LIF), and oncostatin M (OSM), on the production of tissue inhibitor of matrix metalloproteinases 1 (TIMP‐1) in human articular cartilage.

Methods. Effects of IL‐6 family cytokines and growth factors on TIMP‐1 production were studied in human articular chondrocytes, grown as monolayer and cartilage explant culture. TIMP‐1 protein levels in the cultured medium were measured by sandwich enzyme immunoassay. TIMP‐1 messenger RNA levels in the cultured chondrocytes were analyzed by Northern blotting. Western blot analysis was performed to evaluate the release of matrix metalloproteinases (MMPs) in the cultured medium. Cell proliferation of chondrocytes was determined by 3H‐thymidine uptake.

Results. IL‐6 family cytokines induced TIMP‐1 expression in monolayer and explant culture, and the production of TIMP‐1 was most stimulated by OSM. In contrast, OSM did not modulate the release of MMPs and cell proliferation.

Conclusion. These results suggest that OSM may be characterized as one of the chondroprotective mediators in cartilage destruction.
Activation of the transcription factor nuclear factor‐κB in human inflamed synovial tissue
Authors: Marok R. , Winyard P. G. , Coumbe A. , Kus M. L. , Gaffney K. , Blades S. , Mapp P. I. , Morris C. J. , Blake D. R. , Kaltschmidt C. , Baeuerle P. A.
Published: February 2007

Objective. The transcription factor–κB (NF‐κB) has been implicated in the inflammatory response and is known to be activated by a process involving reactive oxygen intermediates. The purpose of the present study was to demonstrate the presence and distribution of activated NF‐κB in synovium samples from patients with rheumatoid arthritis (RA) and osteoarthritis (OA) and from autopsy subjects with no known history of arthritis.

Methods. Immunohistochemical staining was performed using both polyclonal and monoclonal “activity‐specific” antibodies to the Rel‐A (p65) subunit of NF‐κB (anti‐Rel‐A nuclear location sequences). Histologic features of inflammation were also scored.

Results. Both antibodies demonstrated positive staining of synovial tissue, with a cellular distribution that was nuclear. The staining was associated with specific cell types within the tissue, in particular, type A synoviocytes and vascular endothelium. Notably, lymphoid aggregates were unstained. Using the monoclonal antibody, a further study was carried out to investigate the distribution of staining in tissues from patients with different disease activities and clinical diagnoses, as well as in normal control tissue obtained at autopsy. Patients with acute RA more commonly showed vessel staining (P = 0.05) and, conversely, showed less frequent staining of the synovial lining (P < 0.005) compared with OA patients. Synovial tissue from controls exhibited either no staining or only weak staining in the synovial lining.

Conclusion. The activation of NF‐κB in vascular endothelium and type A synovial lining cells is a feature of synovial tissue from both RA and OA patients. The distribution of this staining appears to be related to the clinical diagnosis.
Reduction in long‐term disability in patients with rheumatoid arthritis by disease‐modifying antirheumatic drug–based treatment strategies
Authors: Fries James F. , Williams Catherine A. , Morfeld Dianne , Singh Gurkirpal , Sibley John
Published: February 2007

Objective. Therapeutic strategies for rheumatoid arthritis (RA) have been evolving from the traditional “pyramid” approach toward one based upon early and sustained use of disease‐modifying antirheumatic drugs (DMARDs), in the hope of improving long‐term health outcomes. However, few data to have been presented to document the effects of this approach. We sought to directly assess associations between consistent DMARD use and long‐term functional outcomes.

Methods. We studied 2,888 RA patients who were followed up prospectively for up to 20 years (average 9 years) at 8 databank centers. The independent variable was the proportion of patient encounters that resulted in treatment with ⩾1 DMARD (hydroxychloroquine, sulfasalazine, auranofin, intramuscular gold, D‐penicillamine, methotrexate, and/or azathioprine). The dependent variable was each patient's last recorded Disability Index value from the Health Assessment Questionnaire (HAQ).

Results. Increased DMARD use was strongly associated with better long‐term Disability Index values (P < 0.0001). The association was strengthened when restricted to more seriously affected (rheumatoid factor (RF)–positive) patients. The magnitude of the effect, unadjusted, was a difference of 0.53 HAQ Disability units (scale 0–3) between 100% DMARD use and 0%. Correlation coefficients ranged up to 0.26. Effects were similar for all disease duration periods (0–4, 5–9, 10–14, 15–19, and 20+ years). “Control” correlations, with variables computed to represent the proportion of time in which patients were taking either nonsteroidal antiinflammatory drugs or prednisone, failed to show positive associations. A multiple linear regression model, which controlled for age, disease duration, sex, RF positivity, proportion of visits under a prednisone regimen, and initial disability level, included the proportion of time in which patients were taking DMARDs (P < 0.0001), with a model R2 of 0.54. These results were obtained despite an adverse selection bias in which more severely affected individuals were given DMARDs more frequently, and despite absence of data on drug use early in the disease course of many patients. Thus, these results, which suggest up to a 30% reduction in longterm disability with consistent DMARD use, are most likely conservative.

Conclusion. An association between consistent DMARD use and improvement in long‐term functional outcomes in RA is supported by these data.
A prospective long‐term study of fibromyalgia syndrome
Authors: Kennedy Maura , Felson David T.
Published: February 2007

Objective. To ascertain the long‐term natural history of fibromyalgia syndrome (FMS).

Methods. Patients with a history of FMS, seen in an academic rheumatology referral practice, were originally surveyed soon after onset of symptoms, and then were reinterviewed 10 years later in a prospective followup cohort study. A validated telephone survey was administered that inquired into current symptoms, medical care and treatments used, and work disability. The results were compared with the prior surveys.

Results. Of the original 39 patients, there were 4 deaths. Of the remaining 35 patients, 29 (83%) were reinterviewed. Mean age at current survey was 55 years, and mean duration of symptoms was 15.8 years. All patients had persistence of some fibromyalgia symptoms, although almost half (48%) had not seen a doctor for them in the last year. Moderate to severe pain or stiffness was reported in 55% of patients; moderate to a lot of sleep difficulty was noted in 48%; and moderate to extreme fatigue was noted in 59%. These symptoms showed little change from earlier surveys. In 79% of patients, medications were still being taken to control FMS symptoms. Despite continuing symptoms, 66% of patients reported that FMS symptoms were a little or a lot better than when first diagnosed. Fifty‐five percent of patients said they felt well or very well in terms of FMS symptoms, and only 7% felt they were doing poorly. With the exception of sleep trouble, which was persistent, baseline survey symptoms correlated poorly with symptoms at the 10‐year followup.

Conclusion. FMS symptoms last, on average, at least 15 years after illness onset. However, most patients experience some improvement in symptoms after FMS onset.
Coexistence of two antisynthetases in a patient with the antisynthetase syndrome
Authors: Gelpí Carmen , Kanterewicz Eduardo , Gratacos Jordi , Targoff Ira N. , Rodriguez‐Sanchez José Luis
Published: February 2007

We describe the immunologic findings in a patient with the antisynthetase syndrome characterized by prominent arthritis, lung fibrosis, and subclinical myositis. At disease onset and during the followup, this patient's serum showed 2 different subsets of antisynthetase autoantibodies: anti–Jo‐1, which reacted with histidyl–transfer RNA (tRNA) synthetase by immunoblot and inhibited its enzymatic function; and anti‐OJ, which immunoprecipitated the multi‐enzyme complex of synthetases, and reacted with lysyl‐tRNA synthetase by immunoblot. This is the first report of anti–Jo‐1 and another antisynthetase antibody being found together in the same patient.
Mixed monoclonal cryoglobulinemia and monoclonal rheumatoid factor cross‐reactive idiotypes as predictive factors for the development of lymphoma in primary Sjögren's syndrome
Authors: Tzioufas Athanasios G. , Boumba Dimitra S. , Skopouli Fotini N. , Moutsopoulos Haralampos M.
Published: February 2007

Objective. To prospectively investigate whether mixed monoclonal cryoglobulinemia (MMC) and monoclonal rheumatoid factor (mRF)–associated crossreactive idiotypes (CRI) serve as predictive factors for the development of lymphoma in patients with primary Sjögren's syndrome (SS).

Methods. One hundred three consecutive patients with primary SS were evaluated from 1986 to 1991. In all patients, the amount of cryoglobulin was measured by ultraviolet absorption at 280 nm and 260 nm. The type of cryoglobulinemia was identified by agarose gel electrophoresis, combined with immunofixation. Sera from all patients were evaluated by enzyme‐linked immunosorbent assay, using the corresponding monoclonal or polyclonal antibodies, for the presence of immunoglobulins bearing the idiotypes 17109 (VκIIIb associated), G‐6 (VH1 associated), and 3rd SS (a rabbit polyclonal antibody raised against the Fab fragment of an IgMκ mRF from a patient with primary SS). Data analysis was performed by logistic regression.

Results. Eighteen of the patients with primary SS (17.4%) had MMC during the first evaluation. There was a statistically significant correlation between the presence of MMC and a higher prevalence of autoantibodies to Ro/SS‐A and La/SS‐B, as well as extraglandular manifestations. During a 5‐year period, 7 patients developed lymphoma. Six of the 7 (86%) had MMC before the appearance of lymphoma, compared with 12 of 96 (12.4%) of the remainder (r = 0.421, P < 0.0009). Patients who developed lymphoma had higher amounts of cryoglobulin than those who did not (mean ± SD 53.4 ± 44.7 mg/dl versus 26.8 ± 20.6 mg/dl). CRIs 17109 and G‐6 were also correlated with lymphoma development (r = 0.321, P < 0.006 and r = 0.22, P < 0.03, respectively). For both CRIs, this correlation was dependent on the presence of MMC, since a stepwise multiple comparison analysis revealed that their individual significance was abolished when their correlation with lymphoma in association with MMC was assessed.

Conclusion. The determination of MMC can be used as a laboratory predictive factor for lymphoma development in primary SS. CRIs 17109 and G‐6 may also be used to predict lymphoma development, especially when the monoclonal component is absent.
Increased expression of human thioredoxin/adult T cell leukemia–derived factor in Sjögren's syndrome
Authors: Saito Ichiro , Shimuta Misa , Terauchi Kumiko , Tsubota Kazuo , Yodoi Junji , Miyasaka Nobuyuki
Published: February 2007

Objective. To determine the involvement of human thioredoxi/adult T cell leukemia–derived factor (TRX/ADF) in Sjögren's syndrome (SS) and the correlation with Epstein‐Barr virus (EBV).

Methods. Indirect immunohistochemical techniques and reverse transcriptase polymerase chain reaction were utilized to analyze TRX/ADF expression and the presence of EBV, using 6 normal tissues and 23 surgical specimens. The kinetics of expression of TRX/ADF induced by EBV was examined in vitro with peripheral blood B cells from EBV‐seronegative donors.

Results. Marked expression of TRX/ADF was found in the infiltrating B cells and the epithelial cells of salivary gland tissues from patients with SS (11 of 12 cases), but not in those from patients with other salivary gland inflammatory conditions (0 of 11 cases) or those of normal individuals (0 of 6 cases). In immunohistologic analyses, a striking topographic correlation between TRX/ADF and EBV was found. The coexistence of TRX/ADF messenger RNA and EBV DNA was detected by polymerase chain reaction (r = 0.75, P < 0.01). Peripheral blood B cells from EBV‐seronegative donors showed de novo synthesis of TRX/ADF following in vitro infection with EBV. EBV‐infected B cell lines all expressed TRX/ADF. TRX/ADF was not detected in non‐EBV‐infected cells. Tumors in SCID mice reconstituted with mononuclear cells of salivary glands from SS patients, which were composed of human B cells carrying EBV DNA, were positive for TRX/ADF.

Conclusion. These findings suggest that TRX/ADF expression closely reflects the intracellular event of EBV reactivation in SS. This is also the first report to show the ectopic in vivo expression of TRX/ADF in human autoimmune disease.
Anticytokine treatment of established type II collagen–induced arthritis in DBA/1 mice: A comparative study using anti‐TNFα, anti–IL‐1α/β, and IL‐1Ra
Authors: Joosten Leo A. B. , Helsen Monique M. A. , van de Loo Fons A. J. , van den Berg Wim B.
Published: February 2007

Objective. To examine the role of tumor necrosis factor α (TNFα), interleukin‐1α (IL‐1α), and IL‐1β in collagen‐induced arthritis (CIA), immediately after onset and during the phase of established arthritis.

Methods. Male DBA/1 mice with collagen‐induced arthritis were treated with antibodies against murine TNFα and IL‐1α/β at different time points of the disease. IL‐1 receptor antagonist (IL‐1Ra) was administered using Alzet osmotic minipumps. The effect of anticytokine treatment was monitored by visual scoring. Histology and cytokine reverse transcription polymerase chain reaction (RT‐PCR) analyses were performed at the end of the treatment period.

Results. Anti‐TNFα treatment showed efficacy shortly after onset of the disease, but had little effect on fully established CIA. Histologic analysis after early treatment revealed that anti‐TNFα significantly reduced joint pathology, as determined by infiltration of inflammatory cells and cartilage damage. Anti–IL‐1α/β treatment ameliorated both early and full‐blown CIA. This clear suppression of established arthritis was confirmed by administration of high doses of IL‐1Ra. Dose‐response experiments showed that a continuous supply of 1 mg/day was needed for optimal suppression. Histologic analysis showed markedly reduced cartilage destruction both in the knee and the ankle joints. Autoradiography demonstrated full recovery of chondrocyte synthetic function of articular cartilage. In addition, we found that the IL‐1β isoform plays a dominant role in established CIA. Profound suppression of CIA was observed with anti–IL‐1β, although elimination of both IL‐1α and IL‐1β still gave better protection. Analysis of messenger RNA with RT‐PCR revealed that IL‐1β was highly upregulated in synovium and cartilage at late stages of CIA, whereas anti–IL‐1β treatment markedly reduced IL‐1β message in the synovium.

Conclusion. The present study identified different TNFα/IL‐1 dependencies in various stages of CIA and revealed that blocking of TNFα does not necessarily eliminate IL‐1. Continuous, high doses of IL‐1Ra are needed to block CIA.
An epitope on Ki antigen recognized by autoantibodies from lupus patients shows homology with the sv40 large t antigen nuclear localization signal
Authors: Takasaki Yoshinari , Yano Tetsuro , Hirokawa Kaoru , Takeuchi Ken , Ando Soichiro , Takahashi Toshimitsu , Shimada Kohichiro , Hashimoto Hiroshi
Published: February 2007

Objective. Epitopes on Ki antigen were analyzed using synthetic peptides, including KILT, a 16‐mer peptide with an amino acid sequence homologous to the SV40 large T antigen nuclear localization signal (SV40 T NLS).

Methods. In addition to KILT, 4 synthetic peptides, all potential epitopes on Ki antigen according to computer analysis, were prepared and tested for reactivity with 49 anti‐Ki–positive lupus sera by enzyme‐linked immunosorbent assay.

Results. Eighteen sera reacted with KILT, but not with other peptides. The reaction of anti‐Ki sera with KILT was specifically inhibited by recombinant Ki antigen. Eight of 49 anti‐Ki sera reacted with a 7‐mer synthetic peptide of SV40 T NLS, and the reaction was specifically inhibited by KILT.

Conclusion. The 16‐mer Ki peptide containing the sequence homologous to the SV40 T NLS is one of the antigenic epitopes recognized by anti‐Ki antibodies in lupus sera.
Oligoclonal T cell proliferation in patients with rheumatoid arthritis and their unaffected siblings
Authors: Waase Inge , Kayser Corinna , Carlson Paula J. , Goronzy Jörg J. , Weyand Corelia M.
Published: February 2007

Objective. To analyze whether patients with rheumatoid arthritis (RA) have an intrinsic defect in T cell proliferation and survival, possibly contributing to the infiltration of the synovial membrane with CD4+ T cells.

Methods. Fifteen patients with seropositive RA, 11 patients with psoriatic arthritis, 20 normal controls, and 9 affected and 13 unaffected siblings from 7 multiplex families with RA were analyzed for clonal proliferation. To investigate this clonal T cell proliferation, CD4+ T cells were purified from peripheral blood and synovial fluid by magnetic bead separation. T cell receptor (TCR) β‐chain sequences were amplified by reverse transcriptase–polymerase chain reaction, using TCR BV and BJ gene segment–specific primer sets. Clonally expanded T cell specificities were identified by size fractionation and sequencing of the amplified product.

Results. All RA patients carried clonally expanded CD4+ T cells in the peripheral blood compartment. Such expanded CD4+ T cell clonotypes were only infrequently observed both in normal individuals (P < 0.0001) and in patients with psoriatic arthritis (P = 0.004). Lymphoproliferation of selected CD4+ T cells was shared by affected and unaffected siblings from RA multiplex families (P = 0.005 and P = 0.0003, respectively, compared with normal controls). Expanded clonotypes persisted for several years and contributed to the T cell infiltrate in the joint. Clonal T cell proliferation involved a diverse spectrum of TCR molecules.

Conclusion. RA patients have an abnormality in the homeostasis of CD4+ T cells, characterized by the emergence of clonally proliferating populations. The presence of clonal outgrowth of selected CD4+ T cell specificities in unaffected siblings of RA patients suggests that oligoclonality of CD4+ T cells is inherited and is a risk factor for, rather than a result of, synovial inflammation.
Expression and functional expansion of fibroblast growth factor receptor T cells in rheumatoid synovium and peripheral blood of patients with rheumatoid arthritis
Authors: Byrd Victor , Zhao Xiao‐Ming , McKeehan Wallace L. , Miller Geraldine G. , Thomas James W.
Published: February 2007

Objective. Rheumatoid arthritis (RA) is an inflammatory disorder of the diarthrodial joints, characterized by fibroblast proliferation, angiogenesis, and perivascular CD4+ T cell infiltration. The present study examined the interactions between fibroblast growth factor–1 (FGF‐1) and T cells.

Methods. Synovial tissues from patients with RA or noninflammatory arthritis were examined for the expression of FGF‐1 and its receptor, FGFR‐1, by immunohistology and reverse transcriptase–polymerase chain reaction. Functional assays were used to detect enrichment of FGF‐1–responsive peripheral CD4+ T cells in RA.

Results. FGF‐1 is abundantly expressed by rheumatoid synovium. Enhanced expression of its receptor, FGFR‐1, was found in perivascular CD4+ T cells. In addition, T cells that are activated by FGF‐1 are increased in the peripheral blood of patients with RA, as compared with other inflammatory conditions.

Conclusion. The increased frequency of peripheral T cells that respond to FGF‐1 in RA is consistent with expansion of FGFR‐1‐expressing T cells in the rheumatoid synovium.
Inhibition of synoviocyte collagenase gene expression by adenosine receptor stimulation
Authors: Boyle David L. , Sajjadi Fereydoun G. , Firestein Gary S.
Published: February 2007

Objective. To characterize the regulation of matrix metalloproteinases (MMPs) by adenosine.

Methods. Cultured fibroblast‐like synoviocytes (FLS) were stimulated with interleukin‐1 (IL‐1) in the presence or absence of adenosine receptor agonists. Immunoreactive MMPs were measured using specific enzyme‐linked immunosorbent assays, and gene expression was assessed by Northern blot analysis.

Results. The nonselective adenosine receptor agonist 5′‐N‐ethylcarboxamidoadenosine (NECA) decreased collagenase production by IL‐1‐stimulated synoviocytes from 196 ± 28 ng/ml (mean ± SEM) to 66 ± 9 ng/ml (P < 0.001). There was minimal effect on stromelysin production (decrease from 107 ± 16 ng/ml to 97 ± 15 ng/ml). Selective adenosine receptor agonists implicated the A2b adenosine receptor in this activity, and reverse transcriptase–polymerase chain reaction studies confirmed that FLS express this receptor. Northern blot analysis demonstrated that the mechanism of action was pre‐translational since NECA decreased collagenase, but not stromelysin or tissue inhibitor of metalloproteinases 1 (TIMP‐1), messenger RNA levels. Cyclic AMP levels were increased by NECA, and a direct adenylate cyclase activator (forskolin) also suppressed collagenase gene expression. These data suggest that cAMP mediates the inhibitory effect of NECA on collagenase production.

Conclusion. Stimulation of the A2b receptor on FLS decreases collagenase gene expression, with little or no effect on stromelysin and TIMP‐1. The combination of antiinflammatory and MMP‐regulating properties of adenosine of adenosine‐regulating agents suggests that treatment based on this approach might be useful in rheumatoid arthritis.
Clinical correlations with HLA type in Japanese patients with connective tissue disease and anti–U1 small nuclear RNP antibodies
Authors: Kuwana Masataka , Okano Yutaka , Kaburaki Junichi , Inoko Hidetoshi
Published: February 2007

Objective. To elucidate the roles of HLA genes in the clinical presentation of patients with connective tissue disease and serum anti–U1 small nuclear RNP antibody.

Methods. HLA class I antigens and HLA class II alleles were determined in 43 Japanese patients with anti–U1 RNP antibody alone, by microcytotoxicity testing and DNA typing, respectively. Prospectively recorded clinical and laboratory features were analyzed in relation to HLA class I and class II types.

Results. DQB1*0303 was associated with lupusrelated symptoms including fever, malar rash, oral ulcers, hypocomplementemia, and high‐titer anti–double‐stranded DNA antibody. Other HLA–clinical associations included DR2 with pleuritis, DR4 with hand swelling, and DRB1*0405 with arthritis.

Conclusion. These HLA–clinical associations explain, in part, the heterogeneous clinical presentation in patients with anti–U1 RNP antibody.
Normal expression of type 1 insulin‐like growth factor receptor by human osteoarthritic chondrocytes with increased expression and synthesis of insulin‐like growth factor binding proteins
Authors: Tardif Ginette , Reboul Pascal , Pelletier Jean‐Pierre , Geng Changshan , Cloutier Jean‐Marie , Martel‐Pelletier Johanne
Published: February 2007

Objective. Our previous research demonstrated that, in contrast to normal chondrocytes, human osteoarthritic (OA) chondrocytes were hyporesponsive to stimulation by insulin‐like growth factor 1 (IGF‐1). The aim of the present investigation was to examine whether this finding was due to an alteration in the level of IGF receptors (IGFRs) and/or IGF binding proteins (IGFBP).

Methods. A quantitative reverse transcriptase polymerase chain reaction technique (RT‐PCR) was used to measure the type 1 IGFR messenger RNA (mRNA) level, and Northern blotting was used to measure type 2 IGFR and IGFBP mRNA levels. Western immunoblotting was used to identify and measure IGFBP levels.

Results. There were similar levels of type 1 IGFR mRNA in normal and OA chondrocytes. The level of type 2 IGFR mRNA, in which an increased amount of which can interfere with the biologic effects of IGF‐1, was lower in OA chondrocytes compared with normal chondrocytes. Articular chondrocytes produced IGFBP‐2, IGFBP‐3, and IGFBP‐4, and OA chondrocytes secreted and expressed higher amounts than did normal chondrocytes. There was also an increased level of IGFBP‐3 in the OA chondrocyte lysates. IGFBPs 1, 5, and 6 were not detectable.

Conclusion. OA chondrocytes synthesize and express a larger amount of 3 IGFBPs. This observation, along with a lack of detectable change in type 1 IGFR mRNA level, suggests that the hyporesponsiveness of OA chondrocytes to IGF‐1 might implicate the involvement of IGFBPs in this pathologic process.
Renal vascular damage in systemic sclerosis patients without clinical evidence of nephropathy
Authors: Rivolta Roberto , Mascagni Barbara , Berruti Vittorio , di Palo Filippo Quarto , Elli Attilio , Scorza Raffaella , Castagnone Daniela
Published: February 2007

Objective. To evaluate the use of color‐flow Doppler ultrasonography, a direct, noninvasive technique, for measurement of kidney blood flow in patients with systemic sclerosis (SSc).

Methods. Twenty‐five normal volunteers and 25 SSc patients (median disease duration 8 years, range 2–21 years) were studied. All were free of clinical symptoms of renal damage. The resistance index (RI) was determined on main, interlobar, and cortical vessels.

Results. In SSc patients, the RI was significantly increased at every sampling site examined (P < 0.001). RI values were strongly correlated with disease duration (main artery r = 0.56, P < 0.04; interlobar artery r = 0.63, P < 0.02; cortical artery r = 0.75, P < 0.002). Regression analysis showed no relationship between RI and creatinine clearance values.

Conclusion. Color‐flow Doppler ultrasonography is a sensitive and noninvasive technique for evaluating vascular damage of the kidney in patients with SSc.
Utility of anti‐Sm, anti‐RNP, anti–Ro/SS‐A, and anti–La/SS‐B (extractable nuclear antigens) detected by enzyme‐linked immunosorbent assay for the diagnosis of systemic lupus erythematosus
Authors: Sánchez‐Guerrero Jorge , Lew Robert A. , Fossel Anne H. , Schur Peter H.
Published: February 2007

Objective. To determine the utility of anti‐extractable nuclear antigen (anti‐ENA) antibodies detected by enzyme‐linked immunosorbent assay as a predictor for the diagnosis of systemic lupus erythematosus (SLE).

Methods. Among 2,185 serum samples sent for testing for antinuclear antibodies (ANA) by indirect immunofluorescence, 259 consecutive patients with positive ANA were identified. Medical charts of these patients were reviewed to assess the clinical diagnosis, with the reviewer having no knowledge of the anti‐ENA result. Clinical data were abstracted for all patients, and diagnoses established using American College of Rheumatology criteria. The utility of ENA antibodies in the diagnosis of SLE was determined by univariate and multivariate analysis among all patients who were positive for ANA, patients who were positive for ANA and for anti‐double‐stranded DNA (anti‐dsDNA), and patients who were positive for ANA and negative for anti‐dsDNA. Clinical differences between SLE patients with and those without anti‐ENA antibodies were assessed.

Results. Anti‐ENA antibodies, especially anti‐Ro/SS‐A, showed strong predictive diagnostic value among ANA+/anti‐dsDNA– patients, but were of no utility among ANA+/anti‐dsDNA+ patients. The only clinical manifestations that were more common among anti‐ENA+ SLE patients were pleuritis and the use of hydroxychloroquine.

Conclusion. The presence of anti‐ENA antibodies, especially anti‐Ro/SS‐A, is a useful predictor for the diagnosis of SLE, primarily among patients attending a referral rheumatology center who are positive for ANA and negative for anti‐dsDNA. No major clinical differences were noted among ANA+ SLE patients with versus those without ENA.
Deactivation of vascular endothelium by monoclonal anti–tumor necrosis factor α antibody in rheumatoid arthritis
Authors: Paleolog Ewa M. , Hunt Mary , Elliott Michael J. , Feldmann Marc , Maini Ravinder N. , Woody James N.
Published: February 2007

Objective. To assess whether monoclonal antibody to tumor necrosis factor α (TNFα) reduces endothelial activation in rheumatoid arthritis (RA).

Methods. Levels of serum E‐selectin, intercellular adhesion molecule 1 (ICAM‐1), and vascular cell adhesion molecule 1 (VCAM‐1), and circulating leukocytes (differential counts) were measured in RA patients before and up to 4 weeks after infusion of either placebo or chimeric anti‐TNFα antibody cA2 (1 or 10 mg/kg).

Results. Treatment with anti‐TNFα decreased serum E‐selectin and ICAM‐1 levels, with the earliest detectable changes observed on days 1–3 after anti‐TNFα infusion. No effect on VCAM‐1 levels was detected. In parallel, there was a rapid and sustained increase in circulating lymphocytes. The extent of the decrease in serum E‐selectin and ICAM‐1 levels and the increase in lymphocyte counts was significantly higher (P ≤ 0.05) in patients in whom a clinical benefit of anti‐TNFα was observed (≥20% response, by Paulus criteria, at week 4) compared with that in patients who failed to respond to anti‐TNFα at this time point.

Conclusion. We propose that decreased serum levels of adhesion molecules may reflect diminished activation of endothelial cells in the synovial microvasculature, leading to reduced migration of leukocytes into synovial joints, and thus prolonging the therapeutic effect of anti‐TNFα in RA.
Lack of association between augmentation mammoplasty and systemic sclerosis (scleroderma)
Authors: Hochberg Marc C. , Perlmutter Donna L. , Medsger Thomas A. , Nguyen Katherine , Steen Virginia , Weisman Michael H. , White Barbara , Wigley Fredrick M.
Published: February 2007

Objective. To examine the possible association between augmentation mammoplasty and systemic sclerosis (SSc; scleroderma).

Methods. Eight hundred thirty‐seven women with a clinical diagnosis of SSc, recruited as a volunteer sample from 3 university‐based, tertiary care scleroderma clinical research centers, and 2,507 race‐matched local control women, recruited by the technique of random‐digit‐dialing and frequency‐matched on age, completed a questionnaire providing data on history of augmentation mammoplasty, including possible complications of the procedure. The odds ratio (OR) and 95 percent confidence interval (95% CI) for the association of augmentation mammoplasty with SSc were estimated by multivariate logistic regression analysis with adjustment for age, race and center, and by conditional logistic regression analysis with adjustment for age.

Results. Eleven (1.31%) of the 837 cases reported a history of augmentation mammoplasty prior to diagnosis of SSc, compared with 31 (1.24%) of the 2,507 controls. The adjusted OR from the unmatched analysis was 1.07 (95% CI 0.53–2.13), while that from the matched analysis was 1.11 (95% CI 0.55–2.24).

Conclusion. These results fail to demonstrate a significant association between augmentation mammoplasty and SSc, and are consistent with those reported from other epidemiologic studies.
Pilot study of antithymocyte globulin in systemic sclerosis
Authors: Matteson Eric L. , Shbeeb Mohammad I. , McCarthy Tim G. , Calamia Kenneth T. , Mertz Lester E. , Goronzy Jörg J.
Published: February 2007

Objective. Between June 1, 1992 and August 31, 1994 we conducted an open pilot study of antithymocyte globulin (ATGAM; Upjohn, Kalamazoo, MI) in 10 patients with early systemic sclerosis (SSc) to assess whether this agent might prevent the progression of cutaneous and pulmonary involvement in this disease.

Methods. Adult patients with early SSc (<3 years) and evidence of progressive skin and pulmonary disease were enrolled. All patients were hospitalized and received a single course of intravenous ATGAM, at a dosage of 10 mg/kg over 4 hours, on 5 consecutive days. Patients were followed up at weeks 1, 2, 3, and 4, and months 2, 4, 6, and 12. Patients were considered to be improved if the Rodnan skin score decreased ≥25%, to be worse if the skin score increased ≥25%, and to be not improved if the skin score was within 25% of baseline. For pulmonary involvement, patients were considered to be improved if either the diffusing capacity for carbon monoxide or the forced vital capacity was increased ≥10%, worse if decreased by ≥10%, and stable if within 10% of baseline.

Results. Most patients tolerated the treatment well, although 1 patient developed an allergic reaction necessitating discontinuation of treatment, 1 developed a serum sickness reaction after completion of therapy, and 1 developed a central venous access–related axillary vein thrombosis. Two patients died of SSc‐related complications during the followup period. At 12 months, only 2 patients showed improvement in both skin and pulmonary function measures, whereas 5 patients were worse and 3 were stable.

Conclusion. At the dosage administered in this study, ATGAM appears ineffective in improving the skin and pulmonary features of SSc.
Evidence of free radical–mediated injury (isoprostane overproduction) in scleroderma
Authors: Stein C. Michael , Tanner S. Bobo , Awad Joseph A. , Roberts L. Jackson , Morrow Jason D.
Published: February 2007

Objective. Free radical–induced oxidative stress with consequent lipid peroxidation and resultant tissue damage has been suggested as a potential mechanism of the pathogenesis of scleroderma. However, because reliable measurement of lipid peroxidation in vivo is difficult, it has not been possible to adequately examine this hypothesis. We have previously described a series of bioactive prostaglandin F2–like compounds, termed F2‐isoprostanes, produced in vivo in humans by the noncyclooxygenase, free radical–catalyzed, peroxidation of arachidonic acid and have shown them to be a reliable measure of lipid peroxidation in vivo. In the present study, we determined whether scleroderma is associated with enhanced oxidative stress.

Methods. As a measure of oxidative stress, we determined urinary concentrations of a tetranordicarboxylic acid metabolite of F2‐isoprostanes (F2IP‐M) by mass spectrometry in 8 patients with scleroderma (representing a wide spectrum of disease, including limited disease with refractory digital ulceration or pulmonary hypertension, and diffuse disease) and in 10 healthy control subjects.

Results. F2IP‐M concentrations were significantly higher in patients with scleroderma (mean ± SEM 3.41 ± 0.64 ng/mg of creatinine) than in healthy controls (1.22 ± 0.14 ng/mg of creatinine) (P = 0.002). These elevations occurred in patients with limited disease and in those with diffuse disease.

Conclusion. The increased level of urinary F2IP‐M supports the hypothesis that free radical–induced oxidative injury occurs in scleroderma and provides a biologic marker whose relationship to disease activity and disease therapy may be important. These findings may also provide a rationale for exploring whether antioxidant therapy may influence the natural course of the disease.
Autoantibodies to fibrillarin in systemic sclerosis (scleroderma). An immunogenetic, serologic, and clinical analysis
Authors: Arnett Frank C. , Reveille John D. , Goldstein Rose , Pollard K. Michael , Leaird Kimberly , Smith Edwin A. , Leroy E. Carwile , Fritzler Marvin J.
Published: February 2007

Objective. To determine the frequency, clinical associations, and any major histocompatibility complex correlations of antifibrillarin antibodies in patients with systemic sclerosis (SSc).

Methods. Antifibrillarin antibodies were determined by indirect immunofluorescence, immunoblotting, and immunoprecipitation, and HLA class II alleles by DNA oligotyping, in a large cohort of SSc patients.

Results. Antifibrillarin was found in 8% of 335 SSc sera and was significantly more common in blacks (16%) than whites (5%), in males (33%) than females (14%), and in patients with cardiac, renal, or gut involvement. The HLA class II haplotype DRB1*1302, DQB1*0604 was found significantly more frequently in SSc patients with antifibrillarin compared with racematched normal controls and 260 SSc patients without antifibrillarin. In addition, 1 or more of the HLA–DQB1 alleles *0604, *0301, *0602, and/or *0302 was found in all antifibrillarin‐positive patients, and 62% of the antifibrillarin‐positive patients had 2 of these HLA–DQB1 alleles, a highly significant difference from both race‐matched normal controls and antifibrillarin‐negative SSc patients.

Conclusion. Antifibrillarin, although an infrequent nucleolar autoantibody, is a marker for severe SSc, especially in blacks and males, and is strongly associated with a unique HLA haplotype, as well as with combinations of certain HLA–DQB1 alleles.
Sensitivity and specificity of plasma and urine complement split products as indicators of lupus disease activity
Authors: Manzi Susan , Rairie Joan E. , Carpenter A. Betts , Kelly Robert H. , Jagarlapudi Santhi P. , Sereika Susan M. , Medsger Thomas A. , Ramsey‐Goldman Rosalind
Published: February 2007

Objective. To determine if measurement of serum complement split products (C4d, Bb, C5b–9) is better than conventional C3 and C4 measurements in distinguishing patients with varying degrees of lupus disease activity, and to determine if the presence of C3d in urine is helpful in distinguishing lupus patients with from those without early lupus nephritis.

Methods. Lupus disease activity was prospectively determined at 3 consecutive visits an average of 4 months apart, using the Systemic Lupus Activity Measure (SLAM), the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI), and physician global assessment (PGA). Blood samples were evaluated for the presence of C4d, Bb, and C5b–9 by quantitative microsassay plate enzyme immunoassay at each patient visit. We characterized urinary excretion of C3 fragments (with attention to C3d) by sodium dodecyl sulfate–polyacrylamide gel electrophoresis with Western blotting.

Results. Thirty‐one SLE patients were enrolled in the study. The mean SLAM score and the mean SLEDAI score each correlated well with the PGA at all 3 visits. A SLAM score of 6 and a SLEDAI score of 4 had the best overall sensitivity and specificity for predicting moderate‐to‐severe disease activity by PGA (100% and 73%, respectively, for the SLAM and 86% and 94%, respectively, for the SLEDAI). Serum C4d and Bb were more sensitive indicators of current moderate‐to‐severe lupus disease activity at all 3 visits than were serum C5b–9, C3, and C4. C3 and C4 were more specific indicators of moderate‐to‐severe disease activity. Serum C4d and Bb were more sensitive at predicting moderate‐to‐severe disease activity at subsequent visits than were C5b–9, C3, and C4. Urine C3d was better than C3, plasma C4d, Bb, C5b–9 and anti–double‐stranded DNA antibody in distinguishing patients with from those without acute lupus nephritis (P = 0.02).

Conclusion. C4d and Bb are sensitive indicators of moderate‐to‐severe lupus disease activity and may be most helpful in situations where conventional measurements are not, such as in lupus patients whose C3 and C4 levels remain normal despite evidence of clinical disease activity. It appears from this study that detection of urine C3d may be a simple way of measuring complement activation in the setting of lupus renal disease. The availability of instruments for clinical disease activity measurement such as the SLAM and the SLEDAI may enable more consistent definition of lupus disease activity and may thus provide a means for better examining the role of complement activation products in predicting lupus disease activity in larger patient populations.
Inhibition and prevention of monosodium urate monohydrate crystal–induced acute inflammation in vivo by transforming growth factor β1
Authors: Lioté Frédéric , Prudhommeaux Florence , Schiltz Corinne , Champy Romuald , Herbelin André , Ortiz‐Bravo Esteban , Bardin Thomas
Published: February 2007

Objective. We investigated the effects of transforming growth factor β1 (TGFβ1) on monosodium urate monohydrate (MSU) crystal‐induced acute inflammation in vivo.

Methods. One hour after MSU crystal–induced acute inflammation was produced in the rat subcutaneous air pouch model, the effects of recombinant human TGFβ1 (rHuTGFβ1; 10–100 pg/animal) and ultrapure TGFβ1 (UPTGFβ1; 100 and 500 pg/animal) were assessed, based on absolute and differential white blood cell counts in the exudate. The effects of 10 pg of rHuTGFβ1 preincubated with a specific anti‐TGFβ antibody, and the effects of coinjection of crystals and rHuTGFβ1, were also studied.

Results. UPTGFβ1 and rHuTGFβ1 markedly reduced MSU crystal–induced inflammation. Recombinant human TGFβ1 also reduced inflammation when administered concomitantly with MSU crystals. Moreover, rHuTGFβ1 and UPTGFβ1, injected 1 hour after MSU crystal injection, reduced the inflammatory response in a dose‐dependent manner. Injection of rHuTGFβ1 (100 pg/animal) resulted in a >90% reduction in the maximal white blood cell count, achieved 6 hours after crystal injection. Preincubation of rHuTGFβ1 with a specific anti‐TGFβ1 antibody significantly (P < 0.01) reversed the inhibitory effect of rHuTGFβ1 on the inflammatory response. Consistent with the regulation of inflammatory cell recruitment into the joint, the percentage of monocytes markedly decreased (P < 0.01) following local injection with rHuTGFβ1 6 hours after MSU crystal injection.

Conclusion. Exogenous TGFβ1 prevents and inhibits MSU crystal–induced acute inflammation in vivo. Its role in the self‐limitation of gouty attacks deserves consideration, among the various other factors involved.