Therapeutic potential of russelioside B as anti-arthritic agent in Freund's adjuvant-induced arthritis in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Caralluma species are traditional edible herbs used in folkloric medicine as antidiabetic, antioxidant, antipyretic, antirheumatic, anti-inflammatory and anthelmintic agents. C. quadrangula was selected in this study to document the traditional use of the genus as anti-rheumatic treatment and the possible mechanisms of action. AIM OF THE STUDY: The higher mortality rates and shorter survival among the patients suffering from rheumatoid arthritis (RA) led to the increased interest on searching for new treatments for RA. Russelioside B (RB), a major pregnane glycoside found in C. quadrangula, was evaluated as a new anti-rheumatic agent. MATERIALS AND METHODS: The n-butanol fraction of C. quadrangula was chromatographed on a silica gel column to isolate RB. The adjuvant-induced arthritis (AIA) model was established in rats by intradermal injection of complete Freund's adjuvant (CFA) to evaluate its anti-arthritic effect. Ibuprofen was used as a reference drug. Forty rats were randomly divided into 5 groups (n = 8): normal (NOR); CFA model (CFA); ibuprofen, 5 mg/kg; RB, 25 mg/kg and RB, 50 mg/kg. The treatments were initiated from day 16 when AIA model was established and continued up to day 40. Serum diagnostic rheumatoid markers, inflammatory cytokines, oxidative stress biomarkers, cartilage and bone degeneration enzymes were assessed. RESULTS: RB at 50 mg/kg b. wt., showed significant decreases in the activities of hyaluronidase and ß-glucouronidase enzymes as well significant decreases in the levels of proinflammatory cytokines as nuclear factor-kappa-B (NF-κB), tumour necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1ß (IL-1ß) compared to the CFA group; 11.04 ± 0.61 pg/mg protein, 4.35 ± 0.25 pg/mg protein, 3.32 ± 0.13 pg/mg protein & 2.75 ± 0.14 pg/mg protein for RB, 50 mg/kg b. wt. group vs. 25.33 ± 2.13 pg/mg protein, 25.65 ± 2.1 pg/mg protein, 22.20 ± 1.34 pg/mg protein & 13.27 ± 1.40 pg/mg protein for the arthritic group, respectively. The total antioxidant capacity (TAC) was significantly restored to normal values in RB, 50 mg/kg treated rats (4.01 ± 0.09 nmol/mL vs. 3.71 ± 0.27 nmol/mL) and the levels of myeloperoxidase (MPO) reduced by 10-folds of the CFA arthritic group. Bone histomorphometry revealed that RB treatment significantly attenuated the CFA-induced bone loss in a dose-dependent manner. CONCLUSIONS: These findings suggested that the anti-arthritic effect of RB was mediated through the reduction of the rheumatoid markers, anti-inflammatory and antioxidant action, inhibition of cartilage and bone degenerative enzymes as well as attenuation of bone loss and osteoclastogenesis.

Vasculitis reumatoide: estudio de 41 casos / Rheumatoid vasculitis: study of 41 cases

OBJETIVO: Analizar las características clínicas, tratamiento y evolución de la vasculitis reumatoide. MÉTODOS: Estudio retrospectivo (1975-2017). Pacientes con vasculitis reumatoide diagnosticados en 2 servicios de reumatología. RESULTADOS: Se incluyó a 41 pacientes: 17 (41,5%) varones y 24 (58,5%) mujeres; con una edad media al diagnóstico de 67 ± 9 años y una duración de la artritis reumatoide de 10 ± 8,3 años. La artritis fue erosiva en 33 (80%) pacientes. Tanto el factor reumatoide como los anticuerpos antipéptido citrulinados fueron positivos en todos los casos. Los síntomas constitucionales se presentaron en 30 pacientes (73%) y las manifestaciones extrarticulares en 17 (41%). Las manifestaciones clínicas más frecuentes fueron: cutáneas 28 (68%) y neuropatía periférica 26 (63%). Todos los pacientes fueron tratados con glucocorticoides. En 24 pacientes (58%) se asoció a un inmunosupresor y 5 (12%) pacientes fueron tratados con fármacos biológicos. La mortalidad a los 2 años de seguimiento fue del 33%. Las principales causas de muerte fueron: la infección y la progresión de la vasculitis reumatoide. La frecuencia de la vasculitis reumatoide disminuyó en la última década. CONCLUSIONES: Las manifestaciones clínicas de la vasculitis reumatoide en España son similares a las descritas. La frecuencia disminuye; sin embargo, el cuadro clínico y la gravedad se mantienen invariables

AIM: To describe the clinical manifestations, evolution and treatment of patients with rheumatoid vasculitis. METHODS: Retrospective study (1975-2017) of all patients diagnosed with rheumatoid vasculitis in 2 Rheumatology Services. RESULTS: A total of 41 patients were included, 17 (41.5%) males and 24 (58.5%) females; mean age at diagnosis: 67 ± 9 years; duration of rheumatoid arthritis: 10 ± 8.3 years. Most patients had erosive disease, 33 (80%). Rheumatoid factor and anticitrullinated antibodies were positive in all patients. Constitutional symptoms were present in 30 (73%) patients and extra-articular manifestations in 17 (41%) patients. The clinical manifestations of rheumatoid vasculitis were mainly: cutaneous 28 (68%), and polyneuritis 26 (63%). All patients were treated with glucocorticoids. An immunosuppressant was associated in 24 (58.5%) patients. Five (12%) patients were treated with the association of glucocorticoids and a biologic treatment. The mortality after 2years of follow-up was 33%, the most common causes being infection and progression of the vasculitis. The frequency of rheumatoid vasculitis has decreased over the last decade. CONCLUSION: The clinical manifestations of rheumatoid vasculitis were similar to previous studies. The frequency of rheumatoid vasculitis seems to decrease. However, the clinical picture and severity remains invariable

Does immunological remission, defined as disappearance of autoantibodies, occur with current treatment strategies? A long-term follow-up study in rheumatoid arthritis patients who achieved sustained DMARD-free status.

OBJECTIVES: Sustained disease-modifying antirheumatic drug (DMARD)-free status, the sustained absence of synovitis after cessation of DMARD therapy, is infrequent in autoantibody-positive rheumatoid arthritis (RA), but approximates cure (ie, disappearance of signs and symptoms). It was recently suggested that immunological remission, defined as disappearance of anti-citrullinated protein antibodies (ACPA) and rheumatoid factor (RF), underlies this outcome. Therefore, this long-term observational study determined if autoantibodies disappear in RA patients who achieved sustained DMARD-free remission. METHODS: We studied 95 ACPA-positive and/or RF-positive RA patients who achieved DMARD-free remission after median 4.8 years and kept this status for the remaining follow-up (median 4.2 years). Additionally, 21 autoantibody-positive RA patients with a late flare, defined as recurrence of clinical synovitis after a DMARD-free status of ≥1 year, and 45 autoantibody-positive RA patients who were unable to stop DMARD therapy (during median 10 years) were studied. Anti-cyclic citrullinated peptide 2 (anti-CCP2) IgG, IgM and RF IgM levels were measured in 587 samples obtained at diagnosis, before and after achieving DMARD-free remission. RESULTS: 13% of anti-CCP2 IgG-positive RA patients had seroreverted when achieving remission. In RA patients with a flare and persistent disease this was 8% and 6%, respectively (p=0.63). For anti-CCP2 IgM and RF IgM, similar results were observed. Evaluating the estimated slope of serially measured levels revealed that RF levels decreased more in patients with than without remission (p<0.001); the course of anti-CCP2 levels was not different (p=0.66). CONCLUSIONS: Sustained DMARD-free status in autoantibody-positive RA was not paralleled by an increased frequency of reversion to autoantibody negativity. This form of immunological remission may therefore not be a treatment target in patients with classified RA.

Identifying markers of sustained remission in rheumatoid arthritis patients on long-term tapered biological disease-modifying antirheumatic drugs.

To identify features associated with long-term persistent remission in rheumatoid arthritis (RA) patients on tapered biological disease-modifying antirheumatic drugs (bDMARD) (tap-bDMARD) therapy. We carried out a 40-month (m) extension follow-up study of 77 RA patients from a previous 12 m tap-bDMARD study. Disease activity was assessed at baseline and every 3 months. Doppler US investigation of 42 joints for the presence and grade (0-3) of B-mode synovial hypertrophy (SH) and synovial power Doppler signal (i.e., Doppler synovitis) was performed before starting the tap-bDMARD strategy by a rheumatologist blinded to clinical and laboratory data. At the 40 m mark, 44 (57.1%) patients failed the tap-bDMARD strategy, while 33 (42.9%) succeeded. Patients who presented a failed tap-bDMARD had significantly longer disease duration, a longer time from symptom onset to synthetic (s) DMARD start, longer duration of sDMARD treatment, a greater number of sDMARDs, and a higher baseline DAS28 and SDAI than patients with successful tap-bDMARD at 40 months. In logistic regression analysis, the presence of baseline Doppler synovitis, a DAS28 ≥ 2.2, and the presence of rheumatoid factor were identified as predictors of tap-bDMARD failure at 40 m. In those patients who succeed tap-bDMARD at 12 m, a smoking habit was significantly more frequently found in tap-bDMARD failures at 40 m. Our results showed that DAS28 and the presence of Doppler synovitis, RF and a smoking habit predicted long-term tap-bDMARD failure.

Rheumatoid factor in idiotypic regulation of autoimmunity.

AIM: Rheumatoid factor (RF) is known to be heterogeneous, and RFs detected by various methods exhibit different characteristics. In addition to interacting with the Fc region of immunoglobulin G (IgG), certain RFs are able to recognize idiotypes of antibodies. Given the important role of idiotypic interactions in regulating autoimmunity, we hypothesize that RF is involved in regulation of lymphocyte activity against autoimmune disease-inducing antigens via idiotype-anti-idiotype interactions with these lymphocytes. METHOD: RF level and the existence of idiotype-anti-idiotype interactions between RF and antibodies to autoimmunity-inducing antigens were studied in rats resistant and sensitive to collagen-induced arthritis, encephalomyelitis and atherosclerosis. RF was assayed by agglutination of tanned IgG-loaded erythrocytes. RESULTS: Rat resistance to autoimmune disease is associated with high RF production during the initiation of the immune response, and a low RF level during this period may be a preclinical marker of experimental autoimmune disease manifestation. RF-containing sera compete with an antigen if the RF-containing sera were obtained from rats immunized with that antigen, and they non-specifically inhibit binding of different antigen-antibody pairs. This suggests that RFs are anti-idiotypic antibodies that carry two kinds of paratopes: a particular paratope that recognizes the antigen-binding sites of antibodies, and a shared paratope that serves to recognize the recurrent idiotype on antibodies. Antigenic epitopes for the shared RF paratope can be created in the hinge region of Fc fragments of homologous IgG. CONCLUSION: RF detected by agglutination of tanned IgG-loaded erythrocytes is involved in negative idiotypic regulation of lymphocytes specific to autoimmunity-inducing antigens.

Elevated autoantibody content in rheumatoid arthritis synovia with lymphoid aggregates and the effect of rituximab.

INTRODUCTION: The purpose of this study was to quantitatively evaluate the contribution of synovial lymphoid aggregates to autoantibody (rheumatoid factor [RF] and anti-cyclic citrullinated peptide [anti-CCP]) and total immunoglobulin (IgG and IgM) production in rheumatoid arthritis (RA) patients and the effect thereon of the B-cell-depleting antibody, rituximab, in the ARISE (Assessment of Rituximab's Immunomodulatory Synovial Effects) trial. METHODS: Autoantibodies as well as total IgM and IgG were quantified by enzyme-linked immunosorbent assay in extracts of synovial tissues and matched serum from patients with RA or osteoarthritis (OA). Synovial biopsies and serum were obtained at baseline and 8 weeks following rituximab therapy in 14 RA patients. A synovial/serum index (SSI) was calculated as the ratio of synovial to serum antibody/albumin, with values above 1 representing synovial enrichment. Lymphoid aggregates were evaluated histologically. RESULTS: Anti-CCP IgG, but not RF-IgM, was significantly enriched in RA synovia compared with serum. Total IgM and IgG were also enriched in RA, but not in OA. SSI correlated significantly with mRNA content for both IgM and IgG, demonstrating that it reflected synovial immunoglobulin production. RA synovia with lymphocyte aggregates contained significantly elevated RF-IgM and anti-CCP IgG compared with tissues with diffuse lymphoid infiltration. Rituximab treatment did not affect synovial autoantibody or total immunoglobulin SSI overall. However, in aggregate-containing tissues, rituximab significantly reduced total IgM and IgG SSI as well as IgM and IgG1 mRNA. Surprisingly, RF-IgM and anti-CCP IgG SSIs were unchanged by rituximab in aggregate-containing synovia. CONCLUSIONS: Combined with earlier observations that synovial lymphoid aggregates are unaltered by rituximab treatment, these data suggest that lymphoid aggregates may provide a protective niche for autoantibody-producing cells.

IgM-rheumatoid factor, anti-cyclic citrullinated peptide, and anti-citrullinated human fibrinogen antibodies decrease during treatment with the tumor necrosis factor blocker infliximab in patients with rheumatoid arthritis.

OBJECTIVE: To investigate the effect of treatment with infliximab on serum levels of rheumatoid factor (IgM-RF), antibodies against cyclic citrullinated peptide (anti-CCP), and antibodies against deiminated human fibrinogen, a specific citrullinated peptide (ACF), and their association with disease activity and disease duration in patients with rheumatoid arthritis (RA). METHODS: The study sample included 62 consecutive patients who were treated with infliximab for at least one year. IgM-RF, anti-CCP, and ACF were measured at 0, 14, 30, and 46 weeks. RESULTS: Patients had a mean age of 54 years and median disease duration of 10 years and were predominantly female (81%). At baseline 63%, 77%, and 82% of patients were positive for IgM-RF, anti-CCP, and ACF, respectively. In terms of percentages, the levels of IgM-RF were reduced by 64% at 46 weeks, while anti-CCP and ACF levels were reduced by roughly 25%. The decrease in serum levels of these autoantibodies was not associated with the decrease in disease activity. The change in ACF was significantly related to disease duration, while the changes in IgM-RF or anti-CCP were not. CONCLUSION: In a cohort of patients with RA who responded to infliximab therapy, all autoantibodies decreased significantly, but IgM-RF showed a larger decrease than anti-CCP or ACF. These changes in levels of autoantibodies are not directly related to the change in disease activity. Early in the disease, ACF levels were best influenced by treatment with infliximab.

No effects of adalimumab therapy on the activation of NF-kappaB in lymphocytes from patients with severe rheumatoid arthritis.

The aim of this study was to determine the activation level of the pro-inflammatory transcription factor nuclear factor kappaB (NF-kappaB) in lymphocytes of patients with rheumatoid arthritis (RA) before and during an anti-tumor necrosis factor alpha (TNFalpha) therapy (adalimumab). In addition, we analyzed the inflammatory markers, interleukin 6 (IL-6), and C-reactive protein (CRP) and investigated the expression of rheumatoid factor (RF) and anti-cyclic citrullinated peptide (anti-CCP) autoantibodies in patients' sera. Twenty RA patients and 20 control subjects were investigated. RA patients' characteristics were evaluated by radiography and disease activity score 28 (DAS 28). Twelve weeks of adalimumab therapy was effective in the treatment of RA patients, as shown by a significant improvement of the DAS 28. The inflammatory markers IL-6 and CRP were significantly different in sera of RA patients compared to the control group before the onset of therapy and exhibited a tendency to return to normal levels during the first 12 weeks of therapy. We measured a comparable activation level of NF-kappaB in lymphocytes of control subjects and of RA patients before starting adalimumab therapy. During the following 12 weeks, no significant changes in the activation levels of both NF-kappaB subunits were detected. Serum concentration of RF was significantly lower after 12 weeks, whereas anti-CCP antibody level remained constant.

HCV-NS3 and IgG-Fc crossreactive IgM in patients with type II mixed cryoglobulinemia and B-cell clonal proliferations.

We demonstrate that in three cases of MC (two with immunocytoma), the IgM-RF+ component of their cryoprecipitated represents the circulating counterpart of the B-cell receptor (BCR) of the monoclonal overexpanded B-cell population. These IgMs were isolated and used to demonstrate a crossreactivity against both hepatitis C virus (HCV) NS3 antigen and the Fc portion of IgG. Epitopes were identified in a fraction of exemplary samples by using epitope excision approach (NS(31250-1334) and IgG Fc(345-355)). The same phenomenon of crossreactivity has been shown to occur in vivo after immunization of a mouse with the NS3(1251-1270) peptide. To verify if the same reaction was also present in MC samples characterized by an oligo/polyclonal B-cell proliferation, IgM crossreactivity was tested in 14 additional samples. Five out of the 14 were reactive against HCV NS3 and 11 out of 14 were reactive against IgG-Fc peptide. The data support the role of HCV NS3 antigen in a subset of patients with MC, whereas the high frequency of the IgG-Fc epitope suggests that these B cells originate from precursors strongly selected for auto-IgG specificity. We suggest that engagement of specific BCRs by NS3 (or NS3-immunocomplex) antigen could explain the prevalence of IgM cryoglobulins in these patients.

Low dose interleukin-2 and 13-cis-retinoic acid as maintenance therapy in patients with solid tumors responsive to chemotherapy.

After aggressive surgery and chemotherapy, patients (PTS) with advanced solid tumors have often an impaired immune function that may facilitate tumor relapse. We have demonstrated, in a phase IB study, that administering low-dose interleukin-2 (IL-2) and 13-cis retinoic acid (RA), there was a significant improvement of the immune function, with low toxicity, in PTS with advanced tumors that had been treated with aggressive surgery and chemotherapy (Clin Cancer Res 7: 1251-1257, 2001). A further A phase II study (International J Oncology 20: 1275-82, 2002) showed the efficacy of IL-2/RA in improving immunological parameters and in prolonging response in the same category of PTS. Aim of this study was evaluate the clinical and laboratory outcome of 214 patients, with advanced solid tumors, that, after aggressive surgery and chemotherapy, were treated with: subcutaneous IL-2 1.8 x 10(6) IU and oral RA, 0.5 mg/Kg for 5 days/week for 2 cycles of 3 weeks, with a 1-week rest, for up to 2 years. After a median follow-up time of 31 months, there was a statistically significant improvement in the number of lymphocytes, T4/T8 ratio, NK, with respect to baseline values. Responses were maintained for a median time of 36.4 months, while median survival was not reached yet (57% of PTS alive). These data show that the administration of low-dose IL-2/RA determines, with a low toxicity, a sustained increase in the immunological parameters known to be prognostically relevant.

Interference of rheumatoid factor activity by aspartame, a dipeptide methyl ester.

Circulating autoimmune complexes of IgM rheumatoid factors (RF) bound to the Fc portions of normal, polyclonal IgG antibodies are frequently present in humans with rheumatoid arthritis (RA). The sweet tasting methyl ester of L-Asp-L-Phe (aspartame or APM) was found to relieve pain and improve joint mobility in subjects with osteo- and mixed osteo/rheumatoid arthritis [Edmundson, A. B. and Manion, C. V. (1998). Clin. Pharmac. Ther. 63, 580-593]. These clinical observations prompted the testing of the inhibition by APM of the binding interactions of human IgM RFs with IgG Fc regions. The propensity of APM to inhibit IgM RF binding was assessed by competitive enzyme immunoassays with solid-phase human IgG. Ten RA serum samples and three purified monoclonal cryoglobulins, all of which had RF activity, were tested in this system. We found that the presence of APM significantly reduced the binding of IgM RFs. The inhibitory propensity of APM with monoclonal RF cryoglobulins was increased by the addition of CaCl(2) to the binding buffer. Similar inhibition of the binding of RA derived RFs to IgG was observed for Asp-Phe and its amidated derivative, indicating that the methyl ester is not required for APM's interaction with IgM antibodies. A human (Mez) IgM known to bind octameric peptides derived from the Fc portion of a human IgG(1) antibody was tested for binding of dipeptides by the Pepscan method of combinatorial chemistry. The relative binding constants of Asp-Phe and Phe-Asp were ranked among the highest values for 400 possible combinations of the 20 most common amino acids. Possible blocking interactions of APM were explored by computer-assisted docking studies with the model of a complex of an RF Fab with the Fc of a human IgG(4) antibody. Modeling of ternary immune complexes revealed a few key residues, which could act as molecular recognition sites for APM. A structural hypothesis is presented to explain the observed interference with RF reactivity by APM. Extrapolations of the current results suggest that APM may inhibit the binding of IgG in a substantial proportion of IgM RFs. Interference of RF reactivity, especially in RA patients, may alleviate the pain and immobility resulting from chronic inflammation of the joints.

Interference of rheumatoid factor activity by aspartame, a dipeptide methyl ester.

Circulating autoimmune complexes of IgM rheumatoid factors (RF) bound to the Fc portions of normal, polyclonal IgG antibodies are frequently present in humans with rheumatoid arthritis (RA). The sweet tasting methyl ester of L-Asp-L-Phe (aspartame or APM) was found to relieve pain and improve joint mobility in subjects with osteo- and mixed osteo/rheumatoid arthritis [Edmundson, A. B. and Manion, C. V. (1998). Clin. Pharmac. Ther. 63, 580-593]. These clinical observations prompted the testing of the inhibition by APM of the binding interactions of human IgM RFs with IgG Fc regions. The propensity of APM to inhibit IgM RF binding was assessed by competitive enzyme immunoassays with solid-phase human IgG. Ten RA serum samples and three purified monoclonal cryoglobulins, all of which had RF activity, were tested in this system. We found that the presence of APM significantly reduced the binding of IgM RFs. The inhibitory propensity of APM with monoclonal RF cryoglobulins was increased by the addition of CaCl(2) to the binding buffer. Similar inhibition of the binding of RA derived RFs to IgG was observed for Asp-Phe and its amidated derivative, indicating that the methyl ester is not required for APM's interaction with IgM antibodies. A human (Mez) IgM known to bind octameric peptides derived from the Fc portion of a human IgG(1) antibody was tested for binding of dipeptides by the Pepscan method of combinatorial chemistry. The relative binding constants of Asp-Phe and Phe-Asp were ranked among the highest values for 400 possible combinations of the 20 most common amino acids. Possible blocking interactions of APM were explored by computer-assisted docking studies with the model of a complex of an RF Fab with the Fc of a human IgG(4) antibody. Modeling of ternary immune complexes revealed a few key residues, which could act as molecular recognition sites for APM. A structural hypothesis is presented to explain the observed interference with RF reactivity by APM. Extrapolations of the current results suggest that APM may inhibit the binding of IgG in a substantial proportion of IgM RFs. Interference of RF reactivity, especially in RA patients, may alleviate the pain and immobility resulting from chronic inflammation of the joints.

Is bone turnover a determinant of bone mass in rheumatoid arthritis?

OBJECTIVE: To study the relationship between bone turnover markers and bone mineral density in patients with rheumatoid arthritis. METHODS: We studied 54 patients, 24 of whom were receiving low dose steroids, and compared them to 54 age and sex matched controls. RESULTS: An 8.2% decrease of femoral neck bone mineral density (BMD) was found in patients not taking steroids compared with controls (confidence interval 1.2-15.3%). Serum markers of bone turnover -- namely, procollagen type I C-terminal propeptide (PICP) and procollagen type I N-terminal propeptide (PINP), which reflect bone formation, and procollagen type I C-terminal telopeptide, which reflects bone resorption -- were significantly increased compared with controls (p < 0.05, p < 0.01, p < 0.01, respectively). Both PINP levels and PICP levels were correlated with the femoral neck BMD as well as osteocalcin levels: R = -0.32 (p < 0.05), R = -0.29 (p < 0.05), and R = -0.42 (p < 0.01), respectively. The best independent predictors of bone mass (stepwise multiple regression analysis) at the femoral neck were steroid use, osteocalcin levels, age, height, the presence of rheumatoid factor, and the Health Assessment Questionnaire score, which explained 61.6% of the variance in femoral neck BMD. CONCLUSION: Elderly patients with RA using steroids with severe disease and high levels of osteocalcin have marked osteoporosis at the hip.

Suppression of in vitro IgM rheumatoid factor production by diphtheria toxin interleukin 2 recombinant fusion protein (DAB 486IL-2) in patients with refractory rheumatoid arthritis.

OBJECTIVE: To investigate the effect of diphtheria toxin interleukin 2 recombinant fusion protein (DAB 486IL-2) on in vitro synthesis of immunoglobulin and rheumatoid factor (RF) in patients with severe refractory rheumatoid arthritis (RA) enrolled in a phase II, double blind, placebo controlled study. METHODS: Anticoagulated venous blood samples were obtained before (Day 1) and after (Day 28) intravenous infusion of either DAB 486IL-2 at 0.075 mg/kg/day (12 patients) or saline placebo (10 patients) on Days 1-5. Peripheral blood leukocytes (PBL) were prepared by density gradient centrifugation, cultured in the presence and absence of pokeweed mitogen (PWM) for one week, and culture supernatants assayed for immunoglobulins and IgM RF by ELISA. RESULTS: Compared to placebo treated patients, PWM induced IgM RF synthesis by PBL decreased after treatment with DAB 486IL-2 (p = 0.043). However, there was no apparent correlation with clinical improvement. PWM induced IgM, IgA, and IgG synthesis also tended to decrease, although the changes did not attain statistical significance. In contrast, PWM induced IgM RF, IgM, IgA, and IgG synthesis by PBL from patients treated with placebo tended to increase during the observation period. Spontaneous immunoglobulin and IgM RF production by PBL from either the DAB 486IL-2 or placebo patients remained stable. CONCLUSION: These observations raise the possibility that DAB 486IL-2 may diminish B cell function either directly or indirectly through effects on T cell function, but the change may not correspond to clinical response.

In vitro effects of methotrexate on polyamine levels in lymphocytes from rheumatoid arthritis patients.

OBJECTIVE: Several studies have documented increased levels of polyamines in rheumatoid arthritis (RA). We have suggested that one of the mechanisms of action of methotrexate (MTX) involves the inhibition of polyamine synthesis in lymphocytes. In this study, we sought to establish the inhibitory effect of MTX on polyamine synthesis and its specificity. METHODS: Polyamine levels were determined in stimulated RA lymphocytes incubated in vitro with MTX and compared to levels in lymphocytes incubated with hydrocortisone, D-penicillamine, or medium alone. Lymphocyte polyamine levels were correlated with IgM-rheumatoid factor (RF) synthesis. RESULTS: Incubation with MTX resulted in concentration-dependent decreased intracellular levels of spermidine and spermine, while putrescine levels were not affected. Addition of folinic acid or S-adenosyl-methionine (SAM) prevented this MTX-induced inhibition. Incubation with D-penicillamine or hydrocortisone had no significant effect on polyamine levels. There was a positive correlation between intracellular polyamine levels and the inhibition of IgM-RF synthesis by MTX. CONCLUSION: These data suggest that MTX inhibits the synthesis of spermidine and spermine in stimulated RA lymphocytes through inhibition of the SAM-dependent pathway. This inhibition may be related to the immune-modulating properties of MTX.

Prolactin-induced immunoglobulin and autoantibody production by peripheral blood mononuclear cells from systemic lupus erythematosus and normal individuals.

Prolactin (PRL) has been shown to have immunoregulatory effects on a variety of immune responses. Its effect on B cell immune responses is suggested by in vitro data demonstrating a direct effect on B cell activation and differentiation, and also in vivo data demonstrating a biphasic stimulation of antibody production to sheep red blood cells. In addition, it has been shown both in animal models and patients with hyperprolactinemia that PRL may influence the presence of certain autoantibodies. The objective of this work was to study the effect of PRL on the induction of immunoglobulins, and anti-DNA and rheumatoid factor (RF) autoantibody production from peripheral blood mononuclear cells (PBMCs) from normal individuals and systemic lupus erythematosus (SLE) patients. Six female SLE patients and 10 normal individuals (5 females and 5 males) were studied. Ficoll-Hypaque-isolated PBMCs (1x10(6) cells/ml) with high concentrations of PRL (10(-4)-10(-8)M) and pokeweed mitogen (PWM) diluted to 1:400. An ELISA assay was used for immunoglobulins, RF and anti-dsDNA antibodies. PRL stimulated IgG and IgM production in a biphasic manner in normal PBMCs. Enhanced synthesis was observed at 10(-6) M, and a stimulatory effect was again observed at higher doses of PRL (10(-4))M. In contrast, only a mild stimulatory effect was observed in IgG synthesis by SLE PBMCs. These changes in Ig synthesis, however, did not reach statistical significance. PRL also induced IgG and IgM anti-dsDNA antibodies by both normal and SLE lymphocytes, but no differences were observed when compared to PWM stimulation. PRL induced IgM RF synthesis by normal lymphocytes but had no effect on SLE PBMCs. This study demonstrates that PRL induced immunoglobulin synthesis by normal and to a lesser degree by SLE lymphocytes, and also induced anti-dsDNA antibody by normal and SLE PBMCs, and IgM RF by normal PBMCs. However, the exact mechanism(s) of PRL action on the immune response awaits elucidation.

Inflammatory and immunological parameters of disease activity in rheumatoid arthritis patients treated with minocycline.

The objective of this study was to analyze the anti-inflammatory effect of minocycline in rheumatoid arthritis. Serum samples of 65 RA patients who completed a 26-week randomized double-blind trial of minocycline (100 mg twice a day) versus placebo were studied. In this trial some clinical parameters and in particular the acute phase response decreased significantly in the minocycline-treated group. Serum levels of albumin and interleukin-6 (IL-6) were compared with CRP levels in order to study the acute phase response. Furthermore, rheumatoid factor (RF) and total immunoglobulin isotypes as well as serum levels of soluble interleukin-2 receptor (sIL2-2R) were determined in order to study immunological parameters of the disease. Immunoglobulins and cytokines were measured by ELISA. Serum levels of albumin remained stable, whereas serum CRP levels decreased both in the minocycline- and in the placebo-treated group. Serum levels of IL-6 decreased in the minocycline-treated group only and this decrease was positively correlated with the decrease in CRP levels. Minocycline significantly decreased serum IgM-RF, IgA-RF, total IgM and total IgA levels. In addition the ratio of IgM-RF/total IgM decreased in the minocycline-treated group. No such changes were observed in the placebo-treated group. The anti-inflammatory effect of minocycline in RA patients may be due to the reduction in the synthesis of IL-6 and rheumatoid factor.

Calcitonin inhibits production of immunoglobulins, rheumatoid factor and interleukin-1 by mononuclear cells from patients with rheumatoid arthritis.

OBJECTIVES: Elcatonin (eCT), an eel calcitonin derivative, is shown to considerably improve the clinical signs and symptoms, as well as laboratory data, in patients with rheumatoid arthritis (RA). The therapeutic efficacy of eCT, however, is reduced by preceding and/or concomitant use of corticosteroid. Thus the effects of eCT on the production of immunoglobulins, IgMRF and interleukin-1 (IL-1) by mononuclear cells (MNCs)/monocytes were studied, and compared among patients with RA that received three kinds of treatment and also normal volunteers (NV). METHODS: Ten patients with RA had been treated with a non-steroidal anti-inflammatory drug only (NSAID group), 11 with oral prednisolone (PSL group), and eight with intramuscular eCT (eCT group). MNCs/monocytes from these patients, and also 10 from the NV group, were collected and cultured. IgG, IgA, IgM, IgMRF, IL-1 alpha and IL-1 beta in the supernatants were measured by enzyme-linked immunosorbent assay (ELISA). In the NSAID, PSL and NV groups, eCT was added to the culture medium, and the effects of eCT on production of these substances were studied. RESULTS: Baseline production of IgM, IL-1 alpha and IL-1 beta by MNCs/monocytes in the eCT and NV groups was significantly lower than that in the NSAID group. Furthermore, addition of eCT to the culture medium significantly inhibited the productions of IgG, IgMRF, IL-1 alpha and IL-1 beta by MNCs/monocytes in the NSAID group, whereas production of neither IgG, IgA, IgM, IgMRF nor IL-1 by MNCs/monocytes in the PSL and NV groups was affected by eCT. CONCLUSION: eCT may regulate immune responses through MNC/monocyte function in patients with RA. The present results support our proposal that eCT is an effective agent for the treatment of RA.

Long-term treatment with 5,5-diphenylhydantoin reduces lymphadenopathy and anti-ssDNA autoantibodies in C57BL/6-lpr/lpr mice.

To further the insight in the immunomodulating properties of the anticonvulsant 5,5-diphenylhydantoin (DPH), C57BL/6 (B6), C57BL/6-lpr/lpr (B6-lpr/lpr) and MRL/MpJ- +/+ (MRL) mice received DPH orally for six months to determine weekly urinary biopterin levels, a potential T-cell activation marker, by high performance liquid chromatography. At the end of the experiment serum antibody levels were measured by ELISA and relative lymphoid organ weights determined. DPH treatment resulted in reduced body weight in all strains, reduced spleen weights in B6 and MRL mice, profoundly reduced popliteal lymph node weights in B6-lpr/lpr mice and increased thymus weights in MRL mice. DPH treatment decreased serum IgM, IgG and IgA as well as IgM and IgG anti-ssDNA levels in B6-lpr/lpr mice, but did not affect these parameters in other strains. Effects of DPH on IgM rheumatoid factor levels in B6-lpr/lpr mice were inconsistent. Urinary biopterin levels of untreated B6 and B6-lpr/lpr mice were about equal and lower than those of MRL mice. During the first three months of DPH treatment, persistently elevated biopterin levels were observed in B6 and to a lesser degree in MRL mice, and alternately elevated and control levels in B6-lpr/lpr mice. Thereafter, the effects faded in all strains. Results show that long-term DPH treatment causes only minor lymphoid organ weight changes in B6 and MRL mice, but causes a clear reduction of the lymphadenopathy and (auto)antibody formation in B6-lpr/lpr mice. Observed changes could not be related to altered biopterin excretion indicating that the latter is an inappropriate marker of murine autoimmune disease.

Detección de factor reumatoide por nefelometría en pacientes de un centro hospitalario en México / Determination of rheumatoid factor by nephelometry in a mexican medical center

Se estudió la presencia de factor reumatoide por nefelometría en 6.264 muestras consecutivas de suero, provenientes de pacientes captados de una población hospitalaria de 1988 a 1990. El 52 por ciento de estos pacientes presentaron FR positivo, en comparación con el 1,4 por ciento en el grupo control. Se obtuvieron resultados negativos en 48 por ciento de los sueros captados en población hospitalaria, en comparación al 98 por ciento de muestras negativas en una población control (p<0,000001), captada en una clínica de atención de primer nivel. Ambos grupos mostraron características demográficas y socioeconómicas similares y los resultados no dependieron del número de pacientes estudiados, cuando se divieron a los resultados en negativos (<30Ul/ml), intermedios (30-110 Ul/ml) y altos (>30 Ul/ml), se encontró una frecuencia similar de los sueros positivos (20-27 por ciento), especialmente cuando se evaluaron los límites intermedios. Estos niveles variaron muy poco en los diferentes períodos del estudio. Los resultados de factor reumatoide pueden mostrar gran variación de acuerdo a la edad, sexo, antecedentes inmunogenéticos, o antecedentes patológicos en diferentes poblaciones. Los pacientes de población hospitalaria, muestran una alta incidencia de padecimientos infecciosos e inflamatorios crónicos que pueden cursar con pruebas positivas para factor reumatoide, y esto puede explicar su alta frecuencia en comparación con el grupo control. Falta definir la participación de ésta y otras variables en estudios posteriores