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ID PMID Title PublicationDate abstract
9367846 Fas/Fas ligand interaction regulates cytotoxicity of CD4+ T cells against staphylococcal e 1997 Oct 29 Infiltration of activated CD4+ T cells and apoptosis of endothelial cells are present in the synovium of rheumatoid arthritis (RA). Using staphylococcal enterotoxin B (SEB) as an antigen, we examined the possible role of antigen (Ag)-dependent activation of CD4+ T cells by endothelial cells, in inducing endothelial cell apoptosis. The human endothelial cell line, EA.hy926 cells, was cultured with or without interferon-gamma (IFN-gamma) and further incubated with CD4+ T cells in the presence or absence of SEB. After this cocultivation, the cytotoxicity and Fas ligand (FasL) expression of CD4+ T cells were examined. A small percentage of EA.hy926 cells expressed HLA-DR and -DQ, and this expression was significantly augmented after IFN-gamma stimulation. Anti-Fas IgM-induced apoptosis was exhibited by both unstimulated and IFN-gamma-stimulated EA.hy926 cells. Cytotoxicity of CD4+ T cells toward SEB-pulsed unstimulated EA.hy926 cells was detected. Furthermore, when CD4+ T cells were incubated with IFN-gamma-stimulated, SEB-pulsed EA.hy926 cells with augmented HLA-DR and -DQ expression, this cytotoxicity was more significant. The addition of anti-HLA-DR and -DQ monoclonal antibodies (mAbs) or human Fas chimeric protein (hFas-Fc) reduced the cytotoxicity. FasL expression was induced in CD4+ T cells cocultured with SEB-pulsed EA.hy926 cells, especially when the EA.hy926 cells were IFN-gamma-stimulated. Furthermore, the addition of mAbs against CD54 and CD58 inhibited both the cytotoxicity and FasL expression of CD4+ T cells induced by SEB-pulsed EA.hy926 cells, indicating the importance of costimulatory molecules on EA.hy926 cells in activating CD4+ T cells. Our results suggest that CD4+ T cells are activated by endothelial cells in an Ag-dependent manner and subsequently express FasL, which induces Fas-mediated apoptosis of endothelial cells. This phenomenon may counteract the growth of RA synovium by inhibiting the proliferation of endothelial cells.
9151795 Treatment with depleting CD4 monoclonal antibody results in a preferential loss of circula 1997 May 1 CD4(pos) TH1 T cells are considered to play a central role in a number of human autoimmune diseases such as rheumatoid arthritis (RA) and multiple sclerosis. Experimental treatment protocols aimed at selectively eliminating CD4(pos) T cells thus far have yielded disappointing clinical results. Here we analyzed phenotype and function of circulating T cells in multiple sclerosis patients treated with the chimeric CD4 mAb cM-T412 in a randomized, double-blind, placebo-controlled, magnetic resonance imaging-monitored phase II trial. Treatment resulted in a long-lasting depletion of CD4(pos) T cells but did not affect CD8(pos) T cell numbers. Analysis of CD4(pos) subpopulations showed that unprimed, CD45RA(pos)/R0(neg) lymphocytes were approximately three times more sensitive to the mAb than primed, CD45RA(neg)/R0(pos) T cells. Notably, within the CD45RA(pos) subset, T cells with phenotypic evidence of prior activation, i.e., expressing Fas, were relatively insensitive to cM-T412, compared with Fas(neg) cells. Remarkably, while a decrease in the number of IL-4-producing T helper 2 (TH2)-type cells in the anti-CD4 treated group was observed, numbers of IFN-gamma-producing T helper 1 (TH1)-type cells remained stable, resulting in a significant increase in the TH1/TH2 ratio. Our data show that treatment with depleting CD4 mAb does not eliminate the cells most strongly involved in the disease process, i.e., primed, IFN-gamma-producing TH1-type cells, and may therefore give an explanation for the lack of beneficial clinical effects of depleting CD4 mAb in human chronic autoimmune disease.
11727506 VEGF antagonists. 2001 Jul The majority of cancer have an absolute requirement for angiogenesis, the process by which new blood vessels are formed. The most potent angiogenic cytokine is vascular endothelial growth factor (VEGF) and there has been substantial research into the development of VEGF/VEGF receptor (VEGFR) antagonists. To date these strategies have included gene therapy techniques that deliver antisense oligonucleotides, soluble VEGFRs that function in a dominant negative fashion and ribozymes. Additional strategies have included the development of receptor tyrosine kinase (RTK) inhibitors and monoclonal antibodies (mAbs) directed against VEGF or the signalling receptor. The most promising agents appear to be the monoclonal anti-VEGF antibodies and the RTK inhibitors as these have demonstrated broad spectrum antitumour activity in vivo and single agent activity in early phase clinical trials in patients with advanced pre-treated breast and colorectal carcinoma and Kaposi's sarcoma. The RTK inhibitors are of particular interest as they can be administered by mouth. Collation of the early clinical trial data suggests that VEGF antagonists are largely well-tolerated but may be associated with vascular toxicities such as haemorrhage and thromboembolic events. Combination studies of chemotherapy and VEGF antagonists are underway but the benefit of these regimens will need to be established in adequately powered Phase III studies. Potentially these agents may play a role in the treatment of both early (adjuvant) and advanced cancer. The efficacy of the drugs will be explored in a number of non-malignant conditions including rheumatoid arthritis (RA), psoriasis, diabetic retinopathy and possibly as non-steroidal contraceptives but the overall clinical development of these agents can only be optimised if appropriate biological end points are identified and incorporated into clinical trials.
11529938 Sulphasalazine inhibits macrophage activation: inhibitory effects on inducible nitric oxid 2001 Aug The anti-inflammatory agent sulphasalazine is an important component of several treatment regimens in the therapy of ulcerative colitis, Crohn's disease and rheumatoid arthritis. Sulphasalazine has many immunomodulatory actions, including modulation of the function of a variety of cell types, such as lymphocytes, natural killer cells, epithelial cells and mast cells. However, the effect of this agent on macrophage (M phi) function has not been characterized in detail. In the present study, we investigated the effect of sulphasalazine and two related compounds - sulphapyridine and 5-aminosalicylic acid - on M phi activation induced by bacterial lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma). In J774 M phi stimulated with LPS (10 microg/ml) and IFN-gamma (100 U/ml), sulphasalazine (50-500 microM) suppressed nitric oxide (NO) production in a concentration-dependent manner. The expression of the inducible NO synthase (iNOS) was suppressed by sulphasalazine at 500 microM. Sulphasalazine inhibited the LPS/IFN-gamma-induced production of both interleukin-12 (IL-12) p40 and p70. The suppression of both NO and IL-12 production by sulphasalazine was superior to that by either sulphapyridine or 5-aminosalicylic acid. Although the combination of LPS and IFN-gamma induced a rapid expression of the active forms of p38 and p42/44 mitogen-activated protein kinases and c-Jun terminal kinase, sulphasalazine failed to interfere with the activation of any of these kinases. Finally, sulphasalazine suppressed the IFN-gamma-induced expression of major histocompatibility complex class II. These results demonstrate that the M phi is an important target of the immunosuppressive effect of sulphasalazine.
9714181 A modified human alpha 2-macroglobulin derivative that binds tumor necrosis factor-alpha a 1998 Aug The plasma protein alpha 2-macroglobulin (alpha 2M) has been reported to bind the proinflammatory cytokines, tumor necrosis factor-alpha (TNF-alpha) and interleukin 1 beta (IL-1 beta), which play a central role in the pathogenesis of chronic inflammatory disorders, including Crohn's disease and rheumatoid arthritis. In this study, we chemically modified alpha 2M to stabilize a conformation of the protein (termed MAC, Macroglobulin Activated for Cytokine binding) with greatly increased TNF-alpha- and IL-1 beta-binding activity. The equilibrium dissociation constant (KD) for the binding of TNF-alpha to MAC was 80 +/- 20 nM, reflecting a 100-fold increase in affinity compared with native alpha 2M. To test the ability of MAC to neutralize proinflammatory cytokines in vivo, we treated mice with lipopolysaccharide (LPS) by intravenous injection. When MAC (2.5 mg) was administered by intraperitoneal injection 1 hour before the LPS, 12 of 12 mice survived and were without signs of toxicity at 5 days. None of the mice survived in the untreated control group (0/26) or in the group treated with 2.5 mg of unmodified alpha 2M (0/4). MAC also prevented the large increase in expression of inducible nitric oxide synthase in the liver, kidneys, and heart of LPS-treated mice. A novel property of MAC, compared with previously studied anticytokine agents, was its ability to reverse LPS toxicity in 12 of 24 mice when administered after the plasma level of TNF-alpha was elevated. These studies demonstrate that a naturally occurring protein, alpha 2M, can be modified so that it acquires the properties of clinically active monoclonal antibodies. Thus, MAC may have therapeutic potential in the control of chronic inflammatory disorders.
9653431 Catastrophic antiphospholipid syndrome. Clinical and laboratory features of 50 patients. 1998 May We analyzed the clinical and laboratory characteristics of 50 patients with catastrophic antiphospholipid syndrome (APS) (5 from our clinics and 45 from a MEDLINE computer-assisted review of the literature from 1992 through 1996). Thirty-three (66%) patients were female and 17 (34%) were male. Twenty-eight (56%) patients had primary APS, 15 (30%) had defined systemic lupus erythematosus (SLE), 6 (12%) had "lupus-like" syndrome, and 1 (2%) had rheumatoid arthritis. Mean age of patients in this series was 38 +/- 14 years (range, 11-74 yr). Three (6%) patients developed the clinical picture of catastrophic APS under the age of 15 years, and 11 (22%) were 50 years old or more. In 11 (22%) patients, precipitating factors contributed to the development of catastrophic APS (infections in 3, drugs in 3, minor surgical procedures in 3, anticoagulation withdrawal in 2, and hysterectomy in 1). The presentation of the acute multi-organ failure was usually complex, involving multiple organs simultaneously or in a very short period of time. The majority of patients manifested microangiopathy--that is, occlusive vascular disease affecting predominantly small vessels of organs, particularly kidney, lungs, brain, heart, and liver--with a minority of patients experiencing only large vessel occlusions. Thrombocytopenia was reported in 34 (68%) patients, hemolytic anemia in 13 (26%), disseminated intravascular coagulation in 14 (28%), and schistocytes in 7 (14%). The following antibodies were detected: lupus anticoagulant (94%), anticardiolipin antibodies (94%), anti-dsDNA (87% of patients with SLE), antinuclear antibodies (58%), anti-Ro/SS-A (8%), anti-RNP (8%), and anti-La/SS-B (2%). Anticoagulation was used in 70% of the patients, steroids in 70%, plasmapheresis in 40%, cyclophosphamide in 34%, intravenous gammaglobulins in 16%, and splenectomy in 4%. Most patients, however, received a combination of nonsurgical therapies. Death occurred in 25 of the 50 (50%) patients. In most, cardiac problems seemed to be the major cause of death. In several of these, respiratory failure was also present, usually due to acute respiratory distress syndrome and diffuse alveolar hemorrhage. Among the 20 patients who received the combination of anticoagulation, steroids, and plasmapheresis or intravenous gammaglobulins, recovery occurred in 14 (70%) patients. The use of ancrod and defibrotide appeared to be effective in the 2 respective patients in whom they were used.
9351807 The crystal structure of vascular endothelial growth factor (VEGF) refined to 1.93 A resol 1997 Oct 15 BACKGROUND: Vascular endothelial growth factor (VEGF) is an endothelial cell-specific angiogenic and vasculogenic mitogen. VEGF also plays a role in pathogenic vascularization which is associated with a number of clinical disorders, including cancer and rheumatoid arthritis. The development of VEGF antagonists, which prevent the interaction of VEGF with its receptor, may be important for the treatment of such disorders. VEGF is a homodimeric member of the cystine knot growth factor superfamily, showing greatest similarity to platelet-derived growth factor (PDGF). VEGF binds to two different tyrosine kinase receptors, kinase domain receptor (KDR) and Fms-like tyrosine kinase 1 (Flt-1), and a number of VEGF homologs are known with distinct patterns of specificity for these same receptors. The structure of VEGF will help define the location of the receptor-binding site, and shed light on the differences in specificity and cross-reactivity among the VEGF homologs. RESULTS: We have determined the crystal structure of the receptor-binding domain of VEGF at 1.93 A resolution in a triclinic space group containing eight monomers in the asymmetric unit. Superposition of the eight copies of VEGF shows that the beta-sheet core regions of the monomers are very similar, with slightly greater differences in most loop regions. For one loop, the different copies represent different snapshots of a concerted motion. Mutagenesis mapping shows that this loop is part of the receptor-binding site of VEGF. CONCLUSIONS: A comparison of the eight independent copies of VEGF in the asymmetric unit indicates the conformational space sampled by the protein in solution; the root mean square differences observed are similar to those seen in ensembles of the highest precision NMR structures. Mapping the receptor-binding determinants on a multiple sequence alignment of VEGF homologs, suggests the differences in specificity towards KDR and Flt-1 may derive from both sequence variation and changes in the flexibility of binding loops. The structure can also be used to predict possible receptor-binding determinants for related cystine knot growth factors, such as PDGF.
11838747 Fetal tissue/organ transplant in HLA-randomized adult vascular subcutaneous axillary folds 2001 BACKGROUND: In the year 1902, the first successful experimental organ transplant, i.e., an autotransplant of a dog's kidney from its normal position to the vessels of the neck, which resulted in some urine flow, was performed in the Vienna Physiology Institute under the direction of Hofrath Exner by Dr. Emerich Ullman (1861-1937). Since then, the art of transplant surgery has come a long way in establishing itself as an important discipline with the support disciplines of immunology, molecular biology, etc., for the restoration of a failing organ. Today there is a major discrepancy in the demand and supply of organ grafts. The aim of the present study is to see whether fetal organ and tissue, with its intrinsic advantages of hypo-antigenicity, can survive in a HLA and sex-randomized host in a surgically prepared vascular subcutaneous axillary fold, without any immunosuppressive support. We have earlier reported two cases of fetal thymic transplant, collected from consenting mothers undergoing hysterotomy and ligation. MATERIALS AND METHODS: Fourteen cases were recruited for the present study after thorough informed consent and approval by the Ethical Committee of the Project. Of these, five patients were suffering from advanced cancer, three from diabetic gangrene, three from ischaemic heart disease and three from rheumatoid arthritis, liver abscess and disc prolapse. The ages of the patients varied from 39 to 82 years. Six fetal thymuses, three fetal liver tissues, three fetal cardiac tissues, one fetal pancreas and one fetal lung tissue were transplanted. All the fetuses were dissected and the selected tissues/organs were transplanted within one to three minutes after collecting them from the consenting mothers undergoing hysterotomy and ligation. The fetal tissue graft was placed in a surgically prepared subcutaneous vascular axillary fold, 2x1 cm, under local anaesthesia in the consenting adult recipient. Sequential Hb, Tc, Dc, ESR were done to see the impact of the transplant on the host system. After one month, the transplanted fetal tissue was taken out by an elliptical incision and the tissue was processed for histological staining. RESULTS AND ANALYSIS: All the 14 patients tolerated the transplant procedure well. There was no fever, intractable pain or any other specific serious side-effect justifying removal of the transplant earlier. There was no discharge from the incision site and the healing and scar were by and large normal. There was no unusual leucocytosis or lymphocytosis. The serial histological study did not suggest features of transplant rejection. DISCUSSION AND CONCLUSION: Pregnancy and neoplasm are two outstanding examples of natural tolerance to homograft. In both cases, blocking antibody has an important role in the phenomenon of immunotolerance. From our experiments mentioned above transplantation and our earlier reported studies, we believe that the hypo-antigenic fetal tissue has distinct advantages over adult tissue for transplant purposes.
11732857 IL-1beta-induced expression of matrix metalloproteinases and gliostatin/platelet-derived e 2001 Oct The purpose of this study was to examine how chondrocytes are involved in the molecular mechanism of inflammation in rheumatoid arthritis (RA). A chondrosarcoma cell line (OUMS-27) was cultured and treated with interleukin-1beta (IL-1beta). Changes in the expression levels of matrix metalloproteinase-1 (MMP-1), metalloproteinase-13 (MMP-13), and gliostatin/platelet-derived endothelial cell growth factor (GLS/PD-ECGF) were assessed by reverse transcription polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assays. IL-1beta induced the expressions of MMP-1, MMP-13, and GLS mRNAs and proteins in a dose-dependent manner. Selective inhibition of the p38 mitogen-activated protein kinase (p38 MAPK) pathway with SB 203580 and SB 202190 blocked the expression of MMP-1, MMP-13, and GLS more strongly than selective in hibition of the extracellular signal-regulated kinase 1 and 2 (ERK1/2) pathway by PD 98059. These findings suggest that chondrocytes may intensify cartilage destruction and inflammation in RA by the induction of MMP-1, MMP-13, and GLS by IL-1beta and that the p38 MAPK pathway plays an important role in these inductions.
11319753 IL-1- and TNF-induced bone resorption is mediated by p38 mitogen activated protein kinase. 2001 Jun We have previously shown that p38 mitogen-activated protein kinase (MAPK) inhibitors, which block the production and action of inflammatory cytokines such as tumor necrosis factor (TNF) and interleukin-1 (IL-1), are effective in models of bone and cartilage degradation. To further investigate the role of p38 MAPK, we have studied its activation in osteoblasts and chondrocytes, following treatment with a panel of proinflammatory and osteotropic agents. In osteoblasts, significant activation of p38 MAPK was observed following treatment with IL-1 and TNF, but not parathyroid hormone, transforming growth factor-beta (TGF-beta), 1,25(OH)(2)D(3), insulin-like growth factor-1 (IGF-1), or IGF-II. Similar results were obtained using primary bovine chondrocytes and an SV40-immortalized human chondrocyte cell line, T/C28A4. SB 203580, a selective inhibitor of p38 MAPK, inhibited IL-1 and TNF-induced p38 MAPK activity and IL-6 production (IC(50)s 0.3--0.5 microM) in osteoblasts and chondrocytes. In addition, IL-1 and TNF also activated p38 MAPK in fetal rat long bones and p38 MAPK inhibitors inhibited IL-1- and TNF-stimulated bone resorption in vitro in a dose-dependent manner (IC(50)s 0.3--1 microM). These data support the contention that p38 MAPK plays a central role in regulating the production of, and responsiveness to, proinflammatory cytokines in bone and cartilage. Furthermore, the strong correlation between inhibition of kinase activity and IL-1 and TNF-stimulated biological responses indicates that selective inhibition of the p38 MAPK pathway may have therapeutic utility in joint diseases such as rheumatoid arthritis (RA).
10790090 Serologic diagnosis of Lyme borreliosis by using enzyme-linked immunosorbent assays with r 2000 May Class-specific enzyme-linked immunosorbent assays (ELISAs) with purified recombinant antigens of Borrelia burgdorferi sensu stricto and Western blot analyses with whole cells of this spirochete were used to test human sera to determine which antigens were diagnostically important. In analyses for immunoglobulin M (IgM) antibodies, 14 (82%) of 17 serum samples from persons who had erythema migrans reacted positively by an ELISA with one or more recombinant antigens. There was frequent antibody reactivity to protein 41-G (p41-G), outer surface protein C (OspC), and OspF antigens. In an ELISA for IgG antibodies, 13 (87%) of 15 serum samples had antibodies to recombinant antigens; reactivity to p22, p39, p41-G, OspC, and OspF antigens was frequent. By both ELISAs, serum specimens positive for OspB, OspE, and p37 were uncommon. Analyses of sera obtained from persons who were suspected of having human granulocytic ehrlichiosis (HGE) but who lacked antibodies to ehrlichiae revealed IgM antibodies to all recombinant antigens of B. burgdorferi except OspB and IgG antibodies to all antigens except OspE. Immunoblotting of sera from the study group of individuals suspected of having HGE reaffirmed antibody reactivity to multiple antigens of B. burgdorferi. There was minor cross-reactivity when sera from healthy subjects or persons who had syphilis, oral infections, or rheumatoid arthritis were tested by ELISAs with p37, p41-G, OspB, OspC, OspE, and OspF antigens. Although the results of class-specific ELISAs with recombinant antigens were comparable to those recorded for assays with whole-cell antigen and for individuals with confirmed clinical diagnoses of Lyme borreliosis, immunoblotting is still advised as an adjunct procedure, particularly when there are low antibody titers by an ELISA.
10695405 [Nation-wide survey for the treatment with cyclosporin A of interstitial pneumonia associa 1999 Dec OBJECTIVES: This study was performed to investigate the efficacy and safety of cyclosporin A (CsA) for the treatment of interstitial pneumonia (IP) associated with collagen diseases in Japan. METHODS: Questionnaires were sent to 36 hospitals specializing in collagen diseases. RESULTS: Fifty-eight patients (7 polymyositis (PM), 19 dermatomyositis (DM), 7 systemic sclerosis (SSc), 7 rheumatoid arthritis (RA), 2 mixed connective tissue disease (MCTD), 1 systemic lupus erythematosus (SLE) and 1 Sjögren's syndrome (SS), 1 RA + SSc, 2 PM + SSc, 1 DM + SLE, and 10 idiopathic interstitial pneumonia (IIP) with IP were treated with CsA at 14 hospitals. IP was classified into the acute or chronic type. In the PM/DM group (7 PM, 19 DM, 2 PM + SSC, 1 DM + SLE), 65.5% were the acute type. In the other collagen disease group (7 SSc, 7 RA, 2 MCTD, 1 SLE, 1 SS, and 1 RA + SSc) and IIP group, 36.8% and 50% were the acute type, respectively. Mean dosages of CsA were 3.7 +/- 1.3 mg/kg/day for the PM/DM group, 3.0 +/- 1.0 for the other collagen disease group, and 3.8 +/- 4.8 for the IIP group. Oral corticosteroids were administered in combination with CsA in 100, 73.7, and 70% of the patients with PM/DM, other collagen disease, and IIP groups, respectively. CsA was effective for 72.2, 33.3, and 25% of the acute IP cases in the PM/DM, other collagen disease, and IIP groups, respectively. CsA was effective for 50.0, 50.0, and 60.0% of chronic IP cases in the PM/DM, other collagen disease, and IIP groups, respectively. Twenty-three adverse effects were observed, but most of them ameliorated upon withdrawal or reduction of the CsA dose. CONCLUSION: CsA is effective for the treatment of acute type IP associated with collagen diseases, especially PM/DM. To perform a prospective multi-center trial, standards for the recruitment of patients, efficacy assessments, and trial course and treatment should be determined carefully.
10525088 RWJ 67657, a potent, orally active inhibitor of p38 mitogen-activated protein kinase. 1999 Nov Tumor necrosis factor-alpha (TNF-alpha), a cytokine secreted by activated monocytes/macrophages and T lymphocytes, has been implicated in several disease states, including rheumatoid arthritis, inflammatory bowel disease, septic shock, and osteoporosis. Monocyte/macrophage production of TNF-alpha is dependent on the mitogen-activated protein kinase p38. RWJ 67657 (4-[4-(4-fluorophenyl)-1-(3-phenylpropyl)-5-(4-pyridinyl)-1H-imidazol -2-yl]-3-butyn-1-ol) inhibited the release of TNF-alpha by lipopolysaccharide (a monocyte stimulus)-treated human peripheral blood mononuclear cells with an IC(50) of 3 nM, as well as the release of TNF-alpha from peripheral blood mononuclear cells treated with the superantigen staphylococcal enterotoxin B (a T cell stimulus), with an IC(50) value of 13 nM. This compound was approximately 10-fold more potent than the literature standard p38 kinase inhibitor SB 203580 in all p38 dependent in vitro systems tested. RWJ 67657 inhibited the enzymatic activity of recombinant p38alpha and beta, but not gamma or delta, in vitro and had no significant activity against a variety of other enzymes. In contrast, SB 203580 significantly inhibited the tyrosine kinases p56 lck and c-src (IC(50) = 5 microM). RWJ 67657 did not inhibit T cell production of interleukin-2 or interferon-gamma and did not inhibit T cell proliferation in response to mitogens. RWJ 67657 inhibited TNF-alpha production in lipopolysaccharide-injected mice (87% inhibition at 50 mg/kg) and in rats (91% inhibition at 25 mg/kg) after oral administration. Based on these favorable biological properties, RWJ 67657 may have use as a treatment for inflammatory diseases.
10443681 Cleavage of recombinant human corticotropin-releasing factor (CRF)-binding protein produce 1999 Aug CRF is both a peripheral and a central mediator of inflammation, the activity of which is modified by the presence of a 37-kDa binding protein (CRF-BP). The objective of this study was to measure and characterize this protein in the synovial fluid of rheumatoid arthritis patients and to observe the effects of this inflammatory condition on its structure and properties. Measured by immunoradiometric assays, the mean CRF-BP concentration in synovial fluid from 27 arthritic patients was 0.51 nmol/L (SD = 0.24 nmol/L); that for CRF was 6.31 pmol/L. The mean plasma concentration of CRF-BP in 24 control subjects was 1.38 nmol/L (SD = 0.35 nmol/L) and that for 10 arthritic patients was 2.89 nmol/L (SD = 0.84 nmol/L). Synovial fluids were found by immunoblotting to contain intact CRF-BP and a 10-kDa C-terminal CRF-BP fragment; synovial fluid from healthy controls was not examined. We previously reported that after purification of recombinant CRF-BP, spontaneous cleavage frequently occurs, resulting in a 27-kDa N-terminal and a 10-kDa C-terminal fragment. Because concentrations of native CRF-BP in synovial fluid were insufficient to study the effects of cleavage on ligand binding, they were determined using recombinant human CRF-BP. Tryptophan excitation fluorescence spectra of intact and cleaved recombinant CRF-BP revealed that cleavage was accompanied by conformational change in the N-terminal fragment, leading to exposure of the sole tryptophan residue to polar molecules (emission peak shift from 310 to 250 nm). Using gel filtration chromatography to separate the N- and C-terminal fragments, it was found that the N-terminal fragment of the recombinant protein bound human CRF, although dimerization was somewhat impaired. The C-terminal fragment did not bind CRF. Scatchard analysis confirmed that the affinity of both intact and cleaved CRF-BP for CRF was 1 x 10(10) L/mol. We conclude that synovial fluid contains intact CRF-BP in molar excess to CRF and fragmented CRF-BP. The significance of cleavage and the role of cleavage products have yet to be determined, although they may represent the generation of a novel bioactivity.
10048640 A "case control" study on the rôle of HLA DR4 in severe periodontitis and rapidly progres 1999 Feb HLA DR4 antigens have been considered as a risk factor in periodontal disease. The aim of this "case control" study was to verify and to provide fuller clarification of such data. "Cases" or patients had to be aged between 20 and 48 years. They presented at least 5 sites spread over several teeth with an attachment loss equal or greater than 6 mm, and 10 sites spread over several teeth with periodontal pockets equal to or greater than 5 mm. Verification with a WHO probe showed an individual CPITN score of 4. Moreover, subjects whose average CPITN score for the 6 sextants was less than 3 were excluded from the study. Among these "severe periodontitis" patients, a subgroup was distinguished composed of subjects aged 20-35 years who presented, in accordance with the cases by Katz and co-workers, 5 or more teeth showing pocket depths of 6 mm or more. The dental chartings of these subjects showed an attachment loss of more than 3 mm on certain teeth over an inter-exam period of 1-3 years. They all displayed obvious loss of bony support in the affected sites. This constituted the "rapidly progressive periodontitis" subgroup. The "controls" were all over 20 years of age, and it was clinically verified that they were free of periodontal disease. There were 48 "cases" and 55 "controls". HLA typing of patients and controls was performed using "sequence oligoprobe hybridization after polymerase chain reaction" in accordance with the 11th International Workshop. This method allowed the detection of DR4 alleles as well as DR4 subtypes. The ethno-geographic origin of the subjects, considered as a confounding variable, was neutralized by stratified analysis. Subtypes 0401, 0404, 0405 and 0408 tended to be more frequent (p=0.08) in the cases (Severe Periodontitis). Focusing on analysis of "rapidly progressive periodontitis" in subjects aged 20-35 years, a very significant Mantel-Haenszel chi2 was obtained (p=0.0058) which led to a Mantel-Haenszel standardized odds ratio (OR) equal to 17. The 95% confidence interval was 1.03<0.R.<180.10. In conclusion, this supports previous reports and gives further clarification: in particular subtypes 0401, 0404, 0405 and 0408 can be considered as a risk factor for "rapidly progressive periodontitis". It should be noted that these determinants have been implicated in rheumatoid arthritis.
9972976 Antipolymer antibody reactivity in a subset of patients with fibromyalgia correlates with 1999 Feb OBJECTIVE: To determine the prevalence of antipolymer antibodies (APA) in patients with fibromyalgia (FM) and autoimmune disease control groups and to determine if the presence of these antibodies correlates with severity in patients with FM. METHODS: Sera from patients with FM (n = 47), osteoarthritis (OA) (n = 16), and rheumatoid arthritis (RA) (n = 13) were analyzed. Patients with implants of any kind and patients with concurrent autoimmune conditions were excluded from study. Banked sera from autoimmune disease controls including poly/dermatomyosis (n = 15), RA (n = 30), systemic lupus erythmatosus (SLE) (n = 30), and systemic sclerosis (SSc) (n = 30) were also analyzed. To determine if seroreactivity correlates with severity, banked sera from patients with FM assessed as severe (n = 28) or mild (n = 37) and from controls (n = 21) were assayed. RESULTS: Following analysis, the prevalence of seroreactivity was found to be higher in patients with FM (22/47, 47%) compared to patients with OA (3/16, 19%; p<0.1) or RA (1/13, 8%; p<0.05) and the autoimmune disease control sera from poly/dermatomyosis (2/15, 13%; p<0.05), and patients with RA (3/30, 10%; p<0.01), SLE (1/30, 3%; p<0.01), and SSc (1/30, 3%; p<0.01). The prevalence of APA seroreactivity was also significantly higher in patients with severe FM (17/28, 61%) compared to patients with mild FM (11/37, 30%; p<0.05) and controls (4/21, 19%; p<0.01). In addition, both mean threshold and mean tolerance dolorimetry scores were significantly lower in the seropositive patients with mild FM (1.33+/-0.21, 1.95+/-0.25, respectively) compared to the seronegative patients (1.83+/-0.08, 2.53+/-0.11; p<0.05 for both comparisons, respectively). CONCLUSION: These results reveal that an immunological response, production of anti-polymer antibodies, is associated with a subset of patients with FM. The results also suggest that the APA assay may be an objective marker in the diagnosis and assessment of FM and may provide additional avenues of investigation into the pathophysiological processes involved in FM.
9263156 Nucleotide pyrophosphohydrolase in human synovial fluid. 1997 Aug OBJECTIVE: To identify the molecular forms of ectonucleotide pyrophosphohydrolase (NTPPHase) in human synovial fluid (SF). METHODS: We examined synovial fluids from 32 patients with various joint diseases [10 calcium pyrophosphate dihydrate (CPPD) deposition disease; 7 osteoarthritis (OA); 6 rheumatoid arthritis (RA); 3 after total knee arthroplasty (TKA); 6 olecranon bursa] and 3 normal joint fluids. Joint fluids were analyzed after sequential centrifugation for NTPPHase activity and by Western blot using polyclonal antibodies against 127 kDa porcine articular cartilage vesicle-associated NTPPHase and against PC-1 and 58 kDa, 2 other ecto-NTPPHases. Lysate from human synoviocytes, porcine chondrocytes, and their conditioned media were examined using antibodies to these ecto-NTPPHases. Radiographs of joints from which fluid was obtained were graded for degenerative changes 0-4 using a standard method. RESULTS: NTPPHase activity was found in all pathological and normal SF tested and correlated with the degree of radiographic degeneration (r = 0.55, p < 0.05). NTPPHase specific activity in ultracentrifugation pellets was highest in CPPD deposition disease fluids (p < 0.05). 127 kDa enzyme was found in both sedimentable and soluble fractions from CPPD, OA, TKA, and normal fluids, and was extensively degraded in all inflammatory fluids. Intact 115 kDa PC-1 was found only in the 2 CPPD fluids with the highest NTPPHase activity. 58 kDa enzyme was found in most fluids, predominantly in the soluble fraction. 127 kDa protein was identified in human synoviocyte conditioned media but not in cell lysate, while PC-1 and 58 kDa proteins were found in the cell lysate but not in the conditioned media. CONCLUSION: There was no disease specific association with any one ecto-NTPPHase. Total enzyme activity correlated with the degree of degenerative change. The specific activity of pelletable 127 kDa enzyme was higher in fluids containing CPPD crystals. All 3 ecto-NTPPHases or their presumed degradation products were detectable in some pathologic and normal fluids. A 200 kDa reactive band often accompanied reactivity to the 127 kDa enzyme. PC-1 and 127 kDa proteins were extensively degraded in inflammatory SF, while 58 kDa protein was not. The relative contribution of each of these enzymes to inorganic pyrophosphate production by human joint tissues remains unclear.
9234597 Anemia of chronic disorders in systemic autoimmune diseases. 1997 May BACKGROUND AND OBJECTIVE: Anemia of chronic disorders (ACD) is a mild to moderate anemia characterized by decreased serum iron, decreased total iron-binding capacity and increased iron stores that occurs in a wide variety of diseases including cancer, chronic infections and inflammatory disorders. The reason for this review is two-fold. First, systemic autoimmune diseases are frequently characterized by ACD. Second, advances in our knowledge of the pathophysiology of systemic autoimmune diseases as well as pathogenesis and treatment of ACD have so far been dealt with separately. Consequently, the approach to the evaluation of ACD in systemic autoimmune disorders has usually been either immunology- or hematology-oriented. The aim of this review is to integrate the pertinent information from both these fields in order to arrive at a more complete understanding of a problem common to hematologists and immunologists. INFORMATION SOURCES: The articles reviewed have been published in journals listed in the Science Citation Index and Medline. In addition, the authors have a vast experience in the field of hematology and are actively working in the field of systemic autoimmune disorders. STATE OF ART AND PERSPECTIVES: ACD is a parameter of disease activity in systemic autoimmune diseases. The severe inflammatory stimuli responsible for the pathophysiology of these disorders lead to several systemic changes (referred to as chronic active phase response) through which the organism tries to cope with chronic tissue injuries. These reactions are brought about by inflammation-associated cytokines, like IL-6, IL-1, TNF alpha, TGF beta that regulate hepatic synthesis of acute phase proteins. Many cytokines involved in chronic acute phase response, including IL-1, TNF alpha, TGF beta, have an inhibitory activity on erythroid colony formation in vitro. In addition, circulating TNF alpha is elevated in rheumatoid arthritis (RA), IL-1 beta serum levels are significantly increased in RA with ACD and RA patients treated in vivo with antibodies (Abs) to TNF alpha show disease improvement, including an increase in Hb values. Reduced erythropoietin (EPO) activity, usually the result of reduced production, plays a role in the pathogenesis of ACD observed in systemic autoimmune diseases. Both the production and the action of EPO may fall under the control of IL-1 and IFN-gamma. The most controversial and stimulating aspect of the pathogenesis of ACD in systemic autoimmune disorders is the role of iron metabolism and nitric oxide (NO), which contributes to the regulation of iron cellular metabolism. Both iron deficiency and iron overload may influence the proliferation of B and T lymphocytes and differentially affect T helper (TH)-1 and TH-2 lymphocytes. Furthermore, TH-1 cytokines stimulate and TH-2-type cytokines inhibit NO production. For these reasons, cell-mediated immunity may be expected to have influence on NO synthesis and on the mechanisms leading to iron accumulation in the reticuloendothelial system.
10925319 B and T cell responses to the spliceosomal heterogeneous nuclear ribonucleoproteins A2 and 2000 Aug 15 Autoantibodies directed against spliceosomal heterogeneous nuclear ribonucleoproteins (hnRNPs) are a typical feature of rheumatoid arthritis, systemic lupus erythematosus, and mixed-connective tissue disease. With the aim of investigating a potential pathogenic role of these Abs, we have studied the Ab response to A2/B1 hnRNPs in different murine models of lupus. The specificity of anti-A2/B1 Abs was tested with a series of 14 overlapping synthetic peptides covering the region 1-206 of A2 that contains most of the epitopes recognized by patients' Abs. A major epitope recognized very early during the course of the disease by Abs from most of MRL lpr/lpr mice but not from other lupus mice and from mice of different MHC haplotypes immunized against B1 was identified in residues 50-70. This peptide contains a highly conserved sequence RGFGFVTF also present in other hnRNPs and small nuclear ribonucleoproteins. Abs reacting with a second A2 epitope identified in residues 35-55 were detectable several weeks later, suggesting an intramolecular B cell epitope spreading during the course of the disease. We identified several T cell epitopes within the region 35-175 that generated an effective Th cell response with IL-2 and IFN-gamma secretion in nonautoimmune CBA/J mice sharing the same MHC haplotype H-2k as MRL/lpr mice. None of the peptides stimulated T cells primed in vivo with B1. Because Abs to peptide 50-70 were detected significantly earlier than Abs reacting with other A2 peptides and the protein itself, it is possible that within the protein, this segment contains residues playing an initiator role in the induction of the anti-A2/B1 and antispliceosome Ab response.
10759759 IL-17 regulates gene expression and protein synthesis of the complement system, C3 and fac 2000 Apr Human IL-17 is a cytokine secreted by CD4+-activated memory T cells with the profile of effects of a Th1 cytokine. The effects of IL-17 on many cellular constituents of joints suggest that it may participate in inflammatory joint diseases. Proteins of the complement system are known to be regulated by pro- and anti-inflammatory cytokines. The purpose of this work was to study the effect of IL-17 alone and combined with tumour necrosis factor (TNF) on the expression and synthesis of factor B and C3. Fibroblasts were stimulated with the relevant cytokine or cytokines, pulse labelled with 35S-methionine, and the newly synthesized proteins were immunoprecipitated and subjected to SDS-PAGE. Gene expression was determined by Northern blot analysis. IL-17 10 ng/ml induced increases in gene expression and protein synthesis of C3, 2.25 +/- 0.26- and 2.7 +/- 0.7-fold, respectively with concomitant non-significant effects on factor B, 1.5 +/- 0.45- and 2.2 +/- 1. 2-fold, respectively. When both IL-17 and TNF were present simultaneously, the synthesis of factor B increased by 85% more than the expected additive effects of these cytokines separately, while for C3 the effect of both cytokines was 19% lower than the expected additive effect (observed/expected = 0.81). IL-4 reduced the synergistic effect by 50%. We conclude that IL-17 has a regulatory role on C3 expression and synthesis and an amplifying effect on TNF-induced factor B synthesis. Taken together with the evidence that TNF is a major cytokine involved in the inflammation of rheumatoid arthritis, it suggests that IL-17 has a proinflammatory role in the inflammation process of joints. The distinct effects of IL-4, IL-17 and TNF on the synthesis of factor B in fibroblasts suggest that factor B and the alternative pathway of the complement system may play an important role in joint inflammation.