RESUMO
OBJECTIVE: We recently developed an ex vivo cellular model of pannus, the aberrant overgrowth of human synovial tissue. This study was undertaken to use that model to investigate the role of prostaglandin E2 (PGE2) and its receptor subtypes in the development of pannus growth and osteoclast activity in rheumatoid arthritis (RA). METHODS: Inflammatory cells that infiltrated pannus from patients with RA were collected without enzyme digestion and designated synovial tissue-derived inflammatory cells. Their single-cell suspensions were cultured in medium alone to observe an aberrant overgrowth of inflammatory tissue in vitro. Levels of cytokines produced in culture supernatants were measured using enzyme-linked immunosorbent assay kits. Osteoclast activity was assessed by the development of resorption pits in calcium phosphate-coated slides. RESULTS: Primary culture of the synovial tissue-derived inflammatory cells resulted in spontaneous reconstruction of inflammatory tissue in vitro within 4 weeks, during which tumor necrosis factor α, PGE2, macrophage colony-stimulating factor, and matrix metalloproteinase 9 were produced in the supernatant. This aberrant overgrowth was inhibited by antirheumatic drugs including methotrexate and infliximab. On calcium phosphate-coated slides, synovial tissue-derived inflammatory cells showed numerous resorption pits. In the presence of inhibitors of endogenous prostanoid production such as indomethacin and NS398, exogenous PGE1 and EP4-specific agonists significantly inhibited all these activities of synovial tissue-derived inflammatory cells in a dose-dependent manner. Addition of indomethacin, NS398, or EP4-specific antagonist resulted in the enhancement of these cells' activities. EP2-specific agonist had a partial effect, while EP1- and EP3-specific agonists had no significant effects. CONCLUSION: These results suggest that endogenous PGE2 produced in rheumatoid synovium negatively regulates aberrant synovial overgrowth and the development of osteoclast activity via EP4.
Assuntos
Artrite Reumatoide/metabolismo , Dinoprostona/metabolismo , Osteoclastos/metabolismo , Receptores de Prostaglandina E Subtipo EP4/metabolismo , Membrana Sinovial/metabolismo , Artrite Reumatoide/patologia , Células Cultivadas , Inibidores de Ciclo-Oxigenase/farmacologia , Dinoprostona/farmacologia , Relação Dose-Resposta a Droga , Humanos , Indometacina/farmacologia , Nitrobenzenos/farmacologia , Osteoclastos/efeitos dos fármacos , Osteoclastos/patologia , Sulfonamidas/farmacologia , Membrana Sinovial/efeitos dos fármacos , Membrana Sinovial/patologiaRESUMO
OBJECTIVE: Anti-cyclic citrullinated peptide (CCP) antibodies are a serological marker for rheumatoid arthritis (RA); up to 10%-15% of patients with systemic lupus erythematosus (SLE) are also positive. While anti-CCP in RA is citrulline-dependent, anti-CCP in some other diseases is citrulline-independent and reacts with both CCP and the unmodified (arginine-containing) cyclic arginine peptide (CAP). We investigated the citrulline dependence of anti-CCP and its significance in the arthritis of SLE. METHODS: IgG anti-CCP was compared by ELISA to anti-CAP in sera from patients with SLE (n = 335) and RA (n = 47) and healthy controls (n = 35). SLE patients were divided into 5 groups based on their joint involvement: subset I: deforming/erosive arthritis (n = 20); II: arthritis fulfilling (or likely fulfilling) American College of Rheumatology criteria for RA but without erosions (n = 18); III: joint swelling but not fulfilling RA criteria (n = 39); IV: arthritis without documented joint swelling (n = 194); and V: no arthritis (n = 58). RESULTS: Anti-CCP (> 1.7 units) was found in 68% (32/47) of patients with RA and 17% (55/329) of those with SLE. It was more common in SLE patients with deforming/erosive arthritis (38%). High anti-CCP (> 10 units) was found in RA (26%) and deforming/erosive SLE (12%). High anti-CCP/CAP ratios (> 2, indicating a selectivity to CCP) were found in 91% of anti-CCP-positive RA and 50% of anti-CCP-positive SLE patients with deforming/erosive arthritis. Patients from subset II did not have high anti-CCP/CAP. CONCLUSION: Citrulline dependence or high levels (> 10) of anti-CCP were common in SLE patients with deforming/erosive arthritis, while most anti-CCP in SLE patients was citrulline-independent. This may be useful in identifying a subset of SLE patients with high risk for development of deforming/erosive arthritis.
Assuntos
Artrite Reumatoide , Autoanticorpos , Citrulina/metabolismo , Lúpus Eritematoso Sistêmico/imunologia , Lúpus Eritematoso Sistêmico/patologia , Peptídeos Cíclicos/imunologia , Artrite Reumatoide/sangue , Artrite Reumatoide/imunologia , Artrite Reumatoide/patologia , Povo Asiático , Autoanticorpos/sangue , Autoanticorpos/imunologia , Biomarcadores/sangue , Ensaio de Imunoadsorção Enzimática , Humanos , Articulações/patologia , Lúpus Eritematoso Sistêmico/sangue , Lúpus Eritematoso Sistêmico/classificaçãoRESUMO
Association of SLE and alfalfa was first reported in a volunteer who developed lupus-like autoimmunity while ingesting alfalfa seed for a hypercholesterolemia study. This was corroborated with studies in monkeys fed with alfalfa sprout that developed SLE. Re-challenge with L-canavanine relapsed the disease. Arginine homologue L-canavanine, present in alfalfa, was suspected as a cause. L-canavanine can be charged by arginyl tRNA synthetase to replace L-arginine during protein synthesis. Aberrant canavanyl proteins have disrupted structure and functions. Induction or exacerbation of SLE by alfalfa tablets reported in a few cases remains controversial. Epidemiological studies on the relationship between alfalfa and SLE are sparse. In mice, NZB/W F1, NZB, and DBA/2 mice fed with L-canavanine show exacerbation/triggering of the SLE, however, BALB/c studies were negative. L-canavanine incorporation may be more efficient in the presence of inflammation or other conditions that can cause arginine deficiency. The L-canavanine induced apoptotic cells can be phagocytosed and a source of autoantigens processed by endosomal proteases. Endogenous canavanyl proteins are ubiquitinated and processed via proteasome. Incorporation of L-canavanine into proteasome or endosome can also cause disruption of antigen processing. Alfalfa/L-canavanine-induced lupus will be an interesting model of autoimmunity induced by the modification of self-proteins at the translational level.
Assuntos
Autoimunidade , Canavanina/intoxicação , Lúpus Eritematoso Sistêmico/induzido quimicamente , Medicago sativa/efeitos adversos , Aminoácidos , Animais , Arginina , Autoanticorpos/análise , Feminino , Haplorrinos , Humanos , Lúpus Eritematoso Sistêmico/sangue , Lúpus Eritematoso Sistêmico/imunologia , Camundongos , Camundongos Endogâmicos BALB CRESUMO
OBJECTIVE: Autoantibodies to aminoacyl transfer RNA synthetases, such as histidyl (Jo-1), threonyl (PL-7), alanyl (PL-12), glycyl (EJ), and isoleucyl (OJ), are closely associated with a subset of patients with polymyositis/dermatomyositis (PM/DM) complicated by interstitial lung disease (ILD). Anti-Jo-1 is by far the most common, found in 15-25% of patients with PM/DM, whereas the other types are found in only approximately 3% of these patients. In this study, the clinical associations of these autoantibodies in Japanese patients with PM/DM were investigated. METHODS: The diagnoses of PM/DM and amyopathic DM (ADM) were based on the Bohan and Peter criteria and Sontheimer's definition, respectively. Sera from 36 Japanese patients with PM/DM (13 with PM, 20 with DM, 3 with ADM) were screened by immunoprecipitation and by enzyme-linked immunosorbent assay (for Jo-1). Clinical and laboratory data were collected. RESULTS: The frequencies of autoantibodies to Jo-1 (22%) and to EJ, OJ, and PL-12 (3-6%) were similar to those found in previous studies, including studies of Japanese subjects. However, anti-PL-7 was found in 17% of patients, in contrast to a frequency of 1-4% in previous studies (P < 0.02-0.0002). The 6 anti-PL-7-positive patients were not related, and no skewing in year or month of disease development, place of residence or work, or occupation was found. All patients had ILD, consistent with the clinical features of antisynthetase-positive patients. The patients with anti-PL-7 had lower serum muscle enzyme levels and milder muscle weakness (P < 0.05) compared with anti-Jo-1-positive patients. CONCLUSION: Anti-PL-7 was found at an unusually high frequency in this group of Japanese patients with myositis. Although anti-PL-7, similar to anti-Jo-1, is associated with PM/DM with ILD, muscle involvement in the patients with anti-PL-7 appeared to be milder than that in the anti-Jo-1 subset.
Assuntos
Autoanticorpos/sangue , Dermatomiosite/imunologia , Treonina-tRNA Ligase/imunologia , Adulto , Alanina-tRNA Ligase/imunologia , Povo Asiático , Ensaio de Imunoadsorção Enzimática , Feminino , Glicina-tRNA Ligase/imunologia , Histidina-tRNA Ligase/imunologia , Humanos , Imunoprecipitação , Isoleucina-tRNA Ligase/imunologia , Japão , Doenças Pulmonares Intersticiais/complicações , Masculino , Pessoa de Meia-Idade , Doenças Musculares/imunologia , Polimiosite/imunologiaRESUMO
OBJECTIVE: Most lupus patients produce autoantibodies against small ribonucleoproteins such as Sm/RNP and Ro 60 (containing U1 and Y1-Y5 RNAs, respectively). We undertook this study to investigate whether the RNA components of these antigens, which contain extensive tracts of single- and double-stranded RNA, signatures of viral infection, activate innate immunity. METHODS: U1 and Y RNAs were affinity purified from K562 cells. Murine bone marrow-derived dendritic cells (DCs), human HEK 293 cells, and murine RAW264.7 cells were stimulated with U1 RNA and other known Toll-like receptor (TLR) ligands. Expression of the interferon (IFN)-inducible gene Mx1 and other genes was quantified using real-time polymerase chain reaction, and cytokine production was measured by enzyme-linked immunosorbent assay. DC maturation was assessed using flow cytometry. RESULTS: Purified U1 and Y1-Y5 RNAs and synthetic stem-loop II of U1 RNA stimulated type I IFN (IFN-I) production by cell lines and murine bone marrow-derived DCs and promoted DC maturation (CD86 expression). U1 RNA-stimulated, but not TLR-3 ligand-stimulated, IFN-I was blocked by bafilomycin A1, indicating that immunostimulation by U1 RNA requires endosomal acidification. Myeloid differentiation factor 88-deficient cells responded poorly to U1 RNA, suggesting that an endosomal TLR, probably TLR-7, mediates the stimulatory effects of U1 RNA. U1 RNA-induced IFN-I and interleukin-6 production also were protein kinase R (PKR) dependent (abrogated by 2-aminopurine and greatly reduced in PKR-/- cells). CONCLUSION: We conclude that the RNA components of the Ro 60 (Y1-Y5 RNA) and Sm/RNP (U1 RNA) small ribonucleoproteins act as endogenous adjuvants that could play a role in the pathogenesis of autoimmunity by stimulating DC maturation and IFN-I production.
Assuntos
Autoantígenos/imunologia , Lúpus Eritematoso Sistêmico/imunologia , RNA Citoplasmático Pequeno/imunologia , Ribonucleoproteínas Nucleares Pequenas/imunologia , Ribonucleoproteínas/imunologia , Animais , Células Cultivadas , Endossomos/imunologia , Humanos , Interferon Tipo I/biossíntese , Camundongos , Receptores Toll-Like/fisiologiaRESUMO
Lymphoid neogenesis is associated with antibody-mediated autoimmune diseases such as Sjogren's syndrome and rheumatoid arthritis. Although systemic lupus erythematosus is the prototypical B-cell-mediated autoimmune disease, the role of lymphoid neogenesis in its pathogenesis is unknown. Intraperitoneal injection of 2,6,10,14-tetramethyl-pentadecane (TMPD, pristane) or mineral oil causes lipogranuloma formation in mice, but only TMPD-treated mice develop lupus. We report that lipogranulomas are a form of lymphoid neogenesis. Immunoperoxidase staining of lipogranulomas revealed B cells, CD4(+) T cells, and dendritic cells and in some cases organization into T- and B-cell zones. Lipogranulomas also expressed the lymphoid chemokines CCL21, CCL19, CXCL13, CXCL12, and CCL22. Expression of the type I interferon (IFN-I)-inducible genes Mx1, IRF7, IP-10, and ISG-15 was greatly increased in TMPD- versus mineral oil-induced lipogranulomas. Dendritic cells from TMPD lipogranulomas underwent activation/maturation with high CD86 and interleukin-12 expression. Magnetic bead depletion of dendritic cells markedly diminished IFN-inducible gene (Mx1) expression. We conclude that TMPD-induced lupus is associated with the formation of ectopic lymphoid tissue containing activated dendritic cells producing IFN-I and interleukin-12. In view of the increased IFN-I production in systemic lupus erythematosus, these studies suggest that IFN-I from ectopic lymphoid tissue could play a role in the pathogenesis of experimental lupus in mice.
Assuntos
Granuloma/metabolismo , Interferon Tipo I/biossíntese , Tecido Linfoide/metabolismo , Terpenos , Animais , Linhagem Celular , Quimiocina CXCL10 , Quimiocinas/biossíntese , Quimiocinas CXC/biossíntese , Coristoma/induzido quimicamente , Coristoma/metabolismo , Coristoma/patologia , Citocinas/biossíntese , Células Dendríticas/metabolismo , Feminino , Proteínas de Ligação ao GTP/biossíntese , Granuloma/induzido quimicamente , Granuloma/patologia , Fator Regulador 7 de Interferon/biossíntese , Interleucina-12/biossíntese , Lúpus Eritematoso Sistêmico/induzido quimicamente , Lúpus Eritematoso Sistêmico/patologia , Tecido Linfoide/efeitos dos fármacos , Tecido Linfoide/patologia , Camundongos , Camundongos Endogâmicos BALB C , Óleo Mineral , Proteínas de Resistência a Myxovirus , Peritônio , Ubiquitinas/biossínteseRESUMO
Recent progress in understanding the pathogenesis of rheumatoid arthritis (RA) in parallel with elucidation of the functional role of the prostaglandin receptor subfamily has revealed an important regulatory role of PGE2, in addition to its well-known proinflammatory role in the progression of RA. Characteristic features of RA are synovial proliferation and pannus formation, which result in the destruction of cartilage and bone. Pannus tissue is mainly composed of macrophages and fibroblast-like synoviocytes. Both T cell-derived IL-17 and macrophage-derived TNF-alpha seem to play a central role in the progression of proinflammatory cascades in RA. PGE2 is also produced in response to proinflammatory cytokines, which in turn negatively regulates both IL-17 and TNF-alpha expression and TNF/IL-1-induced activation of fibroblast-like synoviocytes through EP2/EP4 receptors, resulting in the modulation of proinflammatory cascades. IL-17- and TNF-activated macrophages differentiate into osteoclasts in the presence of M-CSF and RANKL expressed by fibroblast-like synoviocytes. PGE2 binding to EP4 stimulates osteoclastogenesis through enhancing RANKL expression. At the same time, PGE2 suppresses osteoclastogenesis by inhibiting M-CSF expression of fibroblast-like synoviocytes as well as both IL-17 and IL-17-induced TNF-alpha expression of macrophages. PGE2-EP4 also activates osteoblastogenesis through increasing cbfa1 and osterix, two essential transcription factors required for bone formation. The net effect of PGE2 may direct toward repair of eroding bone through the suppression of inflammation and enhancement of bone remodeling. Here, we discuss a diverse action of PGE2/EP receptors and their important regulatory roles in the pathogenesis of RA, which may lead to a novel therapeutic strategy.
Assuntos
Antirreumáticos/farmacologia , Antirreumáticos/uso terapêutico , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/patologia , Dinoprostona/fisiologia , Receptores de Prostaglandina/efeitos dos fármacos , Receptores de Prostaglandina/fisiologia , Animais , Artrite Experimental/tratamento farmacológico , Artrite Experimental/patologia , Remodelação Óssea/efeitos dos fármacos , Humanos , Mediadores da InflamaçãoRESUMO
A single intraperitoneal (i.p.) injection of pristane, incomplete Freund's adjuvant (IFA), or the adjuvant oil squalene, but not high molecular weight medicinal mineral oils, induces lupus-related autoantibodies to nRNP/Sm and -Su in non-autoimmune strains of mice. This ability appears to be associated with the low molecular weight and adjuvanticity of hydrocarbon. n-Hexadecane (C(16)H(34)), which is present in petroleum, has adjuvant activity and induces arthritis in rodents like other lupus-inducing oils. In addition to dietary exposure to n-hexadecane in mineral oils, exposure also occurs via inhalation of oil mist, jet fuel, or diesel exhaust or by absorption through the skin. Since n-hexadecane is a low molecular weight adjuvant hydrocarbon oil similar to other lupus-inducing hydrocarbons, the present study examined whether it can also induce lupus-related autoantibodies in mice. Female BALB/cJ mice received a single i.p. injection of 0.5 ml of n-hexadecane, pristane, or saline (control). Pathology and serology (immunoglobulin levels, autoantibodies by immunofluorescence, immunoprecipitation, and ELISA) were examined 3 months later. Unexpectedly, all n-hexadecane-treated mice, but none in the other groups, developed inflammatory ascites within 2.5 months. n-Hexadecane induced hypergammaglobulinemia (IgG1, IgG2a), antinuclear (titer>1:160, 67%) and -cytoplasmic antibodies (58%) and autoantibodies to nRNP/Sm (25%), Su (33%), ssDNA (83%), and chromatin (100%). Therefore, non-specific inflammation caused by n-hexadecane resulted in the production of a limited set of specific autoantibodies. These previously unrecognized immunological effects of n-hexadecane may have implications in monitoring human exposure to hydrocarbons and in the pathogenesis of autoimmune diseases.
Assuntos
Alcanos/toxicidade , Autoanticorpos/imunologia , Lúpus Vulgar/induzido quimicamente , Animais , Ascite/induzido quimicamente , Autoimunidade , Poluentes Ambientais/toxicidade , Feminino , Injeções Intraperitoneais , Fígado/efeitos dos fármacos , Fígado/patologia , Lúpus Vulgar/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Peritonite/induzido quimicamente , Baço/efeitos dos fármacos , Baço/patologiaRESUMO
Hypergammaglobulinemia and autoantibodies are reduced in pristane-treated specific pathogen-free mice vs. conventionally housed controls, consistent with the role of microbial stimulation in this model. To determine whether microbial stimulation is required, BALB/c mice housed under germ-free conditions were treated i.p. with sterile PBS or pristane and examined 6 months later. As in conventional mice, pristane-treated germ-free mice developed peritoneal granulomas and hypergammaglobulinemia with increased IgG2a/IgG1 ratios. LPS stimulation induced more IL-6, IL-12, and TNF-alpha, and anti-CD3 induced more IFN-gamma and IL-4 by peritoneal cells from pristane-treated mice vs. control. Anti-nRNP/Sm and -Su autoantibodies were found in 40% and 43%, respectively, of pristane-treated germ-free mice by immunoprecipitation. Thus, bacterial stimulation was not required for lupus autoantibodies, peritoneal granuloma formation, hypergammaglobulinemia, or cytokine overproduction. Although microbial stimulation acts synergistically with pristane, these results clearly indicate that pristane does not act merely by increasing exposure to microbial products such as LPS.
Assuntos
Autoanticorpos/imunologia , Lúpus Eritematoso Sistêmico/induzido quimicamente , Terpenos/farmacologia , Animais , Peso Corporal , Citocinas/sangue , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Feminino , Granuloma/imunologia , Histocitoquímica , Hipergamaglobulinemia/imunologia , Fígado/imunologia , Fígado/patologia , Lúpus Eritematoso Sistêmico/imunologia , Lúpus Eritematoso Sistêmico/microbiologia , Lúpus Eritematoso Sistêmico/patologia , Camundongos , Camundongos Endogâmicos BALB C , Tamanho do Órgão , Organismos Livres de Patógenos Específicos , Baço/imunologia , Baço/patologia , Estatísticas não Paramétricas , Terpenos/imunologiaRESUMO
OBJECTIVE: Autoantibodies to U1-C have been considered a minor component of anti-snRNP (nRNP, Sm) response based on Western blotting. However, we have previously shown that virtually all human anti-nRNP sera contain antibodies to native U1-C, as well as novel autoantibodies that stabilize the molecular interaction of the U1-C-Sm core particle. The biological significance of stabilizing antibodies was investigated by titering anti-U1-C/U1-A compared to stabilizing antibodies, and by examining the effects of stabilizing antibodies on antigen processing. METHODS: Autoantibodies to individual native components of U1snRNP (immunoprecipitation of RNase-treated cell extract) and stabilizing antibodies (dissociation of snRNP on anti-Sm monoclonal antibodies by 1 M MgCl2) in human autoimmune sera were titered. Effects of stabilizing antibodies on proteolysis were assessed by incubating UsnRNP with anti-snRNP/Sm autoimmune sera prior to protease digestion. RESULTS: Autoantibodies to native U1-C and U1-C-Sm core particle stabilizing antibodies were universally present in human anti-nRNP or anti-nRNP + anti-Sm sera, but not in anti-Sm sera. Antibodies to U1-A were less common. The titers of stabilizing antibodies were higher than those of antibodies to U1-C (p < 0.01), indicating that the stabilizing antibodies were a significant component of the anti-snRNP response. The stabilizing anti-nRNP, but not anti-Sm antibodies, protects the Sm core particle from dissociation and proteolysis. CONCLUSION: Autoantibodies stabilizing the U1-C-Sm core particle were universally present in anti-nRNP sera and delay proteolysis of the Sm core particle. They may suppress spreading of the autoimmune response to Sm by delaying or altering processing of the Sm core proteins by antigen-presenting cells.
Assuntos
Autoanticorpos/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Peptídeo Hidrolases/metabolismo , Ribonucleoproteína Nuclear Pequena U1/imunologia , Ribonucleoproteínas Nucleares Pequenas/imunologia , Especificidade de Anticorpos , Autoanticorpos/análise , Autoantígenos , Biomarcadores/sangue , Feminino , Humanos , Imunoprecipitação , Técnicas In Vitro , Lúpus Eritematoso Sistêmico/sangue , Masculino , Ribonucleoproteína Nuclear Pequena U1/análise , Ribonucleoproteínas Nucleares Pequenas/análise , Estudos de Amostragem , Sensibilidade e Especificidade , Proteínas Centrais de snRNPRESUMO
Adjuvant oils such as Bayol F (Incomplete Freund's adjuvant: IFA) and squalene (MF59) have been used in human and veterinary vaccines despite poor understanding of their mechanisms of action. Several reports suggest an association of vaccination and various autoimmune diseases, however, few were confirmed epidemiologically and the risk of vaccination for autoimmune diseases has been considered minimal. Microbial components, not the adjuvant components, are considered to be of primary importance for adverse effects of vaccines. We have reported that a single intraperitoneal injection of the adjuvant oils pristane, IFA or squalene induces lupus-related autoantibodies to nRNP/Sm and -Su in non-autoimmune BALB/c mice. Induction of these autoantibodies appeared to be associated with the hydrocarbon's ability to induce IL-12, IL-6, and TNF-alpha, suggesting a relationship with hydrocarbon's adjuvanticity. Whether this is relevant in human vaccination is a difficult issue due to the complex effects of vaccines and the fact that immunotoxicological effects vary depending on species, route, dose, and duration of administration. Nevertheless, the potential of adjuvant hydrocarbon oils to induce autoimmunity has implications in the use of oil adjuvants in human and veterinary vaccines as well as basic research.
Assuntos
Adjuvantes Imunológicos/efeitos adversos , Adjuvante de Freund/efeitos adversos , Lúpus Eritematoso Sistêmico/induzido quimicamente , Óleo Mineral/efeitos adversos , Esqualeno/efeitos adversos , Animais , Autoanticorpos/biossíntese , Autoimunidade , Modelos Animais de Doenças , Humanos , Lúpus Eritematoso Sistêmico/imunologia , Camundongos , Terpenos , Vacinas/efeitos adversosRESUMO
Prostaglandin E(2) (PGE(2)) can have pro- or anti-inflammatory effects, depending on engagement of different PGE(2) receptor (EP) subtypes. The role of EPs in regulating autoimmune inflammation was studied in the murine arthritis/lupus model induced by pristane. Peritoneal macrophages were isolated (biomagnetic beads) from BALB/c, DBA/1, or C57BL/6 mice treated with pristane (intraperitoneally, 3 months earlier) or thioglycolate (3 days earlier) or with untreated controls. EPs, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2) mRNA expression was examined by reverse transcriptase-polymerase chain reaction (RT-PCR). Cells were cultured unstimulated or stimulated with lipopolysaccharide (LPS) or LPS + interferon-gamma in combination with EP subtype-specific agonists. Tumor necrosis factor alpha (TNF-alpha) and interleukin (IL)-6 production was tested by enzyme-linked immunosorbent assay (culture supernatant) and flow cytometry. TNF-alpha mRNA levels also were examined. High levels of EPs (EP4/2>EP1>EP3), iNOS, and COX-2 mRNA were expressed in peritoneal macrophages from pristane-treated but not untreated or thioglycolate-treated mice (RT-PCR). TNF-alpha production was inhibited 50-70% at 2-24 h by EP4/2 agonists, whereas IL-6 was enhanced up to approximately 220%. TNF-alpha inhibition is mediated partly via the protein kinase A pathway and partly via IL-6. Intracellular TNF-alpha staining was inhibited 20% by EP4/2 agonists. TNF-alpha mRNA levels were inhibited 50-70% at 2-24 h, indicating that TNF-alpha inhibition was partly at the level of transcription. EP1/3 agonists had little effect. Synovial cells from mice with pristane-induced arthritis (DBA/1) also expressed EP2/4, and the EP2/4 agonist inhibited TNF-alpha production. PGE(2) can modulate inflammatory reactions via the EP2/4 receptor through its regulation of TNF-alpha and IL-6. Modification of EP signaling may be a new therapeutic strategy in inflammatory/autoimmune diseases.
Assuntos
Interleucina-6/biossíntese , Articulações/imunologia , Macrófagos Peritoneais/imunologia , Receptores de Prostaglandina E/metabolismo , Fator de Necrose Tumoral alfa/efeitos dos fármacos , Animais , Artrite/induzido quimicamente , Células Cultivadas , Ciclo-Oxigenase 2 , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Imunossupressores/farmacologia , Isoenzimas/biossíntese , Isoenzimas/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos , Óxido Nítrico Sintase/biossíntese , Óxido Nítrico Sintase/efeitos dos fármacos , Óxido Nítrico Sintase Tipo II , Prostaglandina-Endoperóxido Sintases/biossíntese , Prostaglandina-Endoperóxido Sintases/efeitos dos fármacos , Receptores de Prostaglandina E/efeitos dos fármacos , Receptores de Prostaglandina E Subtipo EP4 , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Membrana Sinovial/citologia , Membrana Sinovial/efeitos dos fármacos , Membrana Sinovial/imunologia , Terpenos/farmacologia , Tioglicolatos/farmacologia , Fator de Necrose Tumoral alfa/biossíntese , Regulação para CimaRESUMO
Although mineral oils are generally considered nontoxic and have a long history of use in humans, the mineral oil Bayol F (incomplete Freund's adjuvant, IFA) and certain mineral oil components (squalene and n-hexadecane) induce lupus-related anti-nRNP/Sm or -Su autoantibodies in nonautoimmune mice. In the present study, we investigated whether medicinal mineral oils can induce other types of autoantibodies and whether structural features of hydrocarbons influence autoantibody specificity. Female 3-month-old BALB/c (16-45/group) mice each received an i.p. injection of pristane (C19), squalene (C30), IFA, three medicinal mineral oils (MO-F, MO-HT, MO-S), or PBS. Sera were tested for autoantibodies and immunoglobulin levels. Hydrocarbons were analyzed by gas chromatography/mass spectrometry. IFA contained mainly C15-C25 hydrocarbons, whereas MO-HT and MO-S contained C20-C40, and MO-F contained C15-C40. Pristane and n-hexadecane were found in IFA (0.17% and 0.10% w/v, respectively) and MOs (0.0026-0.027%). At 3 months, pristane and IFA induced mainly IgG2a, squalene IgG1, and MOs IgG3 and IgM in sera. Anti-cytoplasmic antibodies were common in mice treated with MO-F, as well as those treated with pristane, squalene, and IFA. Anti-ssDNA and -chromatin antibodies were higher in MO-F and MO-S than in untreated/PBS, squalene-, or IFA-treated mice, suggesting that there is variability in the induction of anti-nRNP/Sm versus -chromatin/DNA antibodies. The preferential induction of anti-chromatin/ssDNA antibodies without anti-nRNP/Sm/Su by MO-S and MO-F is consistent with the idea that different types of autoantibodies are regulated differently. Induction of autoantibodies by mineral oils considered nontoxic also may have pathogenetic implications in human autoimmune diseases.
Assuntos
Autoanticorpos/biossíntese , Hipergamaglobulinemia/induzido quimicamente , Óleo Mineral/toxicidade , Alcanos/análise , Animais , Anticorpos Antinucleares/biossíntese , Anticorpos Antinucleares/sangue , Autoanticorpos/sangue , Autoimunidade , Cromatina/imunologia , DNA de Cadeia Simples/imunologia , Feminino , Adjuvante de Freund/química , Adjuvante de Freund/toxicidade , Cromatografia Gasosa-Espectrometria de Massas , Humanos , Hipergamaglobulinemia/sangue , Imunoglobulinas/biossíntese , Imunoglobulinas/sangue , Camundongos , Camundongos Endogâmicos BALB C , Óleo Mineral/química , Esqualeno/análise , Esqualeno/toxicidade , Terpenos/análise , Terpenos/toxicidadeRESUMO
The case of a 27-year-old woman who simultaneously presented with SLE and severe refractory thrombotic microangiopathic hemolytic anemia (TMHA) is reported. She had extremely high levels of platelet-associated IgG (PAIgG), and her TMHA was refractory to plasma exchange and corticosteroid therapy. However, the TMHA was effectively controlled by i.v. cyclophosphamide therapy. ITP and TTP are generally considered distinct diseases; however, TMHA may occur secondary to platelet aggregation via autoimmune mechanisms in certain cases. Immunosuppressive therapy at an early stage of the disease may be beneficial in refractory cases of TMHA with autoimmune features.
Assuntos
Anemia Hemolítica/tratamento farmacológico , Antirreumáticos/administração & dosagem , Ciclofosfamida/administração & dosagem , Lúpus Eritematoso Sistêmico/complicações , Adulto , Anemia Hemolítica/complicações , Anemia Hemolítica/imunologia , Feminino , Humanos , Infusões Intravenosas , Lúpus Eritematoso Sistêmico/imunologia , Trombose/etiologia , Resultado do TratamentoAssuntos
Vírus da Leucemia Murina , Leucemia Experimental/complicações , Nefrite Lúpica/virologia , Neoplasias Mamárias Experimentais/complicações , Vírus do Tumor Mamário do Camundongo , Infecções por Retroviridae/complicações , Infecções Tumorais por Vírus/complicações , Animais , Autoanticorpos/sangue , Feminino , Imunossupressores , Leucemia Experimental/imunologia , Nefrite Lúpica/induzido quimicamente , Nefrite Lúpica/imunologia , Neoplasias Mamárias Experimentais/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Infecções por Retroviridae/imunologia , Terpenos , Infecções Tumorais por Vírus/imunologiaRESUMO
Exposure to the hydrocarbon oil pristane induces lupus specific autoantibodies in non-autoimmune mice. We investigated whether the capacity to induce lupus-like autoimmunity is a unique property of pristane or is shared by other adjuvant oils. Seven groups of 3-month-old female BALB/cJ mice received a single intraperitoneal injection of pristane, squalene (used in the adjuvant MF59), incomplete Freund's adjuvant (IFA), three different medicinal mineral oils, or saline, respectively. Serum autoantibodies and peritoneal cytokine production were measured. In addition to pristane, the mineral oil Bayol F (IFA) and the endogenous hydrocarbon squalene both induced anti-nRNP/Sm and -Su autoantibodies (20% and 25% of mice, respectively). All of these hydrocarbons had prolonged effects on cytokine production by peritoneal APCs. However, high levels of IL-6, IL-12, and TNFalpha production 2-3 months after intraperitoneal injection appeared to be associated with the ability to induce lupus autoantibodies. The ability to induce lupus autoantibodies is shared by several hydrocarbons and is not unique to pristane. It correlates with stimulation of the production of IL-12 and other cytokines, suggesting a relationship with a hydrocarbon's adjuvanticity. The potential to induce autoimmunity may complicate the use of oil adjuvants in human and veterinary vaccines.
Assuntos
Adjuvantes Imunológicos/efeitos adversos , Autoanticorpos/efeitos dos fármacos , Autoanticorpos/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Adjuvantes Imunológicos/farmacologia , Animais , Autoantígenos , Citocinas/metabolismo , Feminino , Adjuvante de Freund/efeitos adversos , Imunoglobulina G/sangue , Imunoglobulina G/efeitos dos fármacos , Interleucina-12/metabolismo , Lipídeos/efeitos adversos , Lúpus Eritematoso Sistêmico/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Peritônio/efeitos dos fármacos , Peritônio/metabolismo , Proteínas/imunologia , Ribonucleoproteínas Nucleares Pequenas/imunologia , Esqualeno/efeitos adversos , Terpenos/farmacologia , Proteínas Centrais de snRNPRESUMO
Murine lupus can occur spontaneously or be induced by hydrocarbons, such as pristane. Spontaneous disease in MRL and NZB/W F1 mice is suppressed by the xid (X-linked immunodeficiency) mutation, which greatly diminishes T cell-independent type 2 responses as well as the number of peritoneal B1 cells. The present study asked whether lupus induced by i.p. injection of pristane likewise is inhibited by the xid defect. Male CBA/N (xid) mice were refractory to the induction of autoantibodies by pristane, whereas 23% of pristane-treated male CBA/CaJ controls produced anti-nRNP/Sm, -Su and/or -OJ (isoleucyl tRNA synthetase) antibodies. Unexpectedly, 43% (12 of 28) of the xid mice spontaneously produced anti-nuclear antibodies that proved highly specific for the lupus antigen RNA helicase A (RHA). Strikingly, this specificity was absent in CBA/CaJ mice (none of 51). Moreover, pristane treatment suppressed the production of anti-RHA antibodies when administered prior to the onset of autoantibody production, but enhanced anti-RHA levels when given after the onset of autoantibody production, suggesting that pristane interferes with anti-RHA production at an early stage. Large amounts of IgG1 anti-RHA autoantibodies were detected in the sera of xid mice, whereas pristane-induced anti-nRNP/Sm and -Su autoantibodies were almost exclusively IgG2a. Cytokine production within the peritoneal cavity reflected the predominant isotypes: IL-12 and IFN-gamma predominated in pristane-treated mice, whereas IL-4 and IL-6 were more predominant in untreated xid mice. The spontaneous production of anti-RHA by xid mice and its suppression by pristane treatment at the level of autoantibody induction supports the idea that lupus autoantibodies may be generated through a variety of mechanisms.