Conjugated linoleic acid (CLA) has been shown to reduce inflammation via Peroxisome Proliferator-Activated Receptor (PPAR)-γ in inflammatory disorders such as Crohn's Disease and Inflammatory Bowel Disease. We sought to determine whether CLA isomers would reduce inflammation via PPAR-γ in cultured mesangial cells, and in murine models of anti-glomerular basement membrane (anti-GBM) glomerulonephritis and Systemic Lupus Erythematosus (SLE). SV40-transformed mouse mesangial cells (MES13) were cultured with pure CLA isomers (c9,t11 or t10,c12-CLA or a 50:50 mixture prior to immune stimulation with lipopolysaccharide and interferon-γ. Next, cultured mesangial cells were transfected with small interfering RNA (siRNA) targeting PPAR-γ and treated with CLA isomers prior to immune stimulation. ELISA, qPCR, Western blot, and Griess reaction were performed to measure cytokine production, mRNA expression, induced nitric oxide synthase (iNOS) and nitrite production, respectively. Next, myeloid-specific (LysM creR2+) PPAR-γ knockout mice were treated with CLA prior to the induction of anti-GBM glomerulonephritis and evaluated for disease. Finally, NZM2410/J mice (a natural model of SLE) were treated with c9,t11-CLA and evaluated for disease progression. Treatment with CLA reduced IL-6 production in cultured mesangial cells, but not in siRNA-treated mesangial cells, supporting a PPAR-γ-mediated mechanism. CLA treatment increased both Transforming Growth Factor (TGF-β) and Interleukin-1 Receptor Antagonist (IL-1RA) mRNA expression independent of PPAR--γ. While CLA treatment reduced nitrite production and iNOS production to some degree, this was an inconsistent finding. Conversely, in the induced anti-GBM mouse model, CLA treatment increased mesangial cell IL-6 mRNA expression, reduced TGF-β expression, and had no effect on IL-1RA. Moreover, NZM2410/J mice that were fed a c9,t11-CLA-supplemented diet had reduced survival times, increased renal inflammation and increased serum IgG2a relative to controls. Taken together, these studies indicate that the in vitro MES13 cell line does not translate to the in vivo mouse model of anti-GBM induced glomerulonephritis. Furthermore, while CLA may have beneficial effects in other mouse models, it worsens disease in NZM2410/J mice. Findings from these models should be interpreted with caution. / Ph. D.

Systemic lupus erythematous (SLE) is an autoimmune and inflammatory disease that can affect all organs of the body. The purpose of this quantitative cross-sectional study was to examine SLE-related issues associated with depression and work-productivity impairment, and to assess if depression mediated the relationship between SLE disease activity and damage and work-productivity impairment. Participants were 257 residents of the state of Georgia in the United States with SLE and were recruited from the Georgians Organized Against Lupus study. Bandura'€™s social cognitive theory was the guiding theoretical framework of the study. Findings showed that the majority of participants worked full time (78.2%), identified as Black (72.8%), female (94.2%), above poverty level (77.4%), and had private health insurance (70.0%). Mean and median score results indicated that participants missed, on average, slightly less than half a day of work every 7 days, and had mild-to-moderate levels of work productivity impairment. Mean and median scores showed that participants reported mild-to-moderate levels of SLE disease activity and damage and depression. Linear regression results revealed significant relationships between SLE activity and damage and work productivity impairment. Hierarchical linear regression for mediation findings indicated that depression partially mediated the relationship between SLE disease activity and damage and work productivity impairment. The findings from this study might help to increase stakeholder awareness of SLE disease activity and damage and SLE effects on depression and work functioning.

ABSTRACT OBJECTIVE: To determine the prevalence and clinical correlates of anti-phospholipid antibodies (aPL), including anti-cardiolipin antibodies (aCL), lupus anti-coagulant (LA), anti- β2-glycoprotein 1 (aβ2GP1) and anti-prothrombin (aPT) antibodies, in Black South African patients with systemic lupus erythematosus (SLE) METHODS: A cross-sectional study of 100 SLE patients in whom clinical characteristics, including features of the anti-phospholipid syndrome (APS), disease activity, and damage were documented, and sera tested for aCL, aβ2GP, and aPT of all isotypes, and LA. RESULTS: Positive aCL, aβ2GP1 and aPT and LA were found in 53, 84, 20, and 2 patients, respectively. Immunoglobulin (Ig)A aCL and IgG aβ2GP1 were the commonest aCL (49.1%) and aβ2GP1 (47%) isotypes, respectively. IgA aβ2GP1 were associated with both a history of thrombosis alone (p<0.05) and a history of any clinical feature, thrombosis and/or spontaneous abortion of the APS (p<0.05); IgA aCL were associated with a history of any clinical APS event (p<0.05); and aβ2GPI of any isotype were associated with a history of arthritis (p<0.001). CONCLUSION: My findings provide further evidence that the screening for aβ2GP1 and IgA aCL isotype may improve the risk assessment for APS in SLE patients of African extraction. Further prospective studies are warranted to determine the clinical utility of these tests and to elucidate the genetic basis for increased IgA aPL response in SLE patients of African extraction.

No description available.

Polymorphisms in the interferon regulatory factor 5 (IRF5) gene have been consistently replicated and shown to confer risk for or protection from the development of systemic lupus erythematosus (SLE). IRF5 expression is significantly upregulated in SLE patients and upregulation associates with IRF5-SLE risk haplotypes. IRF5 alternative splicing has also been shown to be elevated in SLE patients. Given that human IRF5 exists as multiple alternatively spliced transcripts with distinct function(s), it is important to determine whether the IRF5 transcript profile expressed in healthy donor immune cells is different from that expressed in SLE patients. Moreover, it is not currently known whether an IRF5-SLE risk haplotype defines the profile of IRF5 transcripts expressed. Using standard molecular cloning techniques, we identified and isolated 14 new differentially spliced IRF5 transcript variants from purified monocytes of healthy donors and SLE patients to generate an IRF5 variant transcriptome. Next-generation sequencing was then used to perform in-depth and quantitative analysis of full-length IRF5 transcript expression in primary immune cells of SLE patients and healthy donors by next-generation sequencing. Evidence for additional alternatively spliced transcripts was obtained from de novo junction discovery. Data from these studies support the overall complexity of IRF5 alternative splicing in SLE. Results from next-generation sequencing correlated with cloning and gave similar abundance rankings in SLE patients thus supporting the use of this new technology for in-depth single gene transcript profiling. Results from this study provide the first proof that 1) SLE patients express an IRF5 transcript signature that is distinct from healthy donors, 2) an IRF5-SLE risk haplotype defines the top four most abundant IRF5 transcripts expressed in SLE patients, and 3) an IRF5 transcript signature enables clustering of SLE patients with the H2 risk haplotype.

For more than fifty years glucocorticoids (GCs) has been used to treat a wide range of inflammatory diseases, such as allergies, asthma, rheumatoid arthritis, and autoimmune diseases, due to its potentiality on the antiinflammatory and immunomodulatory effects. The anti-inflammation actions of glucocorticoid were mediated by an intracellular receptor, glucocorticoid receptor (GR), a member of the nuclear receptor family of ligand-dependent transcription factor. Upon activation by their ligand, GRs translocated to the nuclear and then bound to glucocorticoid responsive element (GRE) or negative glucocorticoid responsive elemen (nGRE). The administration of GCs depended on the acuity of disease and on the responses of patient clinically. Although some Systemic Lupus Erythematosus (SLE) patients given the maximal steroid doses, the response to the therapy remained poorly, and thus called ¡§glucocorticoid resistance¡¨. Despite the fact that the side effects and complications in SLE patients may result from the restrictions of physic; it has been documented that there were some relationships between the glucocorticoid resistance with the polymorphisms of GR, and the levels of glucocorticoid receptor beta. However, no significant differences in the GR polymorphisns (TthIII, ER22/23EK, N363S, BclI and I559N) between controls and SLE patients were found and there were no significant differences found on the levels of SUMO-2 antibody between patients with active and inactive SLE in this study. Neverthless, a significant association on the the allelic polymorphism of BclI was observed in patients with glucocorticoid resistance. Additionally, the expression of GR£] in patients with SLE was higher than that of controls and the TthIII CT genotype was associated with GR£\ expression.

No description available.

The pathogenesis of systemic lupus erythematosus (SLE) and cardiovascular disease (CVD) are tightly linked, and CVD is one of the leading causes of death in lupus patients. There are many risk factors that increase the risk of CVD in SLE patients, including endothelial dysfunction, lipid dysregulation, and abnormal regulation of innate and adaptive immunity. We have previously investigated the role of interferon regulatory factor 5 (Irf5), on atherosclerosis in lupus mouse models. Irf5 has a pro-inflammatory function by activating macrophage and cytokine recruitment and is thus being considered as a potential therapeutic target for the treatment of SLE. We hypothesized that Irf5 deficiency would ameliorate lupus disease as well as improve cardiovascular disease in the Irf5-deficient mouse model. However, while lupus disease did improve in the mouse model, the atherosclerotic plaques were found to be significantly increased in size. This poses a challenge to our current understanding of Irf5, as well as adds complexity to an already difficult clinical problem. Therefore, our aim of this study is to characterize the cells within the atherosclerotic lesions to examine their inflammatory potential. The focus of this study is the infiltration of macrophages into the mouse aortic root as determined by immunohistochemistry staining. In a time-course study using apoE.Irf5-/- mice, we found that macrophages started to accumulate into aortic leaflets as early as two weeks after starting a Western diet. Macrophage infiltration into the site of leaflet attachment seemed to possibly be a precursor to atherosclerotic lesion formation and appeared as early as 4 weeks after starting Western diet. No apparent differences were found between Irf5 sufficient and Irf5 deficient mice at either two or four weeks on Western diet. In a bone marrow chimera study, we examined the effects of Irf5 from bone marrow- and non-bone marrow-derived cells on the accumulation of macrophages on aortic leaflets and in the tunica intima in the gld.apoE-/- mouse model of lupus and atherosclerosis. Macrophage accumulation did not correlate with differences in Irf5 production. However, the finding of macrophage accumulation on aortic leaflets suggests a role of macrophages in Libman-Sacks endocarditis, an inflammatory disease of the mitral and aortic valves seen in patients with lupus. Together, our results do not support nor refute a role of Irf5 in macrophage infiltration into the aortic root. More samples are needed, as are more methods of identifying macrophages and quantifying them. However, it is still likely that macrophages play a role in the pathogenesis of atherosclerotic lesion formation in a lupus mouse model, and it is an area of study worth exploring. In a time-course study using apoE.Irf5-/- mice, we found that macrophages started to accumulate into aortic leaflets as early as two weeks after starting a Western diet. Macrophage infiltration into the site of leaflet attachment seemed to possibly be a precursor to atherosclerotic lesion formation and appeared as early as 4 weeks after starting Western diet. No apparent differences were found between Irf5 sufficient and Irf5 deficient mice at either two or four weeks on Western diet. In a bone marrow chimera study, we examined the effects of Irf5 from bone marrow- and non-bone marrow-derived cells on the accumulation of macrophages on aortic leaflets and in the tunica intima in the gld.apoE-/- mouse model of lupus and atherosclerosis. Macrophage accumulation did not correlate with differences in Irf5 production. However, the finding of macrophage accumulation on aortic leaflets suggests a role of macrophages in Libman-Sacks endocarditis, an inflammatory disease of the mitral and aortic valves seen in patients with lupus. Together, our results do not support a role of Irf5 in macrophage infiltration into the aortic root. However, it is still likely that macrophages play a role in the pathogenesis of atherosclerotic lesion formation in a lupus mouse model, and it is an area of study worth exploring.

Orientadores: Sandra Cecilia Botelho Costa, Lilian Tereza Lavras Costallat / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-13T20:41:17Z (GMT). No. of bitstreams: 1 Norberto_CristianeMudinutidaSilva_M.pdf: 1242945 bytes, checksum: 8ce75d400c67b263809938aa66fce948 (MD5) Previous issue date: 2008 / Resumo: Doenças Infecciosas são uma das maiores causas de complicações que ameaçam à vida dos pacientes com doenças reumáticas. O citomegalovírus, um betaherpesvírus, é o maior causador de morbidade e mortalidade em indivíduos imunossuprimidos, podendo causar febre e danos a órgãos como, pulmão, fígado, trato gastrointestinal, medula óssea e retina. Pacientes com LES fazem uso de imunossupressores e estão sob risco de reativar a infecção latente pelo HCMV. Os pacientes HCMV-IgG positivos que usam imunossupressores devem ser monitorizados usando a N-PCR e/ou HCMV antigenemia para que o tratamento antiviral precoce possa suprimir a reativação viral e conseqüentemente a doença associada. Diante do exposto, foram monitorizados 40 pacientes com diagnóstico de Lúpus Eritematoso Sistêmico (SLE) em relação à infecção ativa pelo citomegalovírus humano (HCMV) utilizando as técnicas de Nested-PCR (N-PCR) e Antigenemia (AGM). Foi também verificado o status sorológico IgM e IgG anti- HCMV por ELISA. Para tanto, foram necessárias 2 amostras de sangue por paciente, obtidas através de punção venosa. Os pacientes encontravam-se em diferentes estágios da doença lúpica ou tratamento específico. Os objetivos deste estudo foram: detectar a presença do DNA e do antígeno pp65 do HCMV utilizando as técnicas de N-PCR e AGM, respectivamente, em amostras de sangue periférico de pacientes com LES; verificar o impacto clínico causado pela infecção ativa e doença por HCMV nos pacientes desta casuística; verificar a presença das imunoglobulinas IgG e IgM anti-HCMV pelo método de ELISA. Todos os pacientes apresentaram sorologia IgG-HCMV reagente. Dos pacientes estudados, 4/40 (10%) apresentaram infecção ativa pelo HCMV detectado por NPCR e/ou AGM. Destes, 3 estavam com a doença lúpica em atividade (SLEDAI ? 6). Entre os pacientes com infecção ativa pelo HCMV, 2/4 (50%) apresentaram sintomas compatíveis com doença por HCMV. Um destes pacientes foi à óbito por sepse de foco pulmonar, compatível com doença por HCMV. Apesar da infecção ativa por HCMV não ser freqüente nos pacientes com LES, quando ocorre é de extrema gravidade, podendo levar o paciente à óbito. A utilização de diagnóstico precoce da infecção ativa pelo HCMV é muito importante, pois permite um tratamento mais precoce e mais efetivo em relação ao diagnóstico sorológico, principalmente em pacientes com falha imunológica como os pacientes deste estudo / Abstract: Infectious diseases are a major cause of life-threatening in patients with rheumatic disease. The cytomegalovirus, a betaherpesvírus, is the major cause of morbidity and mortality in immunosuppressed hosts may cause fever and damage to organs such as lung, liver, gastrointestinal tract, bone marrow and retina. Patients with SLE make use of immunosuppressive and are at risk of reactivate latent infection by HCMV. The HCMV-IgG seropositive patients using immunosuppressive should be monitored using the N-PCR and/or HCMV antigenemia for the early antiviral treatment can suppress viral reactivation and consequently the associated disease. Given the foregoing, monitored 40 patients were diagnosed with Systemic Lupus Erythematosus (SLE) for the active infection by the human cytomegalovirus (HCMV) using the techniques of nested-PCR (N-PCR) and Antigenemia (AGM). It was also checked the status serum IgM and IgG anti-HCMV by ELISA. For both, were needed 2 samples of blood per patient, obtained by venipuncture. The patients were in various stages of the disease lupus or specific treatment. The objectives of this study were: to detect the presence of DNA and antigen of the pp65 HCMV using the techniques of N PCR and AGM, respectively, in samples of peripheral blood of patients with SLE; verify the clinical impact caused by active infection and disease by HCMV patients in this series; verify the presence of immunoglobulins IgG and IgM anti-HCMV by the method of ELISA. All patients had serology IgG-HCMV reagent. Of the patients studied, 4/40 (10%) had active infection by HCMV detected by N-PCR and/or AGM. Of these, 3 were in lupus activity (SLEDAI ? 6). Among patients with active infection by HCMV, 2/4 (50%) had symptoms consistent with disease by HCMV. One of these patients developed epsis with pulmonar focus and died probably by HCMV. Despite the active HCMV infection were uncommon in these patients, the outcome are othen fatal. The use of early diagnosis tests of active infection by HCMV is very important because it allows a treatment earlier and more effective in relation to the serological diagnosis, especially in patients with immunity fail as the patients in this study / Mestrado / Ciencias Basicas / Mestre em Clinica Medica

Introduction: Patients with Systemic Lupus Erythematosus (SLE) have increased cardiovascular risk, endothelial dysfunction, and abnormal endothelial repair mechanisms. Vitamin D deficiency is common in SLE and has been associated with active disease and increased vascular stiffness. Myeloid angiogenic cells (MACs) repair damaged vessels by secretion of angiogenic factors and may be a target for vitamin D. Vitamin D may therefore be a novel therapy to improve cardiovascular risk in SLE patients. This study aimed to determine the effects of vitamin D on endothelial function and repair in patients with SLE. Methods: The effects of the active form of vitamin D (1,25(OH)2D3) were studied on MACs from vitamin D deficient SLE patients ex vivo. Functional models were developed to study MAC migration, adhesion and interaction with endothelial cells. Additional experiments used a model of healthy MACs treated with IFN-alpha. An observational study of clinically stable vitamin D deficient SLE patients being treated with high dose vitamin D over 3 months was used to investigate the effects of vitamin D on endothelial function as measured by flow-mediated dilatation (FMD). Results: MACs expressed markers consistent with an M2 macrophage phenotype and they enhanced endothelial network formation in vitro. SLE patients had an increased number of MACs; however, these were dysfunctional compared to healthy controls. Vitamin D increased the number, changed the phenotype and improved the functional capacity of SLE MACs ex vivo. In addition, the angiogenic capacity of SLE MACs was restored toward that of healthy controls via a reduction in the anti-angiogenic cytokine IP10. Vitamin D-treated MACs were also more able to protect endothelial cells against TNF-mediated down-regulation of endothelial nitric oxide synthase (eNOS). In SLE patients treated with vitamin D, there was a strong correlation between the change in serum 25(OH)D and the change in the ratio of endothelium-dependent:independent dilatation (r=-0.650, p=0.006). This was accompanied by an increase in the number of MACs at 3 months (p=0.015). These observations were independent of changes in serum PTH, calcium or lupus disease activity. Conclusions: Vitamin D can target MACs and therefore offers a novel approach to improve endothelial repair in patients with SLE. In addition, vitamin D treatment in lupus patients resulted in an improvement in endothelial function, related to the change in vitamin D status. These results suggest that vitamin D could improve surrogate markers of cardiovascular disease and thus reduce cardiovascular risk in this patient group.