Extrapancreatic Autoantibody Profiles in Type I Diabetes

Type I diabetes (T1D) is an autoimmune disease characterized by destruction of insulin-producing β-cells in the pancreas. Although several islet cell autoantigens are known, the breadth and spectrum of autoantibody targets has not been fully explored. Here the luciferase immunoprecipitation systems (LIPS) antibody profiling technology was used to study islet and other organ-specific autoantibody responses in parallel. Examination of an initial cohort of 93 controls and 50 T1D subjects revealed that 16% of the diabetic subjects showed anti-gastric ATPase autoantibodies which did not correlate with autoantibodies against GAD65, IA2, or IA2-β. A more detailed study of a second cohort with 18 potential autoantibody targets revealed marked heterogeneity in autoantibody responses against islet cell autoantigens including two polymorphic variants of ZnT8. A subset of T1D subjects exhibited autoantibodies against several organ-specific targets including gastric ATPase (11%), thyroid peroxidase (14%), and anti-IgA autoantibodies against tissue transglutaminase (12%). Although a few T1D subjects showed autoantibodies against a lung-associated protein KCNRG (6%) and S100-β (8%), no statistically significant autoantibodies were detected against several cytokines. Analysis of the overall autoantibody profiles using a heatmap revealed two major subgroups of approximately similar numbers, consisting of T1D subjects with and without organ-specific autoantibodies. Within the organ-specific subgroup, there was minimal overlap among anti-gastric ATPase, anti-thyroid peroxidase, and anti-transglutaminase seropositivity, and these autoantibodies did not correlate with islet cell autoantibodies. Examination of a third cohort, comprising prospectively collected longitudinal samples from high-risk individuals, revealed that anti-gastric ATPase autoantibodies were present in several individuals prior to detection of islet autoantibodies and before clinical onset of T1D. Taken together, these results suggest that autoantibody portraits derived from islet and organ-specific targets will likely be useful for enhancing the clinical management of T1D.     

Associations between interleukin-6 genetic polymorphisms and levels of autoantibodies to 60-kDa heat-shock proteins

AIMS: Previously we reported an association between levels of anti-Hsp60 autoantibodies and interleukin-6 (IL-6) -174 SNP in Finnish population. The aim of this study was to investigate the same association in an independent population and to study four recently described SNP in IL-6. MATERIALS AND METHODS: 313 healthy Hungarian subjects were recruited and genotyped for IL-6 -174(G-->C), -9316(T-->C), -1363(G-->T), +1753(C-->G), +2954(G-->C). IgG antibodies to Hsp60 were measured by ELISA. LD between SNPs was computed by Haploview 3.2 software. RESULTS: A strong association between IL-6 -174 polymorphism and anti-Hsp60 autoantibody levels was observed. Carriers of -174 CC genotype had significantly lower levels of anti-Hsp60 (p = 0.0052). Eight haplotypes were observed with five SNP-s and autoantibody levels in individuals carrying the most common haplotype (containing allele C of -174) were significantly lower than in all other genotype combinations (p = 0.026). CONCLUSIONS: Allele C of -174 promoter polymorphism of the IL-6 gene was repeatedly shown to be associated with low anti-Hsp60 autoantibody levels. Strong linkage in the IL-6 gene was observed and the most frequent haplotype containing the -174 C allele was significantly associated with autoantibody levels. Since the -174 SNP of IL-6 is a functional polymorphism, our results indicate for a direct regulatory effect of IL-6 genotypes in the determination of autoantibody levels.     

Dysregulation of proinflammatory and regulatory cytokines in HIV infected persons with active tuberculosis

Proinflammatory and regulatory cytokines have been implicated to play important role in immunopathology of HIV and tuberculosis (TB) infection. Capacity of unstimulated and mitogen-stimulated peripheral blood mononuclear cells (PBMCs) to secrete cytokines (interleukin (IL)-2, interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), IL-4, IL-10 and IL-6) was estimated for 15 HIV-TB coinfected patients, 22 HIV seropositives without TB, 32 HIV negative TB patients, and 36 healthy subjects. Dually infected patients had suppression of both Th1 and Th2 cytokine secretion as evidenced by significantly lower production of IL-2, IFN-gamma and TNF-alpha as well as IL-4 and IL-10. Production of IL-2 and TNF-alpha was significantly decreased only in case of HIV infection. Significantly higher IL-6 secretion was found in unstimulated cultures in dually infected patients. The mitogen induced cytokine secretion was generally lower in HIV-TB coinfected patients indicating profound perturbation of both Th1 and Th2 responses.     

Interleukin 1-beta responses to bacterial toxins and sudden infant death syndrome

We tested the hypothesis that significantly higher IL-1beta responses to toxic shock syndrome toxin (TSST) noted for parents of sudden infant death syndrome (SIDS) infants might be due in part to genetic factors such as the IL-1beta (C-511T) and IL-1RN (T+2018C) single nucleotide polymorphisms (SNP). The first objective was to assess the distribution of these polymorphisms among SIDS infants, parents of SIDS infants and controls, and two ethnic groups: Aboriginal Australians who have a high incidence of SIDS; and Bangladeshis who in Britain have a low incidence of SIDS compared with Europeans. The second objective was to assess IL-1beta responses to endotoxin and toxic shock syndrome toxin (TSST) from leukocytes of smokers and non-smokers in relation to these polymorphisms. There were major differences in the distributions of the IL-1beta (C-511T) SNP between Europeans and Bangladeshis (p=0.00) and between Europeans and Aboriginal Australians (p=0.00); however, they were similar for the Bangladeshi and Aboriginal Australian subjects. The allele frequency distribution of the IL-1RN (T+2018C) SNP for the Aboriginal Australians was statistically different from the European group (p=0.00), but it was not different from the Bangladeshi group (p=0.09). Compared with controls of European origin, there were no significant differences in the distribution of these polymorphisms among SIDS infants or parents of SIDS infants. For the IL-1beta (C-511T) SNP, the highest IL-1beta responses to endotoxin were obtained with leukocytes of non-smokers with the heterozygous CT genotype. Smokers had significantly lower levels of IL-1beta in response to endotoxin (p=0.01) and these differences were significant for donors with the wild type CC (p=0.00) and CT (p=0.03) genotypes. Similar patterns were observed for IL-1beta responses to TSST, but the differences were not significant. For the IL-1RN (T+2018C) SNP, the highest IL-1beta responses to endotoxin were obtained with leukocytes from non-smoker donors with the wildtype TT genotype and significantly lower responses were found with leukocytes from donors with the TC genotype (p=0.02). The responses of smokers were lower but the differences were significant only for donors with the TT genotype (p=0.00). Similar patterns were observed for IL-1beta responses to TSST, but the differences were not significant. IL-1beta responses to both endotoxin and TSST were increased for the small number of smokers with the TT genotype of the IL-1beta (C-511T) SNP. The TT genotype of the IL-1beta (C-511T) was found predominantly among Aboriginal Australian and Bangladeshi individuals but only a small proportion of Europeans. Smokers with the AA genotype of the IL-10 (G-1082A) SNP which is found predominantly among these two groups had significantly lower levels of IL-10 responses. If cigarette smoke enhances pro-inflammatory responses and reduces anti-inflammatory responses in individuals with these genotypes, this might partly explain the increased susceptibility of Aboriginal Australian infants to infections and SIDS.     

Cytokines and lymphocyte proliferation in juvenile and adult forms of paracoccidioidomycosis: comparison with infected and non-infected controls

Cellular immune response to Paracoccidiodes brasiliensis antigens (PbAg) was evaluated in patients with the juvenile (JF) and adult (AF) forms of paracoccidioidomycosis as well as in a group of infected individuals living in the endemic area but without any clinical manifestation of the disease. The immune profile of this group of paracoccidioidomycosis-infected individuals was characterized by: 1) a positive skin test to P. brasiliensis antigen; 2) absence of specific antibodies; 3) a vigorous lymphoproliferative response to PbAg; and 4) a typical Th1 pattern of cytokines, with production of IFN-gamma and basal levels of IL-4, IL-5 and IL-10. At the opposite end of the spectrum were the JF patients whose proliferative response to PbAg was significantly impaired and whose cytokine pattern was characteristically Th2, i.e. lower IFN-gamma secretion and significantly higher levels of IL-4, IL-5 and IL-10. These profiles are compatible with forms of higher and lower resistance, respectively. Intermediate immune responses were observed in AF patients, whose specific lymphoproliferative response was lower than in the paracoccidioidomycosis-infected group but higher than in the JF patients. The secretion of IFN-gamma and IL-10 did not differ from the JF group, although IL-4 and IL-5 levels were significantly lower. Since AF patients are able to control fungal dissemination for decades, they can be considered more resistant than JF patients, who manifest the disease soon after infection.     

T-cell responses associated with resistance to Leishmania infection in individuals from endemic areas for Leishmania (Viannia) braziliensis

Subclinical or asymptomatic infection is documented in individuals living in endemic areas for leishmaniasis suggesting that the development of an appropriate immune response can control parasite replication and maintain tissue integrity. A low morbidity indicates that intrinsic factors could favor resistance to Leishmania infection. Herein, leishmanial T-cell responses induced in subjects with low susceptibility to leishmaniasis as asymptomatic subjects were compared to those observed in cured cutaneous leishmaniasis (CCL) patients, who controlled the disease after antimonial therapy. All of them have shown maintenance of specific long-term immune responses characterized by expansion of higher proportions of CD4+ as compared to CD8+ Leishmania reactive T-lymphocytes. Asymptomatic subjects had lower indexes of in vitro Leishmania induced lymphoproliferative responses and interferon-gamma (IFN-gamma) production in comparison to CCL patients. On the other hand, interleukin (IL-10) production was much higher in asymptomatics than in CCL, while no differences in IL-5 levels were found. In conclusion, long lived T-cell responses achieved by asymptomatic individuals differed from those who had developed symptomatic leishmaniasis in terms of intensity of lymphocyte activation (proliferation or IFN-gamma) and regulatory mechanisms (IL-10). The absence of the disease in asymptomatics could be explained by their intrinsic ability to create a balance between immunoregulatory (IL-10) and effector cytokines (IFN-gamma), leading to parasite destruction without producing skin tissue damage. The establishment of profiles of cell-mediated immune responses associated with resistance against Leishmania infection is likely to make new inroads into understanding the long-lived immune protection against the disease.     

Interleukin-1 receptor antagonist gene polymorphism and circulating levels of human immunodeficiency virus type 1 RNA in Brazilian women

Interleukin-1 receptor antagonist (IL-1ra) gene polymorphisms in 83 human immunodeficiency virus (HIV)-seropositive women were evaluated. Fourteen of the subjects (16.9%) were homozygous for IL-1ra allele 2 (IL-1RN*2). These women had a lower median level of HIV RNA than did women homozygous for allele 1 (IL-1RN*1) (P = 0.01) or heterozygous for both alleles (P = 0.04). Among 46 subjects not receiving antiretroviral treatment, HIV levels were also reduced in IL-1RN*2 homozygous individuals (P < 0.05). There was no relation between IL-1ra alleles and CD4 levels.     

Plasma microRNA expression and immunoregulatory cytokines in an Indian population occupationally exposed to cadmium

Following its accumulation in the body, cadmium (Cd) exposure is associated with devastating effects on multiple organ system of the human body. The immune system is one of the sensitive targets for Cd-induced toxicity. Recently, studies have demonstrated a significant role of Cd in inducing epigenetic alterations. With this background, the present study was planned to study the changes in candidate microRNA (miRNA) expression associated with immune regulation in occupationally Cd-exposed workers. One hundred individuals involved in welding and metal handicraft manufacturing, while 80 apparently healthy subjects without any prior history of occupational exposure were recruited for the study. Blood Cd level was determined by atomic absorption spectrometry. Serum cytokine levels were measured using an enzyme-linked immunosorbent assay and serum miRNA expression of candidate miRNAs (miR-146a, miR-210, and miR-222) were determined by real-time polymerase chain reaction. The median Cd level (2.40 μg/L) in the occupationally exposed workers was significantly higher than the nonexposed subjects (0.90 μg/L). Among the cytokines, interleukin-4 (IL-4), and tumor necrosis factor-alpha (TNF-α) were significantly higher while IL-2 and IL-10 were significantly lower in the exposed. The expression level of miR-146a and miR-222 were significantly different between the groups with the former showing downregulation and later showing upregulation. Correlation analysis revealed a positive and negative association of miR-222 and miR-146a with blood cadmium level, IL-17 as well as TNF-α, respectively. Furthermore, the in-silico analysis revealed a significant role of the studied miRNAs in various cellular and genetic pathways. The findings of the present study demonstrate significant involvement of Cd-induced alteration in miRNAs in varied immune regulatory changes in exposed individuals.     

Naturally occurring IgM anti-leukocyte autoantibodies (IgM-ALA) inhibit T cell activation and chemotaxis

The physiological relevance of naturally occurring IgM-ALA remains to be elucidated. These autoantibodies are present from birth and increase in diverse inflammatory states that are both infectious and noninfectious. Clinical observations showing significantly less acute allograft rejections in recipients having high IgM-ALA levels, led us to investigate whether IgM-ALA could have a functional role in attenuating T cell mediated inflammatory responses. In pursuit of this hypothesis, we did studies using IgM purified from the serum of normal individuals, patients with end stage renal disease, and HIV-1 infection. All preparations of IgM immunoprecipitated certain receptors e.g., CD3, CD4, CCR5, and CXCR4 from whole cell lysates but failed to immunoprecipitate IL-2R and HLA Ags. In physiological doses IgM down-regulated CD4, CD2 and CD86 but not CD8 and CD28, inhibited T cell proliferation, decreased production of certain proinflammatory cytokines e.g., TNF-alpha, IL-13 and IL-2, but not IFN- gamma, IL-1beta, GM-CSF, IL-6 and IL-8 and inhibited leukocyte chemotaxis. These inhibitory effects were more pronounced when using IgM from patients with high levels of IgM-ALA and these inhibitory effects were significantly reduced after using IgM preabsorbed with leukocytes. IgM-ALA binding to leukocytes was found to be highly specific, as <10% of IgM secreting B cell clones had IgM-ALA specificity with some clones having specificity for either T cells or monocytes. These findings support the concept that IgM-ALA provides an innate mechanism to regulate T cell mediated inflammatory responses.     

High-density lipoproteins attenuate interleukin-6 production in endothelial cells exposed to pro-inflammatory stimuli

The purpose of the present study was to investigate the ability of high-density lipoproteins (HDL) to attenuate endothelial dysfunction, by assessing down-regulation of cytokine-induced interleukin-6 (IL-6) production in cultured endothelial cells, and measuring plasma IL-6 levels in three groups of healthy individuals with low, average, or high plasma HDL-cholesterol. Human plasma HDL caused a concentration-dependent inhibition of TNFalpha-induced IL-6 production in human endothelial cells (by 58.5+/-1.5% at 2 mg of HDL-protein/ml). Reconstituted HDL made with apolipoprotein A-I (apoA-I) and phosphatidylcholine were as effective as plasma HDL, while lipid-free apoA-I or phosphatidylcholine liposomes had no effect. HDL attenuated IL-6 mRNA levels, an effect which occurs through inhibition of p38 MAP kinase. The median plasma IL-6 concentration was significantly higher in subjects with low HDL-cholesterol (2.54 pg/ml) compared with those with average or high HDL-cholesterol (1.31 pg/ml and 1.47 pg/ml, respectively). When all subjects were considered together, a lower HDL-cholesterol was the strongest independent predictor of higher IL-6 (F=25.38, P<0.001). By inhibiting IL-6 production and lowering plasma IL-6 concentration, HDL may limit the pro-atherogenic effects of both acute and chronic inflammatory states, of which IL-6 is a key orchestrator.     

Impaired expression of high affinity interleukin 2 receptor on activated lymphocytes from patients with systemic lupus erythematosus

Peripheral lymphocytes stimulated with phytohemagglutinin (PHA-blasts) were examined for their responsiveness to exogenous interleukin 2 (IL-2). The proliferative response of PHA-blasts to IL-2 was significantly lower in patients with systemic lupus erythematosus (SLE) than in normal subjects. To clarify the reason for this defect, the expression of IL-2 receptor (IL-2R) on PHA-blasts was investigated using anti-Tac antibody and purified IL-2. Cytofluorometric analysis showed no statistical differences in the Tac positivity of PHA-blasts among normal subjects and patients with active and inactive SLE. Scatchard analysis using 125I-labeled anti-Tac monoclonal antibody revealed that the number of Tac epitopes on PHA-blasts was also not different among them. Next, the affinity of IL-2R expressed on PHA-blasts was determined by Scatchard analysis using radiolabeled IL-2 as a ligand. The number of high affinity IL-2R on the PHA-blasts was significantly decreased in patients with active and inactive SLE, as compared with normal subjects. The responsiveness of PHA-blasts to exogenous IL-2 was well correlated to the number of high affinity IL-2R, but not to the number of Tac epitopes or total IL-2R. Inasmuch as high affinity components of IL-2R are functionally active, the defective expression of high affinity IL-2R may be responsible for the T cell dysfunctions in SLE.     

Cytokine response following transthoracic and transhiatal esophagectomy in patients with esophageal cancer

Curative esophageal resection is usually performed using either a transthoracic (TT) or transhiatal (TH) approach. Forty patients with esophageal squamous cell carcinoma who underwent esophagectomies (24 TT and 16 TH), 12 patients who underwent surgery for gastric cancer, and 16 healthy individuals were enrolled in this study. Blood samples were taken from the patients, pre- and post-surgery. The levels of synthesis of T-helper 1 and 2 cytokines were assessed in vitro in the presence of mitogen. Our initial data indicated that at admission, 24 h before surgery, blood cells from both groups of esophageal cancer patients produced significantly lower levels of the Th1 cytokines, IFN-gamma and IL-2 than those from cells of healthy donors. Cells collected from gastric cancer patients prior to surgery produced intermediate levels of IFN-gamma and IL-2: significantly lower than healthy donors, and slightly more (non-significant) than esophageal cancer patients. These results contrast with those for the production of Th2 cytokines prior to surgery, which did not differ significantly between any groups: either the esophageal or gastric cancer patients, or healthy donors. Th1 and Th2 cytokine production was then studied using blood cells collected seven days after surgery. Cells from both groups of esophageal cancer patients, undergoing either TT or TH surgery, produced significantly lower levels of the Th1 cytokines, IFN-gamma and IL-2 than those from cells of gastric cancer patients who had undergone surgery. Postoperative and preoperative production was compared. For patients who had undergone TT esophageal resection, we observed that the post-operative production of IL-2, IL-5 and IL-13 was significantly lower than the pre-operative production of those cytokines. Such reduced post-operative compared to pre-operative production was only significant in patients who had undergone TT esophagectomy. A similar, but non-significant trend was observed in patients who had undergone either TH esophagectomy, or gastrectomy. The results indicate that digestive tract cancer patients, both esophageal and gastric, have (prior to surgery), a significantly reduced, basal, mitogen-induced production of Th1 but not of Th2 cytokine. Post-operatively, a significantly reduced production of Th1 and Th2 cytokines, except for IFN-gamma, was observed only in patients who had undergone surgical esophageal resection using the TT method.     

Bullous pemphigoid: interaction of interleukin 5, anti-basement membrane zone antibodies and eosinophils. A preliminary observation.

Bullous pemphigoid (BP) is an autoimmune blistering skin disease characterized by the presence of autoantibodies to normal components of the hemidesmosome (BPAg1 and BPAg2). Histology of skin lesions demonstrate a subepidermal vesicle and a predominantly eosinophilic dermal cellular infiltrate. Interleukin 5 (IL-5) plays a major role in eosinophil recruitment and function. Using an ELISA, we investigated the levels of IL-5 in the sera and blister fluid of BP patients with active disease and those in prolonged clinical remission treated with intravenous immunoglobulin, and compared it to that in normal controls. Significantly increased levels of IL-5 were detected in the serum and particularly the blister fluid of patients with active disease (P=0.0043) when compared to levels in normal controls. There was no significant difference in IL-5 levels in patients in prolonged clinical remission compared to normal control serum. In an immunoblot assay, using bovine gingival lysate as substrate, we determined the presence of IgG and IgE autoantibodies specific to basement membrane zone proteins in blister fluid and serum of BP patients. IgG autoantibodies to BPAg1 and BPAg2 were detected in both blister fluid and serum of patients, whereas IgE autoantibodies, in much lower titers, were detected to only BPAg1. Elevated levels of blood and tissue eosinophilia were observed in these patients. Based on these data, we present a preliminary hypothesis for the initiation, progression and localization of blister formation in BP.     

Lymphocyte subsets and cytokines in women with gestational diabetes mellitus and their newborn

This study aimed to identify potential immunological markers for predicting type 1 diabetes in patients with gestational diabetes mellitus (GDM) and any immunological impairment in their newborn. In 62 GDM patients and 74 women with normal glucose tolerance (NGT), and their babies, we assessed total lymphocytes, T lymphocyte subsets CD3 and CD8 expressing T cell receptor (TCR) alpha/beta or gamma/delta, CD16 and CD19, pancreatic autoantibodies and cytokines (IL-5, IL-2, soluble receptor IL-2). At delivery, umbilical cord blood samples were taken for lymphocyte subpopulations and cytokine measurements. GDM mothers had higher levels of total lymphocytes, CD8 expressing TCR gamma/delta, and lower levels of CD3 expressing TCR alpha/beta than NGT controls. Insulin-treated GDM mothers had lower CD4 and CD4/CD8 ratios, and higher CD8 and IL-5 than diet-treated GDM or controls. Five women were positive for pancreatic autoantibodies, with lower CD4 (p<0.01) and CD4/CD8 ratios (p<0.05), and higher CD8 (p<0.03) and CD19 than GDM and control mothers negative for autoantibodies. GDM newborn had higher CD8 gamma/delta and lower CD16 than NGT babies. There were no significant differences in TNF-alpha concentrations in the cord blood obtained from the GDM and NGT newborn. In conclusion, GDM women and their newborn have lymphocyte subset impairments, which are more important in patients positive for autoantibodies and/or treated with insulin.     

Differences in the genotypes and plasma concentrations of the INTERLEUKIN-1 receptor antagonist in black and white South African asthmatics and control subjects

The allelic frequency of a variable tandem repeat (VNTR) polymorphism in intron 2 of the IL-1 Ra gene was studied in black and white patients with asthma as well as control individuals. The plasma IL-1 Ra concentration was also determined in asthmatics and compared to control individuals. The 410-bp allele of the IL-1 Ra was significantly increased in all black subjects (90%) as compared to all white subjects (74%, P<0.0001), while the 240-bp allele was significantly reduced in all black subjects (11%) as compared to all white subjects (27%, P<0.0001). There was no difference in the frequency of the VNTR of the IL-1 Ra between black asthmatics and black controls and between white asthmatics and white controls. The IL-1 Ra levels were significantly increased in black and white patients with severe or moderate asthma as compared to patients with mild asthma. Increased plasma concentrations of the IL-1 Ra was found to be associated with disease severity in all asthmatic patients.     

IL-10 Dependent Suppression of Type 1, Type 2 and Type 17 Cytokines in Active Pulmonary Tuberculosis

Background

Although Type 1 cytokine responses are considered protective in pulmonary tuberculosis (PTB), their role as well as those of Type 2, 17 and immunoregulatory cytokines in tuberculous lymphadenitis (TBL) and latent tuberculosis (LTB) have not been well studied.

Aim and Methods

To identify cytokine responses associated with pulmonary tuberculosis (TB), TB lymphadenitits and latent TB, we examined mycobacterial antigen-specific immune responses of PTB, TBL and LTB individuals. More specifically, we examined ESAT-6 and CFP-10 induced Type 1, Type 2 and Type 17 cytokine production and their regulation using multiplex ELISA.

Results

PTB individuals exhibited a significantly lower baseline as well as antigen-specific production of Type 1 (IFNγ, TNFα and IL-2); Type 2 (IL-4) and Type 17 (IL-17A and IL-17F) cytokines in comparison to both TBL and LTB individuals. TBL individuals exhibited significantly lower antigen-specific IFNγ responses alone in comparison to LTB individuals. Although, IL-10 levels were not significantly higher, neutralization of IL-10 during antigen stimulation resulted in significantly enhanced production of IFNγ, IL-4 and IL-17A in PTB individuals, indicating that IL-10 mediates (at least partially) the suppression of cytokine responses in PTB.

Conclusion

Pulmonary TB is characterized by an IL-10 dependent antigen-specific suppression of Type 1, Type 2 and Type 17 cytokines, reflecting an important association of these cytokines in the pathogenesis of active TB.     

Increased pro-inflammatory cytokines (TNF-alpha and IL-6) and anti-inflammatory compounds (sTNFRp55 and sTNFRp75) in Brazilian patients during exanthematic dengue fever.

Pro-inflammatory cytokines, tumor necrosis factor (TNF-alpha), interleukin-6 (IL-6) and interleukin-1beta (IL-1beta) as well as anti-inflammatory compounds, soluble TNF-Receptor p55 (sTNFRp55), sTNFRp75 and IL-1 receptor antagonist (sIL-1Ra), were investigated in 34 Brazilian cases of dengue fever (DF) originated from a study of exanthematic virosis. The presence of pro-inflammatory cytokines was detected in sera from these patients by ELISA. TNF-alpha and IL-6 levels were significantly higher than control subjects in 32% and 52% patients, respectively. To our knowledge this was the first time a receptor antagonist and soluble receptors for cytokines were detected in sera obtained during exanthematic DF without hemorrhagic manifestations. Both sTNFRp55 and sTNFRp75 were consistently elevated in 42% and 84% patients, respectively. Most patients had IL-1beta levels not different from those of normal subjects, except for one case. Only 16% patients had altered levels of IL-1Ra. Previous studies in dengue hemorrhagic fever patients demonstrated production of these soluble factors; here we observed that they are found in absence of hemorrhagic manifestations. The possible role of these anti-inflammatory compounds in immune cell activation and in regulating cytokine-mediated pathogenesis during dengue infection is discussed.     

Effect of azacytidine in the release of leukemia inhibitory factor,oncostatin m,interleukin (IL)-6, and IL-11 by mononuclear cells of patients with refractory anemia

5-azacytidine (AZA) yields hematologic improvement in patients with myelodysplastic syndromes (MDS). Ineffective hemopoiesis in MDS produce the paradox of high intramedullary cellularity with peripheral cytopenias. Leukemia inhibitory factor (LIF), oncostatin M (OSM), interleukin (IL)-6, and IL-11 regulate hemopoiesis and LIF, OSM, and IL-6 also inhibit the proliferation of myeloid leukemic cell lines through the signal-transducing subunit gp130. These IL-6-type cytokines were measured by enzyme-linked immunosorbent assay in cell culture supernatants (SN) obtained from peripheral blood mononuclear cells (MNC) and monocyte-depleted MNC of patients with refractory anemia (RA; n=12) and healthy individuals (n=10). AZA down-regulated OSM, IL-6, and IL-11 release by MNC of patients but not by MNC from healthy individuals. Patient's SN had significantly lower concentrations of LIF, OSM, and IL-11 than SN of normal subjects. When monocyte-depleted MNC of patients were stimulated with phytohemagglutinin a significant increment in OSM levels was observed. In contrast, monocyte depletion in healthy subjects did not cause any significant change in OSM values. We conclude that: (a) AZA inhibits the release of OSM, IL-6, and IL-11 exclusively in RA-diseased MNC, (b) Patient's MNC release subnormal amounts of LIF, OSM, and IL-11, and (c) RA-derived monocytes probably down-regulate OSM release by phytohemagglutinin-activated MNC.     

Expression of three cross-reactive idiotypes on rheumatoid factor autoantibodies from patients with autoimmune diseases and seropositive adults

Approximately one-half of human monoclonal IgM anti-IgG autoantibodies (rheumatoid factors (RF] from unrelated individuals with cryoglobulinemia coordinately express three cross-reactive idiotypic antigens (CRI). The CRI are detected with: 1) monoclonal antibody 17.109, which recognizes a conformation-dependent CRI on K-light chains; and 2) two rabbit anti-peptide antibodies that react with primary sequence-dependent CRI (PSL2 and PSL3) corresponding to the conserved second and third K-chain complementarity-determining regions, respectively. In the present experiments, the structural features of polyclonal RF autoantibodies from diverse patients with rheumatoid arthritis and from those with primary Sj?gren's syndrome, and from seropositive elderly subjects without overt autoimmune diseases, were investigated with these three defined anti-CRI reagents. The pattern of expression of the CRI differed among patient groups. Only the RF autoantibodies from Sj?gren's syndrome patients frequently displayed all three CRI. However, the RF from nearly every subject tested, including patients with rheumatoid arthritis, were enriched in the primary sequence-dependent PSL2-CRI as compared to RF-depleted Ig from the same subjects. Amino acid sequence analysis of monoclonal IgM-RF indicates that PSL2-CRI-positive light chains probably represent the products of a single Vk gene. Therefore, a proportion of the polyclonal RF from different autoimmune states may represent somatic variants of this germ-line RF Vk gene which retain the PSL2 sequence as a common element.