Contribution of Complement System pathways to the killing of Leptospira spp.

The Complement System (CS) plays an important role in the immune response against leptospirosis and can be activated by the Alternative and Lectin Pathways (Innate Immunity) and by the Classical Pathway (Acquired Immunity). Here we analyzed a broad range of nonpathogenic and pathogenic Leptospira strains considering their interaction with each CS pathway. We determined bacterial survival rate and CS protein deposition in the presence of purified proteins, specific component depleted sera and NHS treated with the chelating agents EDTA (inhibits all three activation pathways) or EGTA (inhibits the Classical and Lectin Pathways). We suggest that the Lectin and the Alternative Pathways have an important role to eliminate saprophytic leptospires since i) approximately 50% survival of both saprophytic strains was observed in the presence of MBL-deficient serum; ii) approximately 50% survival of Leptospira biflexa Patoc I was observed in the presence of NHS - EGTA and iii) C1q-depleted serum caused significant bacterial lysis. In all serovars investigated the deposition of C5-C9 proteins on saprophytic Leptospira strains was more pronounced when compared to pathogenic species confirming previous studies in the literature. No difference on C3 deposition was observed between nonpathogenic and pathogenic strains. In conclusion, Leptospira strains interact to different degrees with CS proteins, especially those necessary to form MAC, indicating that some strains and specific ligands could favor the binding of certain CS proteins.

Primary Immunodeficiencies in a Mesoregion of São Paulo, Brazil: Epidemiologic, Clinical, and Geospatial Approach.

Background: Primary immunodeficiencies (PIDs) are rare genetic disorders leading to immunologic abnormalities that can affect different organs and systems. We determined the epidemiology, clinical, and geospatial characteristics of PID disorders among patients diagnosed over a 5 year period in a reference hospital covering a mesoregion in São Paulo, Brazil. Methods: A retrospective analysis of 39 patients with recognizable PIDs according to the criteria of the European Society of Primary Immunodeficiencies were enrolled. Thirty-four patients came from outpatient immunodeficiency clinics and five patients from active search. Demographic, clinical, and immunologic data were collected, and maps were constructed using a geographic information system. Results: The ratio of females to males was 1.4:1, and 48.7% of patients were younger than 17 years of age. The mean age at the onset of symptoms in children was 2.0 years [standard error of the mean (SEM), 1.7 years] and the diagnosis lag was 5.1 years (SEM, 3.1 years); the mean age at diagnosis in adults was 16.3 years (SEM, 11.8 years) and the lag was 10.8 years (SEM, 10.9 years). Antibody deficiency and common variable immunodeficiencies were the most common categories and phenotypes, respectively. The need for intravenous antibiotics and respiratory tract infections were the most prevalent warning signs, with an overall mortality rate of 15.3%. Autoimmune diseases were diagnosed in 56.4% and visceral leishmaniasis in 5.1% of patients. In the active search, 29 patients were investigated and 17.2% were diagnosed; early diagnosis, the involvement of multidisciplinary professionals, and dissemination of knowledge achieved milestone benefits. The distribution of PID networks in Brazil shows great asymmetry between regions and at a regional level; it was shown that the patients lived mainly in Presidente Prudente municipality. Conclusions: The implementation of an immunodeficiency outpatient clinic in a referral hospital covering a mesoregion with a large population has led to the generation of policies and practices to improve the diagnosis, quality of life, and care of patients with PIDs and their families. Furthermore, the search for hospitalized patients with warning signs for PIDs showed great benefits. Inequality in the distribution of PID network centers in Brazil was demonstrated.

Th17 pathway in recent-onset autoimmune diabetes.

AIMS: Evaluate the participation of IL-17 pathway in T1D pathogenesis. T helper 17 cells are potent, highly inflammatory cells that produce interleukin 17A (IL-17A), considered a mediator of various immune disorders. However, their role in Type 1 diabetes (T1D) pathogenesis in humans is not totally elucidated. METHODS: The expression of IL-17 Receptor A (IL-17RA) in peripheral T lymphocytes and IL-17A serum levels in recent-onset patients with T1D were compared with healthy controls. IL-17A gene variants were evaluated in a greater cohort. RESULTS: Patients with recent-onset T1D (less than 6 months of diagnosis) exhibited lower expression of IL-17RA in CD3+ T (% of cells = 31.3% × 43.6%; p = .041) and CD4+ T cells (11.1% × 25.2%; p = .0019) and lower number of IL-17RA in CD4+ T cells (MFI = 1.16 × 4.56; p = .03) than controls. IL-17RA expression in CD8+ T cells and IL-17A serum levels were similar in both groups. The coding regions and boundary intron sequences of IL17A were sequenced. Seventeen allelic variants, including three novel variants in exon 3 (3'UTR n) were identified, but no one was associated with T1D susceptibility, as well as the resulting haplotypes and diplotypes. The expression of IL-17RA was not correlated with metabolic variables (glucose and HbA1c levels) or pancreatic autoantibodies titers. CONCLUSIONS: The lower expression of IL-17RA in CD3+ and CD4+ T cells suggests a reduced effect of IL-17A in immune response of recent-onset T1D patients, at least at peripheral tissues. IL-17A allelic variants were not related with T1D susceptibility.

Binding of human complement C1 sterase inhibitor to Leptospira spp.

Leptospirosis is an important zoonosis of global importance caused by bacteria Leptospira spp. Pathogenic Leptospira is resistant to Complement System killing while non-pathogenic Leptospira is rapidly killed by exposure to normal human serum (NHS). Pathogenic Leptospira interact with Complement Regulators such as Factor H, C4b binding protein and Vitronectin avoiding Complement activation and killing by Alternative and Classical Pathways. One important regulator is C1-inhibitor (C1INH) that interacts with C1s or MASPs controlling the cleavage of C4 and C2 molecules, thereby inhibiting the activation of the Classical and Lectin Pathways. In this study, we demonstrate that attenuated, saprophytic and pathogenic Leptospira interact with C1INH that maintain its regulatory capacity of interaction with C1s preventing the activation of Complement system. Although the interaction with C1INH is not crucial for pathogenic Leptospira survival, it seems to be important for the survival of attenuated and saprophytic Leptospira in normal human serum.

Chemokine, cytokine and type I interferon production induced by Toll-like receptor activation in common variable immune deficiency.

Common variable immunodeficiency (CVID) is the most common symptomatic primary antibody deficiency and is associated with recurrent infections and chronic inflammatory diseases. We evaluated the ability of Toll-like receptor (TLR) ligands to induce secretion of chemokines, cytokines and type I interferons by peripheral blood mononuclear cells (PBMCs) from CVID patients. High levels of CXCL10, CCL2, CXCL9, CCL5, CXCL8, and IL-6 were detected in sera of CVID patients compared with healthy controls. Increased chemokine levels were observed in unstimulated PBMCs, but after stimulation with TLR2 and TLR4 agonists, equivalent chemokine and pro-inflammatory cytokine secretion, as in healthy controls, was observed, whereas TLR4 agonist induced a decreased secretion of CCL2 and CXCL8 and increased secretion of TNF. Decreased IFN-α secretion induced by TLR7/TLR8 activation was observed in CVID, which was recovered with TLR9 signaling. Our findings revealed that TLR9 activation has an adjuvant effect on the altered type I response in CVID.

Impaired CD8(+) T cell responses upon Toll-like receptor activation in common variable immunodeficiency.

BACKGROUND: Infections caused by bacteria or viruses are frequent in common variable immunodeficiency (CVID) patients due to antibody deficiencies, which may be associated with altered T cell function. CVID patients are frequently in contact with pathogen-associated molecular patterns (PAMPs), leading to the activation of innate immunity through Toll-like receptors (TLR) affecting T cell activation. We evaluated the effect of TLR activation on T cells in CVID patients undergoing intravenous immunoglobulin (IVIg) replacement using synthetic ligands. METHODS: Expression of exhaustion, activation and maturation markers on T cells from peripheral blood as well as regulatory T cells and follicular T cells in peripheral blood mononuclear cells (PBMCs) from CVID and healthy individuals were evaluated by flow cytometry. PBMCs cultured with TLR agonists were assessed for intracellular IFN-γ, TNF, IL-10, IL-17a or IL-22 secretion as monofunctional or polyfunctional T cells (simultaneous cytokine secretion) by flow cytometry. RESULTS: We found increased expression of the exhaustion marker PD-1 on effector memory CD4(+) T cells (CD45RA(-)CCR7(-)) in the peripheral blood and increased expression of CD38 in terminally differentiated CD8(+) T cells (CD45RA(+)CCR7(-)). Furthermore, a decreased frequency of naïve regulatory T cells (CD45RA(+)Foxp3(low)), but not of activated regulatory T cells (CD45RA(-)Foxp3(high)) was detected in CVID patients with splenomegaly, the non-infectious manifestation in this CVID cohort (43.7 %). Moreover, the frequency of peripheral blood follicular helper T cells (CD3(+)CD4(+)CXCR5(+)PD-1(+)ICOS(+)) was similar between the CVID and control groups. Upon in vitro TLR3 activation, a decreased frequency of CD8(+) T cells secreting IFN-γ, IL-17a or IL-22 was detected in the CVID group compared to the control group. However, a TLR7/TLR8 agonist and staphylococcal enterotoxin B induced an increased Th22/Tc22 (IL-22(+), IFN-γ(-), IL-17a(-)) response in CVID patients. Both TLR2 and TLR7/8/CL097 activation induced an increased response of CD4(+) T cells secreting three cytokines (IL-17a, IL-22 and TNF)in CVID patients, whereas CD8(+) T cells were unresponsive to these stimuli. CONCLUSION: The data show that despite the unresponsive profile of CD8(+) T cells to TLR activation, CD4(+) T cells and Tc22/Th22 cells are responsive, suggesting that activation of innate immunity by TLRs could be a strategy to stimulate CD4(+) T cells in CVID.

Chronic mucocutaneous candidiasis and systemic lupus erythematosus: a new variant of chronic mucocutaneous candidiasis?

Chronic mucocutaneous candidiasis (CMC) is characterized by susceptibility to Candida infection of skin, nails, and mucous membranes. Autoimmune endocrinopathies are common in CMC patients, but there are no reports of the involvement of systemic autoimmune disorders. We present here the first case of this kind of association in a patient with an autosomal dominant variant of CMC. The individual had had this disorder since childhood and systemic lupus erythematosus with secondary antiphospholipid syndrome, as well as renal, articular and hepatic manifestations without thymoma.

Retinoic acid inhibits dendritic cell differentiation driven by interleukin-4.

All-trans-retinoic acid (atRA) appears to affect Th1-Th2 differentiation and its effects on immune responses might also be mediated by dendritic cell (DC). Nonetheless, studies have been showing contradictory results since was observed either induction or inhibition of DC differentiation. Our aim was to investigate atRA action on human monocyte derived DC differentiation. For this purpose we tested pharmacological and physiological doses of atRA with or without cytokines. Cell phenotypes were analyzed by flow cytometry and function was investigated by phagocytosis and respiratory burst. DC, positive control group, was differentiated with GM-CSF and IL-4 and maturated with TNF-alpha. We demonstrated that atRA effects depend on the dose used as pharmacological doses inhibited expression of all phenotypic markers tested while a physiological dose caused cell differentiation. However, atRA combined or not with cytokines did not promote DC differentiation. In fact, atRA was detrimental on IL-4 property as a DC inductor.

Esophageal cancer associated with chronic mucocutaneous candidiasis. Could chronic candidiasis lead to esophageal cancer?

Chronic mucocutaneous candidiasis (CMC) is a rare disease associated with immunodeficiency and characterized by persistent and refractory infections of the skin, appendages and mucous membranes caused by members of the genus Candida. Several different disorders are classified under this common denominator, including chronic and recurrent mucocutaneous infections due to Candida spp., which are sometimes linked to autoimmune endocrinopathies. These fungal infections are usually confined to the mucocutaneous surface, with little propensity for systemic disease or septicemia. We describe a patient with CMC who had an esophageal candidiasis refractory to treatment for decades and who developed an epidermoid esophageal cancer. No risk factors such as familiar susceptibility, smoking, alcohol drinking, or living in an endemic area were verified. This case report suggests the participation of nitrosamine compounds produced by chronic Candida infections as a risk factor for esophageal cancer in a patient with autosomal-dominant chronic mucocutaneous candidiasis.

Evaluation of dendritic cell-tumor cell hybrid vaccine storage.

Clinical trials using dendritic cells (DCs) to treat cancer patients have generated promising results in recent years. However, even simple aspects of this therapy are still not well understood, including the storage and distribution of manufactured vaccines. These processes are essential and must be elucidated in order to reduce costs. We evaluated the effects of different storage conditions on vaccine functionality using mixed lymphocyte reaction (MLR). Vaccine storage at 4°C for up to 72 h had no significant effect on vaccine activity. Shipping to distant places is possible, if vaccines are kept at 4°C and used up to 3 days after manufacture date.

Low penetrance, broad resistance, and favorable outcome of interleukin 12 receptor beta1 deficiency: medical and immunological implications.

The clinical phenotype of interleukin 12 receptor beta1 chain (IL-12Rbeta1) deficiency and the function of human IL-12 in host defense remain largely unknown, due to the small number of patients reported. We now report 41 patients with complete IL-12Rbeta1 deficiency from 17 countries. The only opportunistic infections observed, in 34 patients, were of childhood onset and caused by weakly virulent Salmonella or Mycobacteria (Bacille Calmette-Guérin -BCG- and environmental Mycobacteria). Three patients had clinical tuberculosis, one of whom also had salmonellosis. Unlike salmonellosis, mycobacterial infections did not recur. BCG inoculation and BCG disease were both effective against subsequent environmental mycobacteriosis, but not against salmonellosis. Excluding the probands, seven of the 12 affected siblings have remained free of case-definition opportunistic infection. Finally, only five deaths occurred in childhood, and the remaining 36 patients are alive and well. Thus, a diagnosis of IL-12Rbeta1 deficiency should be considered in children with opportunistic mycobacteriosis or salmonellosis; healthy siblings of probands and selected cases of tuberculosis should also be investigated. The overall prognosis is good due to broad resistance to infection and the low penetrance and favorable outcome of infections. Unexpectedly, human IL-12 is redundant in protective immunity against most microorganisms other than Mycobacteria and Salmonella. Moreover, IL-12 is redundant for primary immunity to Mycobacteria and Salmonella in many individuals and for secondary immunity to Mycobacteria but not to Salmonella in most.