Inflammatory biomarkers in infective endocarditis: machine learning to predict mortality.

Infective endocarditis (IE) is the cardiac disease with the highest rates of mortality. New biomarkers that are able to identify patients at risk for death are required to improve patient management and outcome. This study aims to investigate if cytokines, chemokines and growth factors measured at IE diagnosis can predict mortality. Patients with definite IE, according to the Duke's modified criteria, were included. Using high-performance Luminex assay, 27 different cytokines, chemokines and growth factors were analyzed. Machine learning techniques were used for the prediction of death and subsequently creating a decision tree, in which the cytokines, chemokines and growth factors were analyzed together with C-reactive protein (CRP). Sixty-nine patients were included, 41 (59%) male, median age 54 [interquartile range (IQR) = 41-65 years] and median time between onset of the symptoms and diagnosis was 12 days (IQR = 5-30 days). The in-hospital mortality was 26% (n = 18). Proinflammatory cytokines interkeukin (IL)-15 and C-C motif chemokine ligand (CCL4) were found to predict death, adding value to CRP levels. The decision tree predicted correctly the outcome of 91% of the patients at hospital admission. The high-risk group, defined as CRP ≥ 72 mg/dL, IL-15 ≥ 5·6 fg/ml and CCL4 ≥ 6·35 fg/ml had an 88% in-hospital mortality rate, whereas the patients classified as low-risk had a mortality rate of 8% (P = < 0·001). Cytokines IL-15 and CCL4 were predictors of mortality in IE, adding prognostic value beyond that provided by CRP levels. Assessment of cytokines has potential value for clinical risk stratification and monitoring in IE patients.

Fcγ-RI, Fcγ-RII and IL-10 as predictive biomarkers for post-therapeutic cicatrization time in monocytes from cutaneous leishmaniasis patients.

Cutaneous leishmaniasis (CL) treatment is based on therapy with Glucantime® , yet, there are few laboratory methods to monitor its success. In this study, ex vivo and in vitro evaluations of peripheral blood monocytes were performed in a longitudinal study to characterize the impact of Glucantime® on overall phenotypic/functional features of these cells from CL patients to identify predictive biomarkers for post-therapeutic monitoring by flow cytometry. The ex vivo evaluation from CL patients demonstrated a modulatory profile before treatment, with a decrease in TLR-2, FcγRII, HLA-DR, CD86, IFN-γR, TNF, IL-12, NO, and an increase in FcγRIII and IL-10R. Conversely, treatment changes some of these biomarker expressions by decreasing FcγRIII and IL-10R and increasing IFN-γR, IL-12 and NO. Moreover, an in vitro analysis of these patients showed a reduced phagocytic capacity of Leishmania braziliensis and higher levels of IL-10 and TGF-ß modulating functional profile. Regardless of the compromised L. braziliensis phagocytic capacity, treatment re-established the production of IL-12, IL-10, TGF-ß and NO at the basal level. Notably, monocytes from patients with early cicatrization showed enhanced FcγRI and FcγRII expressions and reduced IL-10, which was further corroborated by a baseline fold change analysis. Finally, the logistic regression model emphasized the performance of FcγRI, FcγRII and IL-10 as robust predictive biomarkers for post-therapeutic cicatrization during cutaneous leishmaniasis.

Protective Profile Involving CD23/IgE-mediated NO Release is a Hallmark of Cutaneous Leishmaniasis Patients from the Xakriabá Indigenous Community in Minas Gerais, Brazil.

In this study, we described, for the first time, specific aspects of an anti-Leishmania immune response in a Brazilian Xakriabá indigenous community. Induction of an intracellular NO pathway, triggered by the binding of IgE to CD23 receptor in IFN-γ/IL-4 cytokines environment, was evaluated in localized cutaneous leishmaniasis (LCL) carriers and positive Montenegro skin test (MST) individuals without skin lesion (MT(+) SL(-)). Our data demonstrated that the higher frequency of CD23(+) CD14(+) monocytes and the increased serum levels of IgE observed in the LCL group were even higher in LCL carriers with late lesions (LCL≥60). Furthermore, patients with LCL presented increased NO production after Leishmania (Viannia) braziliensis stimulation and this NO profile was independent of the time of the lesion (recent LCL<60 or late LCL≥60). We also showed that the increased frequency of IFN-γ(+) and IL-4(+) CD4(+) T cells is related to the MT(+) SL(-) group. The results of biomarker signature curves demonstrated that in the MT(+) SL(-) group, the index signature was characterized by DAF-2T(+) CD14(+)/IL-4(+) CD8(+)/IFN-γ(+) CD4(+)/IL-4(+) CD4(+). On the other hand, the LCL group presented a higher index of DAF-2T(+) CD14(+)/CD23(+) CD14(+)/IL-4(+) CD8(+), associated with a lower index of IFN-γ(+) CD8(+). Considering the time of lesion, data analysis demonstrated that the main differences observed were highlighted in LCL<60 patients, with a higher index of CD23(+) CD14(+), which was also present in LCL≥60 patients. In conclusion, our data suggest that the protective immune response involving CD23-IgE-mediated NO release is a hallmark of patients with LCL. However, in MT(+) SL(-) individuals, another different leishmanicidal mechanism seems to be involved.

Cross-reactivity of anti-human cytokine monoclonal antibodies used as a tool to identify novel immunological biomarkers in domestic ruminants.

Eleven commercially available PE-labeled anti-human (IL-1-α, IL-6, IL-8, TNF-α, IL-17A, IL-5, IL-10, IL-12 and IL-13) and anti-mouse (IL-10, TNF-α) cytokine monoclonal antibodies (mAbs) were tested for cross-reactivity with cattle, goat, and sheep cytokines. Cross-reactivity was assessed by comparative analysis with the standard reactivity of the target species. Our data demonstrated that anti-human IL-1-α, IL-6, IL-8, IL-17A and IL-10 mAbs cross-react with all ruminant species tested. Anti-human IL-5 mAb showed a strong cross-reactivity with cattle and goat IL-5, while anti-human TNF-α mAb showed a selective cross-reactivity with goat TNF-α. No cross-reactivity with the ruminant cytokines was observed for anti-human IL-12 and IL-13 mAbs or for the two anti-mouse cytokine mAbs tested. The present study demonstrated the cross-reactivity of various anti-human cytokine mAbs with cattle, sheep, and goat cytokines, increasing the range of immunological biomarkers for studies in veterinary medicine.

Distinct pattern of immunophenotypic features of innate and adaptive immunity as a putative signature of clinical and laboratorial status of patients with localized cutaneous leishmaniasis.

In this study, we have analysed the phenotypic features of innate/adaptive immunity of patients with localized cutaneous leishmaniasis (LCL), categorized according to their clinical/laboratorial status, including number of lesion (L1; L2­4), days of illness duration (≤60;>60) and positivity in the Montenegro skin test (MT−;MT+). Our findings highlighted a range of phenotypic features observed in patients with LCL (↑%HLA-DR+ neutrophils; ↑CD8+ HLA-DR+/CD4+ HLA-DR+ T cell ratio; ↑HLA-DR in B lymphocytes, ↑%CD23+ neutrophils, monocytes and B cells; ↑α-Leishmania IgG and ↑serum NO2⁻ + NO3⁻). Selective changes were observed in L1 (↑%HLA-DR+ neutrophils, ↑CD8+ HLA-DR+/CD4+ HLA-DR+ T cell ratio and ↑serum NO2⁻ + NO3⁻) as compared to L2­4 (↑%CD5− B cells; ↑CD23+ B cells and ↑α-Leishmania IgG). Whilst ≤60 presented a mixed profile of innate/adaptive immunity (↓%CD28+ neutrophils and ↑%CD4+ T cells), >60 showed a well-known leishmanicidal events (↑CD8+ T cells; ↑serum NO2⁻ + NO3⁻ and ↑α-Leishmania IgG). MT+ patients showed increased putative leishmanicidal capacity (↑%HLA-DR+ neutrophils; ↑%CD23+ monocytes; ↑CD8+ HLA-DR+/CD4+ HLA-DR+ T cell ratio and ↑ serum NO2⁻ + NO3⁻). Overall, a range of immunological biomarkers illustrates the complex immunological network associated with distinct clinical/laboratorial features of LCL with applicability in clinical studies.

Distribution of hepatitis C virus in circulating blood components from blood donors.

BACKGROUND AND OBJECTIVES: Current nucleic acid tests (NAT) for blood donor screening use plasma as the test sample and, consequently, cannot detect virions bound to blood cells of infected donors. Hepatitis C virus (HCV) RNA and infectious virions have been detected in association with the cellular components of blood of patients with active liver disease; however, studies comparing HCV viral loads in whole blood and plasma have generated contradictory results. The aim of this study was to investigate the distribution of HCV in different compartments of the peripheral blood from HCV-infected blood donors, which may differ from that observed in patients with HCV-associated liver disease. MATERIALS AND METHODS: Hepatitis C virus-positive donor specimens were identified by NAT and antibody testing. HCV RNA was extracted from samples of whole blood and their corresponding components (RBC and plasma). Viral RNA was quantified by real-time qRT-PCR. RESULTS: Hepatitis C virus was present in all blood components from infected donors from which RNA could be amplified. For the majority of samples, plasma (34/46) had the highest detectable concentration of HCV RNA, and RBC (37/46) had the lowest. Specimens with negative NAT and positive antibody assays also produced qRT-PCR negative results. CONCLUSION: These results indicate that including the RBC fraction in the tested sample will not increase assay sensitivity. Although 10% of the specimens had a higher viral load in whole blood, there was no significant overall increase in sensitivity to justify changes in the specimen format. Thus, plasma specimens are well suited for blood donor screening for HCV.

Living in endemic area for infectious diseases accelerates epigenetic age.

Inflammaging is a low-grade inflammatory state generated by the aging process that can contribute to frailty and age-related diseases in the elderly. However, it can have distinct effects in the elderly living in endemic areas for infectious diseases. An increased inflammatory response may confer protection against infectious agents in these areas, although this advantage can cause accelerating epigenetic aging. In this study, we evaluated the inflammatory profile and the epigenetic age of infected and noninfected individuals from an endemic area in Brazil. The profile of cytokines, chemokines and growth factors analyzed in the sera of the two groups of individuals showed similarities, although infected individuals had a higher concentration of these mediators. A significant increase in IL-1ra, CXCL8, CCL2, CCL3 and CCL4 production was associated with leprosy infection. Notably, elderly individuals displayed distinct immune responses associated with their infection status when compared to adults suggesting an adaptive remodelling of their immune responses. Epigenetic analysis also showed that there was no difference in epigenetic age between the two groups of individuals. However, individuals from the endemic area had a significant accelerated aging when compared to individuals from São Paulo, a non-endemic area in Brazil. Moreover, the latter cohort was also epigenetically aged in relation to an Italian cohort. Our data shows that living in endemic areas for chronic infectious diseases results in remodelling of inflammaging and acceleration of epigenetic aging in individuals regardless of their infectious status. It also highlights that geographical, genetic and environmental factors influence aging and immunosenescence in their pace and profile.

Immunological/virological peripheral blood biomarkers and distinct patterns of sleeping quality in chronic hepatitis C patients.

The rational of this study we intended to investigate whether the peripheral blood immunological/virological biomarkers were associated with distinct patterns of sleeping quality in patients with chronic hepatitis C-(HCV). Distinct well-established indexes/scores were used to categorize the sleeping quality of HCV patients, including the Pittsburgh Sleep Quality Index (PSQI), Epworth Sleepiness Scale and Fatigue Severity Scores. Our findings demonstrated that HCV patients classified as 'good sleeper' displayed an enhanced frequency of circulating CD8(+) T cells, lower frequency of activated (CD69(+)) neutrophils and eosinophils but enhanced frequency of activated lymphocytes besides lower seric levels of IL-4/IL-8/IL-10 but higher levels of IL-12, besides lower HCV virus load and lower anti-HCV IgG levels. In contrast, HCV patients classified as 'poor sleeper' displayed enhanced levels of activated neutrophils and eosinophils but lower frequency of activated lymphocytes, higher seric levels of IL-6/TNF-α/IL-10 but lower levels of IL-12 besides higher HCV virus load and increased anti-HCV IgG levels. Positive correlation was further confirmed by the relationship between the leucocyte activation status, the cytokine levels, the HCV viral load and the anti-HCV IgG reactivity with the PSQI indexes. Analysis of cytokine signature curves demonstrated that lower frequency of IL-10 was observed in HCV patients classified as 'good sleepers', whereas enhanced frequency of IL-6 was found HCV patients classified as 'poor sleepers'. In conclusion, our data suggest that immunological biomarkers (leucocytes activation status and seric cytokines levels) are likely to be associated with sleeping quality patterns in HCV patients, suggesting their putative use for clinical monitoring purposes.

A shift towards a T cell cytokine deficiency along with an anti-inflammatory/regulatory microenvironment may enable the synthesis of anti-FVIII inhibitors in haemophilia A patients.

Despite the clinical relevance of anti-factor VIII (FVIII) antibodies (anti-FVIII inhibitors) impairing haemostatic activity of haemophilia A (HA) patients, the immunological mechanisms underlying their production are unknown. Aiming to understand more clearly the immune response in patients with [HAα-FVIII(+)] and without [HAα-FVIII(-)] anti-FVIII inhibitors, we have characterized the cytokine pattern of peripheral blood leucocytes, using an in vitro stimulation of whole blood samples with plasma-derived (pFVIII) or recombinant FVIII (rFVIII). The results highlighted decreased levels of tumour necrosis factor (TNF)-α(+) neutrophils with higher interleukin (IL)-5/TNF-α ratio in HAα-FVIII(+). All HA samples displayed decreased levels of IL-10(+) monocytes when compared to the blood donor (BD) samples. HAα-FVIII(+) showed lower levels of TNF-α(+) monocytes and increased IL-10/TNF-α ratio. Analysis of adaptive immunity revealed increased levels of interferon (IFN)-γ(+) , TNF-α(+) and IL-4(+) T-cells, from both CD4(+) and CD8(+) T cells, in HAα-FVIII(-) when compared to BD. Moreover, increased frequency of IL-10(+) B cells and higher levels of α-FVIII IgG1 were observed in HAα-FVIII(-). Basal levels of cytokine(+) B-cells, similar to BD, and higher levels of α-FVIII IgG4 are major features in HAα-FVIII(+). The global cytokine profile demonstrated a major anti-inflammatory/regulatory pattern in HAα-FVIII(+), confirmed by the in vitro stimuli with pFVIII or rFVIII. The polarized anti-inflammatory/regulatory immune response in HAα-FVIII(+) and the mixed pattern with a bias towards an inflammatory cytokine profile, modulated by IL-4 in HAα-FVIII(-), may be the key element to drive the development of distinct subclasses of anti-FVIII antibodies. These finding have implications for the design of safe and effective therapeutic protocols to control inhibitors synthesis in HA patients.

Ageing down-modulates liver inflammatory immune responses to schistosome infection in mice.

Ageing is associated with several alterations in the immune system. Our aim in this study was to compare the development of immunity to Schistosoma mansoni infection in young versus aged C57Bl/6 mice using the liver as the main organ to evaluate pathological alterations and immune responses. In the acute phase, young mice had large liver granulomas with fibrosis and inflammatory cells. Chronic phase in young animals was associated with immunomodulation of granulomas that became reduced in size and cellular infiltrate. On the other hand, aged animals presented granulomas of smaller sizes already in the acute phase. Chronic infection in these mice was followed by no alteration in any of the inflammatory parameters in the liver. In concert with this finding, there was an increase in activated CD4+ T, CD19+ B and NK liver cells in young mice after infection whereas old mice had already higher frequencies of activated B, NK and CD4+ T liver cells and infection does not change these frequencies. After infection, liver production of inflammatory and regulatory cytokines such as IFN-gamma, IL-4 and IL-10 increased in young but not in old mice that had high levels of IL-4 and IL-10 regardless of their infection status. Our data suggest that the unspecific activation status of the immune system in aged mice impairs inflammatory as well as regulatory immune responses to S. mansoni infection in the liver, where major pathological alterations and immunity are at stage. This poor immune reactivity may have a beneficial impact on disease development.

CD4 and CD8 distribution profile in individuals infected by Schistosoma mansoni.

RATIONALE: Patients with chronic Schistosoma mansoni infection show lower anti-soluble egg antigen (SEA) proliferation responses and higher responses to soluble worm antigen preparation (SWAP). OBJECTIVE: To compare the activation status and proliferation response of peripheral blood mononuclear cells (PBMC) of infected (XTO) and egg-negative individuals (NI) living in the same endemic area. METHODS: XTO (n = 51) and NI individuals from the same geographical area (n = 37) and healthy blood donors (n = 22) were evaluated before and after stimulation with SEA and SWAP. The expression of activation markers (CD4(+) HLADR(+), CD8(high+)HLA-DR(+) and CD8(+) CD28(+)) and proliferation assay was assessed by flow cytometry. FINDINGS: PBMC from infected patients showed lower frequency of CD4(+) but no change in CD8(+) T cells when compared with the healthy donor group. The ratio CD4(+)/CD8(+) was 1.3, 0.6 and 0.5 in healthy donors, infected and non-infected individuals, respectively. The HLA-DR(+) expression on CD8(+) was higher in PBMC from infected and non-infected individuals than from healthy donors, but similar in both total lymphocytes and CD4(+) populations. No intergroup proliferation response differences were observed in CD4(+) and CD8(+) PBMC unstimulated and stimulated with SEA and SWAP. The SEA but not SWAP-stimulated cells showed a decrease in the expression of phosphorylated extracellular signal-regulated kinase (ERK1/2). CONCLUSIONS: XTO and NI individuals living in the same area presented a smaller per cent of CD4(+) and a higher per cent of CD8(+) cells. The activation by either CD8(high+)HLA-DR(+) or CD8(high+)HLA-DR(+)/CD8(+) was enhanced and decreased in XTO and NI by CD8(+) CD28(+) and CD8(+) CD28(+)/CD8(+) when compared with healthy donor. ERK phosphorylation was attenuated in XTO and NI individuals when stimulated with SEA but not SWAP.

Short-term administration of ethanol in mice deviates antigen presentation activity towards B cells.

Alcohol has a variety of short- and long-term effects on cell-mediated and humoral immune response. Herein, we have characterized the impact of high-dose EtOH administration on phenotypic and functional features of murine APC subsets, including dendritic cell (DC), macrophages and B cells. Impaired cytokine synthesis and Leishmania-phagocytosis was observed in peritoneal macrophages following EtOH administration. Moreover, EtOH exposure led to decreased levels of splenic myeloid DC and increased percentage of macrophages with no changes in splenic lymphoid DC and B cells. Adverse effects of short-term EtOH administration also resulted in impaired OVA-endocytosis by DC and macrophages. In contrast, EtOH consumption upregulates OVA-internalization by B cells. These changes on APC hierarchy may play a role shifting the fate of the immune response after EtOH ingestion. In addition to an overall downregulation of Toll-like receptor-TLR-4 expression by splenic APC, a downregulation of TLR-2 expression in macrophages was observed. Moreover, EtOH exposure altered the expression of co-signalling molecules on splenic APC, downregulating CD40 on macrophages and upregulating CD80 on B cells, with no impact on DC subsets. The net result of changes in TLR-mediated and co-stimulatory signals may determine the altered immunological status induced by acute consumption of alcohol. A direct impact of high-dose EtOH administration in the activation status of splenic CD4(+) T cells was observed. Together, our results demonstrated that short-term high-dose EtOH administration has differential impact on APC populations, downregulating splenic macrophages and DC activity but up-regulating B lymphocyte function as APC, and ultimately yielding a micro-environment that led to increased activation of CD4(+) T cells.

Effect of chemotherapy with praziquantel on the production of cytokines and morbidity associated with schistosomiasis mansoni.

The objective of the present study was to test the hypothesis that treatment of schistosomiasis mansoni with praziquantel can alter significantly the immune response of patients and generate a reversal of the level of fibrosis. Peripheral blood mononuclear cell (PBMC) samples were collected from, and abdominal ultrasound examinations conducted on, volunteers infected with Schistosoma mansoni and living in an area where the disease is endemic, both prior to and one year after treatment with praziquantel. Subjects were classified into groups according to the level of pathology (i.e., absent, incipient, moderate, or severe fibrosis). PBMCs were stimulated with schistosome soluble egg antigens (SEA), and the levels of production of the cytokines gamma interferon (IFN-gamma), tumor necrosis factor alpha, transforming growth factor beta, and interleukin-4 (IL-4), IL-10, and IL-13 were determined. The chemotherapy was effective in reducing morbidity, particularly for individuals presenting with severe fibrosis. When levels of cytokine production in posttreatment PBMC cultures stimulated by SEA were categorized as low or high, significant differences in the distribution of IL-13 levels between groups presenting with or not presenting with fibrosis were established. Comparison of pre- and posttreatment SEA-induced cytokine levels in individuals who had experienced no change in the grade of fibrosis following chemotherapy revealed that the level of IFN-gamma decreased in subjects with fibrosis whereas that of IL-10 decreased in individuals with and without fibrosis. The data suggest that chemotherapy is effective in reducing the morbidity of the disease and that the level of IL-13 may be a useful indicator of the persistence of fibrosis following treatment.

Strategy to assess the overall cytokine profile of circulating leukocytes and its association with distinct clinical forms of human Chagas disease.

Herein we have employed an alternative strategy to assess the cytokine patterns of circulating leukocytes and correlate dominant cytokine profiles with indeterminate-IND and cardiac-CARD clinical forms of Chagas disease. We have first calculated median percentages of cytokine-positive leukocytes of our study sample to establish, for each cytokine-positive cell population, the cut-off edge that would segregate 'low' and 'high' cytokine producers to build colour diagrams and draw a panoramic cytokine chart. Using this approach we demonstrated that most IND individuals presented a dominant regulatory cytokine profile, whereas CARD individuals displayed a dominant inflammatory cytokine pattern. In addition, radar chart analysis confirmed the dichotomic cytokine balance between IND and CARD groups and further allowed the identification of the relative contribution of each cell population for the global cytokine pattern. Data analysis demonstrated that CD4+ T cells were the major cell population defining the regulatory profile in IND, whereas monocytes and CD4+ T cells determined the inflammatory cytokine pattern in CARD individuals. Interestingly, in vitro stimulation with trypomastigote Trypanosoma cruzi antigen was able to invert the cytokine balances in IND and CARD groups. Upon antigenic stimulation, changes in the frequencies of IL-10-producing CD4+ T cells and monocytes drove IND individuals towards an inflammatory pattern and CARD towards a regulatory cytokine profile. A similar inversion could be found after in vivo treatment of IND and CARD individuals with benzonidazole. Altogether, these findings shed some light into the complex cytokine network underlying the immunopathogenesis of Chagas disease and provide putative immunological biomarkers of disease severity and therapeutic response.

Trypanosoma cruzi: immunoglobulin isotype profiles during the acute phase of canine experimental infection with metacyclic or blood trypomastigotes.

A detailed investigation has been carried out about the serological profiles of groups of dogs experimentally infected with metacyclic (MT) or blood (BT) trypomastigotes of Berenice-78 Trypanosoma cruzi strain. Peripheral blood was collected from infected dogs and uninfected controls, weekly during 35 days following the acute phase of infection, and immunoglobulin profiles were determined by ELISA. Dogs infected with BT exhibited unaltered levels of IgG2, increases in IgM, IgE, IgA, IgG and IgG1. In contrast, dogs infected with MT presented unaltered levels of IgE and IgG1 and an increase in IgM, IgA, IgG and IgG2 levels. Compared with the MT group, animals infected with BT showed significant increases in IgM on days 7, 14 and 28, in IgA on days 7, 14 and 21, in IgE on days 7 and 14, in IgG on days 14 and 28, and in IgG1 on days 7, 14 and 21. Parasitemia levels of the infected animals were measured over the same time period. No correlations were found between the immunoglobulin profiles and the parasitemia levels. The results demonstrated that the inoculum source (BT or MT) influence the immunoglobulin isotype profile that may drive distinct outcome of acute canine Chagas disease.

Plasma biomarkers profile of female dogs with mammary carcinoma and its association with clinical and pathological features.

The immunological biomarkers profiles were evaluated using Luminex as putative measures to monitor canine mammary carcinomas (MCs). Forty female dogs were categorized into benign mixed tumour (MC-BMT = 28) and mammary carcinoma (MC=12). The ascendant biomarker signatures were used to compare the groups. For example, a higher frequency of MC-BMT animals producing IL-6, CXCL-8 and CXCL-10 was observed, whereas for the MC group IL-2 and CXCL-8 were detected. MC-BMT animals without metastasis had an increase in the levels of IL-2, CXCL-8, CXCL-10, IL-6, TNF-α, IL-15 and a decrease in IL-10 and CXCL-8. MC-BMT animals with metastasis showed only an increase in CXCL-10 and a decrease in IL-18. After comparing the ascendant signatures following the presence of metastasis in both groups, a higher frequency of dogs exhibiting IL-10 production was observed. Pearson correlation (P = 0.0273) and receiver operating characteristic (ROC) curve analysis revealed that this pattern was associated with worse outcome and lower survival rates in MC animals.

Etiological treatment of Chagas disease patients with benznidazole lead to a sustained pro-inflammatory profile counterbalanced by modulatory events.

In the present study, we characterized the phagocytic capacity, cytokine profile along with the FCγ-R and TLR expression in leukocytes from Chagas disease patients (indeterminate-IND and cardiac-CARD) before and one-year after Bz-treatment (INDT and CARDT). A down-regulation of IL-17, IFN-γ and IL-10 synthesis by neutrophils was observed in CARDT. The Bz-treatment did not impact on the expression of phagocytosis-related surface molecules or monocyte-derived cytokine profile in INDT. Although CARDT showed unaltered monocyte-phagocytic capacity, up-regulated expression of Fcγ-RI/III and TLR-4 may be related to their ability to produce IL-10 and TGF-ß. Down-regulation of lymphocyte-derived cytokine was observed in INDT whereas up-regulated cytokine profile was observed for lymphocytes in CARDT. Analysis of cytokine network revealed that IND displayed a multifaceted cytokine response characterized by strong connecting axes involving pro-inflammatory/regulatory phagocytes and lymphocytes. On the other hand, CARD presented a modest cytokine network. The Bz-treatment leads to distinct cytokine network: decreasing the links in INDT, with a pivotal role of IL-10(+) monocytes and expanding the connections in CARDT. Our findings highlighted that the Bz-treatment contributes to an overall immunomodulation in INDT and induces a broad change of immunological response in CARDT, eliciting an intricate phenotypic/functional network compatible with beneficial and protective immunological events.

Impaired phagocytic capacity driven by downregulation of major phagocytosis-related cell surface molecules elicits an overall modulatory cytokine profile in neutrophils and monocytes from the indeterminate clinical form of Chagas disease.

The distinct ability of phagocytes to present antigens, produce cytokines and provide co-stimulatory signals may contribute to the severity of the outcome of Chagas disease. In this paper, we evaluate the phenotypic features of phagocytes along with the cytokine signature of circulating T-cells from Chagas disease patients with indeterminate (IND) and cardiac (CARD) clinical forms of the disease. Our data demonstrated that neutrophils from IND patients displayed an impaired ability to produce cytokines. A lower Trypanosoma cruzi phagocytic index and higher nitric oxide levels were characteristics of monocytes from IND. The impaired phagocytic capacity did not reflect on the levels of anti-T. cruzi IgG, but was detectable in the downregulation of Fc-γR, TLR and CR1 molecules. The monocyte-derived cytokine signature demonstrated that a down-regulated synthesis of IL-12 and a modulatory state were evidenced by a positive correlation between IL-12 and IL-10 with a lower synthesis of TNF-α. The down-regulation of MHC-II and CD86 in monocytes supports the occurrence of particularities in the APC-activation-arm in IND, and may be involved in the T-cell pro-inflammatory pattern counterbalanced by a potent IL-10 response. Our findings support the hypothesis that differential phenotypic features of monocytes from IND may be committed to the induction of a distinct immune response related to low morbidity in chronic Chagas disease.

Foxp3+CD25(high) CD4+ regulatory T cells from indeterminate patients with Chagas disease can suppress the effector cells and cytokines and reveal altered correlations with disease severity.

Immunoregulatory mechanisms are important to control the intense immune activity induced in Chagas disease. We evaluated the phenotypic profile and the mechanisms by which Treg cells function in patients with the indeterminate (IND) and cardiac (CARD) clinical forms of Chagas disease. The frequency of Foxp3(+)CD25(high) CD4(+)-T cells is augmented and correlated with the maintenance of a better cardiac function in IND. Treg cells from IND present suppressive activity, although the mechanism is not IL-10 or CTLA-4 dependent and are able to produce augmented levels of IL-17, IL-10 and granzyme B being its frequency correlated with percentage of Annexin V(+) CD4(+)-cells. In contrast, CARD presents higher frequency of IL-6(+), IFN-gamma(+), TNF-alpha(+) and CTLA-4(+) Treg-cells than IND. Thus, our data suggest that Treg cells have an important role in controlling the exacerbated immune response and morbidity in Trypanosoma cruzi infection, probably modulating the cytokine environment and/or killing effector cells.

Humoral and cellular immune responses in dogs with inapparent natural Leishmania infantum infection.

Molecular analysis, serology and immunophenotyping for T lymphocytes and their subsets, B lymphocytes and monocytes were performed on dogs naturally infected with Leishmania infantum. Animals were categorised as asymptomatic dogs I (AD-I), with negative serology and positive molecular results, and asymptomatic dogs II (AD-II), with positive serology and positive molecular results, and these were compared to symptomatic dogs (SD) and control dogs (CD). AD-I exhibited immunophenotypic features similar to those of CD, including isotype profiles and concentrations of monocytes. Similar biomarkers were found in AD-II and SD, such as, higher levels of immunoglobulins IgG, IgG2, IgM and IgA and higher concentrations of eosinophils. High frequencies of T lymphocytes and CD4(+) T cells were observed in both AD-I and AD-II compared to SD, whereas CD8(+) T cells were higher only in AD-II compared with SD. Analysis of B lymphocytes revealed an increased frequency of this cell type only in AD-II animals compared with SD. Asymptomatic dogs appear to have a dichotomous infection spectrum that can influence the humoral and cellular immunological status during canine visceral leishmaniasis.