Analysis of iron superoxide dismutase-encoding DNA vaccine on the evolution of the Leishmania amazonensis experimental infection.

The present work aimed to evaluate the immunogenicity of Leishmania amazonensis iron superoxide dismutase (SOD)-encoding DNA experimental vaccine and the protective properties of this DNA vaccine during infection. The SOD gene was subcloned into the pVAX1 plasmid, and it was used to immunize BALB/c mice. Twenty-one days after the last immunization, mice were sacrificed (immunogenicity studies) or subcutaneously challenged with L. amazonensis (studies of protection), and alterations in cellular and humoral immune responses were evaluated, as well as the course of infection. Mice only immunized with pVAX1-SOD presented increased frequencies of CD4(+) IFN-γ(+), CD8(+)IFN-γ(+) and CD8(+)IL-4(+) lymphocytes; moreover, high levels of IgG2a were detected. After challenge, mice that were immunized with pVAX1-SOD had increased frequencies of the CD4(+)IL-4(+), CD8(+)IFN-γ(+) and CD8(+)IL-4(+) T lymphocytes. In addition, the lymph node cells produced high amounts of IFN-γ and IL-4 cytokines. Increased IgG2a was also detected. The pattern of immunity induced by pVAX1-SOD partially protected the BALB/c mice from a challenge with L. amazonensis, as the animals presented reduced parasitism and lesion size when compared to controls. Taken together, these results indicate that leishmanial SOD modulates the lymphocyte response, and that the elevation in IFN-γ possibly accounted for the decreased skin parasitism observed in immunized animals.

Imbalanced expression of functional surface molecules in regulatory and effector T cells in systemic lupus erythematosus.

Regulatory T (TREG) cells play an important role in maintaining immune tolerance and avoiding autoimmunity. We analyzed the expression of membrane molecules in TREG and effector T cells in systemic lupus erythematosus (SLE). TREG and effector T cells were analyzed for the expression of CTLA-4, PD1, CD28, CD95, GITR, HLA-DR, OX40, CD40L, and CD45RO in 26 patients with active disease, 31 with inactive disease, and 26 healthy controls. TREG cells were defined as CD25+/high CD127 Ø/low FoxP3+, and effector T cells were defined as CD25+CD127+FoxP3 Ø. The ratio of TREG to effector T cells expressing GITR, PD1, HLA-DR, OX40, CD40L, and CD45RO was determined in the three groups. The frequency of TREG cells was similar in patients with SLE and controls. However, SLE patients had a decreased frequency of CTLA-4+TREG and CD28+TREG cells and an increased frequency of CD40L+TREG cells. There was a decrease in the TREG/effector-T ratio for GITR+, HLA-DR+, OX40+, and CD45RO+ cells, and an increased ratio of TREG/effector-T CD40L+ cells in patients with SLE. In addition, CD40L+TREG cell frequency correlated with the SLE disease activity index (P=0.0163). In conclusion, our findings showed several abnormalities in the expression of functionally critical surface molecules in TREG and effector T cells in SLE that may be relevant to the pathogenesis of this disease.

Imbalanced expression of functional surface molecules in regulatory and effector T cells in systemic lupus erythematosus

Regulatory T (TREG) cells play an important role in maintaining immune tolerance and avoiding autoimmunity. We analyzed the expression of membrane molecules in TREG and effector T cells in systemic lupus erythematosus (SLE). TREG and effector T cells were analyzed for the expression of CTLA-4, PD1, CD28, CD95, GITR, HLA-DR, OX40, CD40L, and CD45RO in 26 patients with active disease, 31 with inactive disease, and 26 healthy controls. TREG cells were defined as CD25+/highCD127Ø/lowFoxP3+, and effector T cells were defined as CD25+CD127+FoxP3Ø. The ratio of TREG to effector T cells expressing GITR, PD1, HLA-DR, OX40, CD40L, and CD45RO was determined in the three groups. The frequency of TREG cells was similar in patients with SLE and controls. However, SLE patients had a decreased frequency of CTLA-4+TREG and CD28+TREG cells and an increased frequency of CD40L+TREG cells. There was a decrease in the TREG/effector-T ratio for GITR+, HLA-DR+, OX40+, and CD45RO+ cells, and an increased ratio of TREG/effector-T CD40L+ cells in patients with SLE. In addition, CD40L+TREG cell frequency correlated with the SLE disease activity index (P=0.0163). In conclusion, our findings showed several abnormalities in the expression of functionally critical surface molecules in TREG and effector T cells in SLE that may be relevant to the pathogenesis of this disease.

Peripheral blood telomere content is greater in patients with endometriosis than in controls.

UNLABELLED: The etiology of endometriosis remains poorly understood but circulating stem cells may contribute. Telomeres shorten with cell divisions and age. Stem cells attempt to compensate for telomere attrition through the action of telomerase. Since circulating stem cells may contribute to endometriosis, we compared telomere content in lymphocytes of patients with and without endometriosis. METHODS: Observational study comparing peripheral lymphocytes telomere content, measured by quantitative polymerase chain reaction, in patients with (n = 86) and without endometriosis (n = 21). FINDINGS: Patients with endometriosis had longer telomeres than that of matched, endometriosis-free controls (telomere to single copy gene ratio [T/S ratio] of 1.62 vs 1.34, respectively, P = .00002). Patients with endometriosis were 8.1-fold more likely to have long telomeres. (odds ratio = 8.1, 95% confidence interval: 1.28-51.57, P = .0264). INTERPRETATION: Longer telomeres could be consistent with a stem cell origin of endometriosis.

HCV viremia drives an increment of CD86 expression by myeloid dendritic cells.

The host immune response, including innate and adaptive immunity, plays a critical role in determining the outcome of viral infection. Nevertheless, little is known about the exact reasons for the failure of the host immune system in controlling hepatitis C virus (HCV) infection. Impairment of dendritic cells (DCs) function is probably one of the mechanisms responsible for immune evasion of HCV. In this study, the frequency and phenotype of DCs subsets were analyzed in three groups: HCV-infected individuals who developed viral persistence (1), HCV-infected individuals who spontaneously cleared the virus (2) and HCV-seronegative uninfected subjects (3). The results showed that the frequency of DCs subsets was not statistically significant between groups. Plasmacytoid DCs circulating exhibited an immature phenotype characterized by low expression of CD86. On the other hand, CD86 expression in myeloid DCs was significantly higher in chronic infected individuals compared to healthy controls (P=0.037). A positive correlation was observed between CD86(+) myeloid DC (mDC) and HCV viral load (r=0.4121, P=0.0263). These results suggest that HCV did not have an inhibitory effect on mDC maturation and the HCV viremia drives the increase of CD86 expression in mDC. The regulation of DCs maturation and migration lies at the level of intracellular signaling. HCV can activate or block intracellular signaling pathways and alter DC function. In conclusion, the present study suggests that imbalance of DC maturation by the virus represents a mechanism of evasion of the immune system despite the fact that HCV viremia appears to exert a "stimulatory" effect on cell-surface immune phenotype.

Chemokines in the pathogenesis of endometriosis and infertility.

Endometriosis is a chronic benign disease that affects women of reproductive age causing abdominal pain and infertility. Its pathogenesis remains obscure despite all the research conducted over the past 100 years. However, there is a consensus among the specialists that the basis of its pathophysiology would be multifactorial. Many publications have demonstrated that chemokines are somehow associated with the development of endometriosis and infertility. In this study, we reviewed all PubMed literature using MeSH terms "chemokines" and "endometriosis" as well as "chemokines" and "female infertility" to establish what we know and what we do not yet know about this relationship.

Investigation of human parvovirus B19 occurrence and genetic variability in different leukaemia entities.

Human parvovirus B19V (B19V) has been associated with various haematological disorders, but data on its prevalence in leukaemia are scarce. In this cross-sectional study, we investigated patients in Sao Paulo, Brazil with leukaemia to determine the molecular frequency of B19 variants and characterize the viral genetic variability by partial and complete sequencing of the coding of non-structural protein 1 (NS1)/viral capsid proteins 1 and 2 (VP1/VP2). The presence of B19V infections was investigated by PCR amplification of the viral NS1 gene fragment and confirmed by sequencing analysis. The NS1/VP1/VP2 and partially larger gene fragments of the NS1-positive samples were determined by overlapping nested PCR and direct sequencing results. The B19V NS1 was detected in 40 (16%) of 249 bone marrow samples including 12/78 (15.4%) acute lymphoblastic leukaemia, 25/155 (16.1%) acute myeloid leukaemia and 3/16 (18.7%) chronic myeloid leukaemia samples. Of the 40 participants, 25 (62.5%) were infected with genotype 1a and 15 (37.5%) with genotype 3b. The phylogenetic analysis of other regions revealed that 12/40 (30%) of the patients with leukaemia were co-infected with genotypes 1a and 3b. In addition, a new B19V intergenotypic recombinant (1a/3b) and an NS1 non-recombinant genotype 1a were detected in one patient. Our findings demonstrated a relatively high prevalence of B19V monoinfections and dual infections and provide, for the first time, evidence of inter-genotypic recombination in adults with leukaemia that may contribute to the genetic diversity of B19V and may also be a source of new emerging viral strains with future implications for diagnosis, therapy and efficient vaccine development.

Lower numbers of circulating Natural Killer T (NK T) cells in individuals with human T lymphotropic virus type 1 (HTLV-1) associated neurological disease.

Human T lymphotropic virus type 1 (HTLV-1) infects 10-20 million people worldwide. The majority of infected individuals are asymptomatic; however, approximately 3% develop the debilitating neurological disease HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). There is also currently no cure, vaccine or effective therapy for HTLV-1 infection, and the mechanisms for progression to HAM/TSP remain unclear. NK T cells are an immunoregulatory T cell subset whose frequencies and effector functions are associated critically with immunity against infectious diseases. We hypothesized that NK T cells are associated with HAM/TSP progression. We measured NK T cell frequencies and absolute numbers in individuals with HAM/TSP infection from two cohorts on two continents: São Paulo, Brazil and San Francisco, CA, USA, and found significantly lower levels when compared with healthy subjects and/or asymptomatic carriers. Also, the circulating NK T cell compartment in HAM/TSP subjects is comprised of significantly more CD4(+) and fewer CD8(+) cells than healthy controls. These findings suggest that lower numbers of circulating NK T cells and enrichment of the CD4(+) NK T subset are associated with HTLV-1 disease progression.

Autoimmune diseases in the TH17 era: [review]

A new subtype of CD4+ T lymphocytes characterized by the production of interleukin 17, i.e., TH17 cells, has been recently described. This novel T cell subset is distinct from type 1 and type 2 T helper cells. The major feature of this subpopulation is to generate significant amounts of pro-inflammatory cytokines, therefore appearing to be critically involved in protection against infection caused by extracellular microorganisms, and in the pathogenesis of autoimmune diseases and allergy. The dynamic balance among subsets of T cells is important for the modulation of several steps of the immune response. Disturbances in this balance may cause a shift from normal immunologic physiology to the development of immune-mediated disorders. In autoimmune diseases, the fine balance between the proportion and degree of activation of the various T lymphocyte subsets can contribute to persistent undesirable inflammatory responses and tissue replacement by fibrosis. This review highlights the importance of TH17 cells in this process by providing an update on the biology of these cells and focusing on their biology and differentiation processes in the context of immune-mediated chronic inflammatory diseases.

Autoimmune diseases in the TH17 era.

A new subtype of CD4+ T lymphocytes characterized by the production of interleukin 17, i.e., TH17 cells, has been recently described. This novel T cell subset is distinct from type 1 and type 2 T helper cells. The major feature of this subpopulation is to generate significant amounts of pro-inflammatory cytokines, therefore appearing to be critically involved in protection against infection caused by extracellular microorganisms, and in the pathogenesis of autoimmune diseases and allergy. The dynamic balance among subsets of T cells is important for the modulation of several steps of the immune response. Disturbances in this balance may cause a shift from normal immunologic physiology to the development of immune-mediated disorders. In autoimmune diseases, the fine balance between the proportion and degree of activation of the various T lymphocyte subsets can contribute to persistent undesirable inflammatory responses and tissue replacement by fibrosis. This review highlights the importance of TH17 cells in this process by providing an update on the biology of these cells and focusing on their biology and differentiation processes in the context of immune-mediated chronic inflammatory diseases.

Imbalance of naive and memory T lymphocytes with sustained high cellular activation during the first year of life from uninfected children born to HIV-1-infected mothers on HAART.

The immune consequences of in utero HIV exposure to uninfected children whose mothers were submitted to highly active antiretroviral therapy (HAART) during gestation are not well defined. We evaluated 45 HIV-exposed uninfected (ENI) neonates and 45 healthy unexposed control (CT) neonates. All HIV-infected mothers received HAART during pregnancy, and the viral load at delivery was <50 copies/mL for 56.8%. Twenty-three ENI neonates were further evaluated after 12 months and compared to 23 unexposed healthy age-matched infants. Immunophenotyping was performed by flow cytometry in cord and peripheral blood. Cord blood lymphocyte numbers did not differ between groups. However, ENI neonates had a lower percentage of naive T cells than CT neonates (CD4+, 76.6 vs 83.1%, P < 0.001; CD8+, 70.9 vs 79.6%, P = 0.003) and higher percentages of central memory T cells than CT neonates (CD4+, 13.9 vs 8.7%, P < 0.001; CD8+, 8.6 vs 4.8%, P = 0.001). CD38 mean fluorescence intensity of T cells was higher in ENI neonates (CD4+, 62.2 vs 52.1, P = 0.007; CD8+, 47.7 vs 35.3, P < 0.001). At 12 months, ENI infants still had higher mean fluorescence intensity of CD38 on T cells (CD4+, 34.2 vs 23.3, P < 0.001; CD8+, 26.8 vs 19.4, P = 0.035). Despite effective maternal virologic control at delivery, HIV-exposed uninfected children were born with lower levels of naive T cells. Immune activation was present at birth and remained until at least 12 months of age, suggesting that in utero exposure to HIV causes subtle immune abnormalities.

Imbalance of naive and memory T lymphocytes with sustained high cellular activation during the first year of life from uninfected children born to HIV-1-infected mothers on HAART

The immune consequences of in utero HIV exposure to uninfected children whose mothers were submitted to highly active antiretroviral therapy (HAART) during gestation are not well defined. We evaluated 45 HIV-exposed uninfected (ENI) neonates and 45 healthy unexposed control (CT) neonates. All HIV-infected mothers received HAART during pregnancy, and the viral load at delivery was <50 copies/mL for 56.8 percent. Twenty-three ENI neonates were further evaluated after 12 months and compared to 23 unexposed healthy age-matched infants. Immunophenotyping was performed by flow cytometry in cord and peripheral blood. Cord blood lymphocyte numbers did not differ between groups. However, ENI neonates had a lower percentage of naive T cells than CT neonates (CD4+, 76.6 vs 83.1 percent, P < 0.001; CD8+, 70.9 vs 79.6 percent, P = 0.003) and higher percentages of central memory T cells than CT neonates (CD4+, 13.9 vs 8.7 percent, P < 0.001; CD8+, 8.6 vs 4.8 percent, P = 0.001). CD38 mean fluorescence intensity of T cells was higher in ENI neonates (CD4+, 62.2 vs 52.1, P = 0.007; CD8+, 47.7 vs 35.3, P < 0.001). At 12 months, ENI infants still had higher mean fluorescence intensity of CD38 on T cells (CD4+, 34.2 vs 23.3, P < 0.001; CD8+, 26.8 vs 19.4, P = 0.035). Despite effective maternal virologic control at delivery, HIV-exposed uninfected children were born with lower levels of naive T cells. Immune activation was present at birth and remained until at least 12 months of age, suggesting that in utero exposure to HIV causes subtle immune abnormalities.

Antiretroviral drug therapy alters the profile of human immunodeficiency virus type 1-specific T-cell responses and shifts the immunodominant cytotoxic T-lymphocyte response from Gag to Pol.

Antiretroviral drug therapy and cytotoxic T lymphocytes (CTL) both exert selective pressures on human immunodeficiency virus type 1, which influence viral evolution. Compared to chronically infected, antiretroviral-untreated patients, most chronically infected, treated patients with detectable viremia lack a cellular immune response against the Gag 77-85(SL9) epitope but show a new immunodominant response against an epitope in protease PR 76-84. Hence, mutations induced by antiretroviral therapy likely alter the profile of epitopes presented to T cells and thus the direction of the response. The consequences of dual pressures from treatment and CTL need to be considered in monitoring of drug therapy.

Interferon-gamma and tumour necrosis factor-alpha production by CD4+ T and CD8+ T lymphocytes in AIDS patients with tuberculosis.

Tuberculosis (TB) is usually more severe in HIV-infected patients, and the immune derangement found in co-infected patients may differ from that in each isolated disease. Following mitogen stimulation of peripheral blood mononuclear cells (PBMC), interferon (IFN)-gamma and tumour necrosis factor (TNF)-alpha production was evaluated in T cells by flow cytometry, and in culture supernatants by enzyme-linked immunosorbent assay (ELISA) in 33 individuals: 11 AIDS patients with tuberculosis, six asymptomatic HIV-1-infected patients, eight patients with tuberculosis and eight healthy controls. The proportion of CD4+ T lymphocytes expressing IFN-gamma did not differ between the groups, whereas a trend towards increased proportions of TNF-alpha-expression in CD4+ T cells was observed in the TB compared to the HIV group, while intermediate values were observed in co-infected patients. Detection of IFN-gamma and TNF-alpha in CD8+ T lymphocytes was higher in TB than in HIV individuals. Co-infected patients presented intermediate values for IFN-gamma, while TNF-alpha detection was similar to that in HIV mono-infection. In conclusion, the proportion of T cells expressing IFN-gamma was relatively preserved in co-infected patients compared to TB patients, while the percentage of T cells expressing TNF-alpha was decreased, mainly in CD8+ T lymphocytes. However, the marked reduction in T lymphocyte numbers in co-infected patients led to a striking reduction of both cytokines in PBMC supernatants, a finding that is consistent with the impaired response to Mycobacterium tuberculosis.

Leptospira interrogans activation of peripheral blood monocyte glycolipoprotein demonstrated in whole blood by the release of IL-6.

Glycolipoprotein (GLP) from pathogenic serovars of Leptospira has been implicated in the pathogenesis of leptospirosis by its presence in tissues of experimental animals with leptospirosis, the inhibition of the Na,K-ATPase pump activity, and induced production of cytokines. The aims of the present study were to investigate the induction of IL-6 by GLP in peripheral blood mononuclear cells (PBMC) and to demonstrate monocyte stimulation at the cellular level in whole blood from healthy volunteers. PBMC were stimulated with increasing concentrations (5 to 2500 ng/ml) of GLP extracted from the pathogenic L. interrogans serovar Copenhageni, lipopolysaccharide (positive control) or medium (negative control), and supernatants were collected after 6, 20/24, and 48 h, and kept at -80 degrees C until use. Whole blood was diluted 1:1 in RPMI medium and cultivated for 6 h, with medium, GLP and lipopolysaccharide as described above. Monensin was added after the first hour of culture. Supernatant cytokine levels from PBMC were measured by ELISA and intracellular IL-6 was detected in monocytes in whole blood cultures by flow-cytometry. Monocytes were identified in whole blood on the basis of forward versus side scatter parameters and positive reactions with CD45 and CD14 antibodies. GLP ( > or = 50 ng/ml)-induced IL-6 levels in supernatants were detected after 6-h incubation, reaching a peak after 20/24 h. The percentage of monocytes staining for IL-6 increased with increasing GLP concentration. Thus, our findings show a GLP-induced cellular activation by demonstrating the ability of GLP to induce IL-6 and the occurrence of monocyte activation in whole blood at the cellular level.

Leptospira interrogans activation of peripheral blood monocyte glycolipoprotein demonstrated in whole blood by the release of IL-6

Glycolipoprotein (GLP) from pathogenic serovars of Leptospira has been implicated in the pathogenesis of leptospirosis by its presence in tissues of experimental animals with leptospirosis, the inhibition of the Na,K-ATPase pump activity, and induced production of cytokines. The aims of the present study were to investigate the induction of IL-6 by GLP in peripheral blood mononuclear cells (PBMC) and to demonstrate monocyte stimulation at the cellular level in whole blood from healthy volunteers. PBMC were stimulated with increasing concentrations (5 to 2500 ng/ml) of GLP extracted from the pathogenic L. interrogans serovar Copenhageni, lipopolysaccharide (positive control) or medium (negative control), and supernatants were collected after 6, 20/24, and 48 h, and kept at -80°C until use. Whole blood was diluted 1:1 in RPMI medium and cultivated for 6 h, with medium, GLP and lipopolysaccharide as described above. Monensin was added after the first hour of culture. Supernatant cytokine levels from PBMC were measured by ELISA and intracellular IL-6 was detected in monocytes in whole blood cultures by flow-cytometry. Monocytes were identified in whole blood on the basis of forward versus side scatter parameters and positive reactions with CD45 and CD14 antibodies. GLP ( > or = 50 ng/ml)-induced IL-6 levels in supernatants were detected after 6-h incubation, reaching a peak after 20/24 h. The percentage of monocytes staining for IL-6 increased with increasing GLP concentration. Thus, our findings show a GLP-induced cellular activation by demonstrating the ability of GLP to induce IL-6 and the occurrence of monocyte activation in whole blood at the cellular level.

An imbalance of naive and memory/effector subsets and altered expression of CD38 on T lymphocytes in two girls with hyper-IgM syndrome.

In this report we evaluated CD4(+) T, CD8(+) T and natural killer (NK) cell counts, the levels of naive/memory subsets within the CD4(+) T lymphocyte population, expression of CD38 on T lymphocytes, and CD4(+) and CD8(+) T cell cytokine production in two girls with hyper-IgM (HIM) syndrome. Both girls developed recurrent infections early in infancy, presenting a wide spectrum of clinical manifestations, with a strikingly different disease severity between them. CD4(+) T cell counts were low in both children (patient 1: 214 cells/mm(3) and patient 2: 392 cells/mm(3)), and the CD4/CD8 T cell ratio was 0.4 for patient 1, the patient with the more severe disease, and 1.4 for patient 2. NK cell numbers were low in patient 1 (60 cells/mm(3)) and borderline (286 cells/mm(3)) with regard to normal levels in patient 2. An imbalance of naive and memory/effector cell subsets was found in both girls, with the percentage of CD45RA(+) 27(+) (naive) CD4(+) T lymphocytes being 5.8 and 12.4 for patients 1 and 2, respectively. Expression of CD38 on the surface of T lymphocytes was low in patient 1. Detection of intracellular interferon (IFN)-gamma and tumour necrosis factor (TNF)-alpha in CD4(+) and CD8(+) T lymphocytes upon PMA-Io stimulus was preserved in both children. In conclusion, we found low numbers of CD4(+) T lymphocytes and a dramatic redistribution of naive and memory/effector CD4(+) T lymphocytes in two girls with non-X-linked HIM syndrome. Furthermore, we found low expression of CD38 on T lymphocytes and low numbers of NK cells in the patient with the more severe disease, indicating a possible role for these cells in the pathogenesis of this immunodeficiency.