High interleukin-4 expression and interleukin-4 gene polymorphisms are associated with susceptibility to human paracoccidioidomycosis.

Paracoccidioidomycosis (PCM) is caused by dimorphic fungi from the Paracoccidioides brasiliensis complex. Previous studies have demonstrated that the severity of disease is associated with a T-helper 2 immune response characterised by high interleukin (IL)-4 production. In the present study we analysed two polymorphisms in the IL-4 gene (-590 C/T and intron-3 microsatellite) in 76 patients with PCM and 73 control subjects from an endemic area. The production of IL-4 by peripheral blood mononuclear cells after antigen or phytohaemagglutinin stimulation was determined by ELISA. A significant correlation was observed between the RP2/RP2 intron-3 genotype and infection with Paracoccidioides sp.(p = 0.011), whereas the RP1/RP1 genotype was correlated with resistance. No significant correlation was observed for the IL-4 promoter polymorphism. Furthermore, the low IL-4 expression observed in the control group compared with patients was associated with the RP1/RP1 genotype. These results suggest that IL-4 polymorphisms might be associated with the ability of the host to control Paracoccidioides sp.infection. The relevance of this polymorphism is supported by the observation that patients with disease produce high levels of IL-4 following mitogen or antigen stimulation. The IL-4 gene is located in the cytokine cluster region of chromosome 5 where other polymorphisms have also been described.

Genetic variation of FcγRIIa induces higher uptake of Leishmania infantum and modulates cytokine production by adherent mononuclear cells in vitro.

Introduction: Single nucleotide variations (SNVs) are specific genetic variations that commonly occur in a population and often do not manifest phenotypically. However, depending on their location and the type of nucleotide exchanged, an SNV can alter or inhibit the function of the gene in which it occurs. Immunoglobulin G (IgG) receptor genes have exhibited several polymorphisms, including rs1801274, which is found in the FcgRIIa gene. The replacement of A with T results in a Histidine (H) to Arginine (R) substitution, altering the affinity of the IgG receptor for IgG subtypes and C-reactive protein (CRP). In this study, we analyzed rs1801274 and its functional implications concerning L. Infantum uptake and cytokine production. Methods: We genotyped 201 individuals from an endemic area for visceral leishmaniasis to assess the presence of rs1801274 using Taqman probes for a candidate gene study. Additionally, we included seventy individuals from a non-endemic area for a functional study. Subsequently, we isolated and cultivated one-week adherent mononuclear cells (AMCs) derived from the peripheral blood of participants residing in the non-endemic region in the presence of L. infantum promastigotes, with and without antigen-specific IgG and/or CRP. We analyzed the rate of phagocytosis and the production of nitric oxide (NO), tumor necrosis factor (TNF)-a, interleukin (IL)-10, IL-12 p70, IL-1b, IL- 6, and IL-8 in the culture supernatants. Results and discussion: In participants from the endemic region, the A/G (H/R isoform) heterozygous genotype was significantly associated with susceptibility to the disease. Furthermore, SNVs induced a change in the phagocytosis rate in an opsonin-dependent manner. Opsonization with IgG increased the production of IL-10, TNF-a, and IL-6 in AMCs with the H/R isoform, followed by a decrease in NO production. The results presented here suggest that the rs1801274 polymorphism is linked to a higher susceptibility to visceral leishmaniasis.

Infection with Leishmania (Leishmania) infantum of 0 to 18-Month-old children living in a visceral leishmaniasis-endemic area in Brazil.

The diagnosis of Leishmania (Leishmania) infantum infection in children from birth may serve as a reference for the early identification of cases that would progress to classical visceral leishmaniasis (VL) in endemic areas. This study prospectively evaluated newborns of mothers living in the municipality of Paracatu, Minas Gerais, Brazil. The infants were followed up at 6-month intervals by clinical examination, serological tests (immunofluorescence [IIF] and enzyme-linked immunosorbent assay with rK39 [ELISA-rK39]) and polymerase chain reaction (PCR) until they had completed 18 months of age. A total of 166 pregnant women were included to evaluate the possible transfer of antibodies or even congenital transmission. Twenty-two of the women tested positive by IIF, four by ELISA-rK39, and one by PCR. Three infants of the 25 women with some positive test results were also positive in the first test (one by IIF, one by ELISA-rK39, and the third by ELISA-rK39 and PCR). One hundred and sixty infants were included in the study; of these, 43 had at least one positive sample over time. However, agreement between tests was low. Follow-up of children with a positive result in the tests studied revealed no progression to classical disease within a period of 18 months. In contrast, two children with negative IIF, PCR, and ELISA-rK39 results developed classical VL at 9 and 12 months of age. In conclusion, a positive test result was variable and sometimes temporary and agreement between tests was low. Therefore, the early diagnosis of Leishmania infection was not associated with the early identification of cases that would progress to classical VL in the endemic area studied.

Association between the lymphotoxin-alpha gene polymorphism and chagasic cardiopathy.

Lymphotoxin-alpha (LT-alpha or LTA) is an inflammatory cytokine that is involved in the organization and maintenance of the inflammatory process and in the arrangement of cells at the site of inflammation. These features suggest an important role in the development of chronic Chagas' disease, especially the cardiac form. The objective of this study was to evaluate LT-alpha genetics and its biological role in chronic Chagas' disease. A total of 284 subjects were studied. The LT-alpha single-nucleotide polymorphism (+252) was analyzed by the polymerase chain reaction-restriction fragment length polymorphism and expression by enzyme-linked immunosorbent assay in culture supernatants and in individual T lymphocytes by flow cytometry. The risk of developing the cardiac form was 2.8 times higher among carriers of genotype GG and 2.4 times among carriers of genotype GA when compared to subjects carrying genotype AA. Seropositive subjects carrying the G allele produced significantly higher levels of LT-alpha. The cytokine was mainly expressed by CD8(+) T lymphocytes in the absence of any stimulus and after stimulation with the Trypanosoma cruzi antigen. This study provides genetic and biological evidence for an important role of LT-alpha in the development of the cardiac form of Chagas' disease.

Genetic and functional role of TNF-alpha in the development Trypanosoma cruzi infection.

TNF-alpha plays an important role in trypanocidal mechanisms and is related to tissue injury. This cytokine has been detected in the heart of human chagasic patients where it is associated with tissue damage. This study investigated whether TNF-alpha levels and the presence of genetic polymorphisms are associated with the presence of T. cruzi infection and/or with the development of the cardiac form in chronic chagasic patients. Genomic DNA of 300 subjects from an endemic area was extracted and analyzed by PCR using specific primers. TNF-alpha was assayed in culture supernatants by ELISA. An association was observed between the absence of the TNF-238A allele and negative serology. Furthermore, seropositive individuals carrying the TNF-238A allele produced significantly higher TNF-alpha levels without stimulation (p=0.04) and after stimulation with LPS (p=0.007) and T. cruzi antigens (p=0.004). The present results suggest that the polymorphism at position -238 influences susceptibility to infection and that this allele is associated with higher TNF-alpha production in seropositive individuals.

High interleukin-4 expression and interleukin-4 gene polymorphisms are associated with susceptibility to human paracoccidioidomycosis

Paracoccidioidomycosis (PCM) is caused by dimorphic fungi from theParacoccidioides brasiliensis complex. Previous studies have demonstrated that the severity of disease is associated with a T-helper 2 immune response characterised by high interleukin (IL)-4 production. In the present study we analysed two polymorphisms in the IL-4gene (-590 C/T and intron-3 microsatellite) in 76 patients with PCM and 73 control subjects from an endemic area. The production of IL-4 by peripheral blood mononuclear cells after antigen or phytohaemagglutinin stimulation was determined by ELISA. A significant correlation was observed between the RP2/RP2 intron-3 genotype and infection with Paracoccidioides sp.(p = 0.011), whereas the RP1/RP1 genotype was correlated with resistance. No significant correlation was observed for the IL-4promoter polymorphism. Furthermore, the low IL-4 expression observed in the control group compared with patients was associated with the RP1/RP1 genotype. These results suggest that IL-4polymorphisms might be associated with the ability of the host to control Paracoccidioides sp.infection. The relevance of this polymorphism is supported by the observation that patients with disease produce high levels of IL-4 following mitogen or antigen stimulation. The IL-4gene is located in the cytokine cluster region of chromosome 5 where other polymorphisms have also been described.

[Association between the plasma levels of TNF-alpha, IFN-gamma, IL-10, nitric oxide and specific IgG isotypes in the clinical forms of chronic Chagas disease]. / Associação entre os níveis plasmáticos de TNF-alpha, IFN-gamma, IL-10, óxido nítrico e os isotipos de IgG específicos nas formas clínicas da doença de Chagas crônica.

Chagas disease is an important chronic parasitic disease that affects around 9-11 million people in Latin America. A combination of parasite and host-related factors are probably responsible for pathogenesis in the chronic phase of the disease. Among the host-related factors, the immunological response is a parameter of special interest. Our aim here was to evaluate the plasma levels of the cytokines interferon gamma, interleukin 10 and tumor necrosis factor alpha and the immunoglobulins total IgG and its subclasses 3 and 4, by means of ELISA, and the levels of nitric oxide by means of the Griess reaction, among individuals who were seropositive for Trypanosoma cruzi, presenting the cardiac, indeterminate and digestive clinical forms of the disease, and among seronegative individuals. The seropositive individuals produced significantly higher levels of total IgG and IgG-3. Individuals with the digestive form presented higher levels of IgG-4 and interleukin 10. However, these individuals presented lower levels of nitric oxide than the controls did. The results suggest that the higher levels of interleukin 10 observed among individuals with the digestive form may contribute towards the higher levels of the specific IgG-4 that were seen.

Associação entre os níveis plasmáticos de TNF-α, IFN-γ, IL-10, óxido nítrico e os isotipos de IgG específicos nas formas clínicas da doença de Chagas crônica / Association between the plasma levels of TNF-α, IFN-γ, IL-10, nitric oxide and specific IgG isotypes in the clinical forms of chronic Chagas disease

A doença de Chagas é uma importante doença parasitária crônica, que acomete cerca de 9-11 milhões de pessoas na América Latina. Provavelmente, uma combinação de fatores relacionados ao parasito e ao hospedeiro podem ser os responsáveis pela patogênese na fase crônica da doença. Dentre os fatores relacionados ao hospedeiro, a resposta imunológica é um parâmetro de especial interesse. Objetivamos avaliar os níveis plasmáticos das citocinas interferon gama, interleucina 10, fator de necrose tumoral alfa e das imunoglobulinas G total, 3 e 4, por ELISA e do óxido nítrico, pela reação de Griess, entre indivíduos soronegativos e soropositivos para Trypanosoma cruzi, com as formas clínicas cardíaca, indeterminada e digestiva. Os indivíduos soropositivos para Trypanosoma cruzi produziram níveis significativamente mais elevados de imunoglobulinas G total e G3. Indivíduos com a forma digestiva apresentam níveis mais elevados de imunoglobulina G4 e interleucina 10. Entretanto, tais indivíduos apresentaram menores níveis de óxido nítrico do que controles. Os resultados sugerem que os maiores níveis de IL-10 observados nos indivíduos com a forma digestiva poderiam contribuir com os maiores níveis de IgG4 específicos observados.

Chagas disease is an important chronic parasitic disease that affects around 9-11 million people in Latin America. A combination of parasite and host-related factors are probably responsible for pathogenesis in the chronic phase of the disease. Among the host-related factors, the immunological response is a parameter of special interest. Our aim here was to evaluate the plasma levels of the cytokines interferon gamma, interleukin 10 and tumor necrosis factor alpha and the immunoglobulins total IgG and its subclasses 3 and 4, by means of ELISA, and the levels of nitric oxide by means of the Griess reaction, among individuals who were seropositive for Trypanosoma cruzi, presenting the cardiac, indeterminate and digestive clinical forms of the disease, and among seronegative individuals. The seropositive individuals produced significantly higher levels of total IgG and IgG-3. Individuals with the digestive form presented higher levels of IgG-4 and interleukin 10. However, these individuals presented lower levels of nitric oxide than the controls did. The results suggest that the higher levels of interleukin 10 observed among individuals with the digestive form may contribute towards the higher levels of the specific IgG-4 that were seen.