The production of interferon gamma (IFNgamma) guarantees effective T cell-mediated immunity against Mycobacterium tuberculosis infection. In the present study, we simply compare the in vitro immune responses to Mycobacterium antigens in terms of IFNg production in a total of 10 healthy Brazilian volunteers. Whole blood and mononuclear cells were cultivated in parallel with PPD, Ag85B, and M. bovis hsp65, and five-days supernatants were harvested for cytokine detection by ELISA. The inter-assay result was that the overall profile of agreement in response to antigens was highly correlated (r² = 0.9266; p = 0.0102). Potential analysis is in current progress to dictate the usefulness of this method to access the immune responses also in tuberculosis patients and its contacts.

The phenotypic features acquired subsequent to antigen-specific stimulation in vitro were evaluated by means of the kinetic expressions of CD69 and CD25 activation molecules on T lymphocytes and assayed by flow cytometry in response to PPD, Ag85B, and ferritin in PPD-positive healthy control individuals. In response to PHA, CD69 staining on both CD4+ and CD8+ T cells became initially marked after 4 h, peaked at 24 h, and quickly decreased after 120 h. For CD25, a latter expression was detected around 8 h, having increased after 96 h. As expected, the response rate to the mycobacterial antigens was much lower than that to the mitogen. Positive staining was high after 96 h for CD25 and after 24 h for CD69. CD69 expression was significantly enhanced (p < 0.05) on CD8+ as compared to CD4+ T cells. High levels were also found between 96-120 h. Regarding Ag85B, CD25+ cells were mostly CD4+ instead of CD8+ T cells. Moreover, in response to ferritin, a lower CD25 expression was noted. The present data will allow further characterization of the immune response to new mycobacterial-specific antigens and their evaluation for possible inclusion in developing new diagnostic techniques for tuberculosis as well in a new vaccine to prevent the disease.

Dados recentes sugerem que o curso clínico dos estados reacionais na lepra está estritamente relacionado à liberação local ou sistêmica de citocinas. Neste estudo, pacientes com ENL (erythema nodosum leprosum) foram grupados segundo a intensidade de seus sintomas clínicos. Os aspectos clínicos e imunológicos do ENL e os efeitos desses parâmetros na carga bacilar foram estabelecidos em conjunção com a resposta imune in vitro desses pacientes, a antígenos microbacterianos. Em 10 de 17 pacientes testados, o índice bacteriano (IB) foi reduzido cm pelo menos 1 log desde o diagnóstico da lepra até o aparecimento do primeiro episódio reacional, comparado a uma redução esperada de 0.3 log no grupo não reacional no mesmo período de MDT (multidrogaterapia). Entretanto, nenhuma diferença na taxa de redução do IB foi notada no final da MDT entre os pacientes ENL e os lepromatosos não reacionais. Assim também, embora os níveis de TNF-alfa (fator de necrose tumoral) estivessem aumentados no soro de 70,6% dos pacientes com ENL testados, nenhuma relação foi notada entre os níveis circulantes de TNF-alfa e a diminuição do IB detectada no aparecimento do primeiro episódio reacional. A avaliação da viabilidade bacteriana do M. leprae isolado de lesões reacionais não mostrou crescimento na pata de camundongo. Somente 20% dos pacientes demonstraram resposta imune específica ao M. leprae durante o ENL. Além disso altos níveis de IL-2R (receptor de interleucina 2) solúvel estiveram presentes em 78% dos pacientes. Anticorpos circulantes anti-nervo (anti-ceramídio e anti-galactocerebrosídio) e anti-micobacterianos foram também detectados no soro dos pacientes, e não estiveram relacionados ao curso clínico da doença. Nossos dados sugerem que a morte bacteriana está aumentada durante as reações. A emergência de resposta imune específica ao M. leprae e o papel efetivo do TNF-alfa na mediação de fragmentação bacteriana necessita ainda ser esclarecida

Recent data suggest that the clinical course of reactional states in leprosy is closely related to the cytokine profile released locally or systemically by the patients. In the present study, patients with erythema nodosum leprosum (ENL) were grouped according to the intensity of their clinical symptoms. Clinical and immunological aspects of ENL and the impact of these parameters on bacterial load were assessed in conjunction with patients' in vitro immune response to mycobacterial antigens. In 10 out of the 17 patients tested, BI (bacterial index) was reduced by at least 1 log from leprosy diagnosis to the onset of their first reactional episode (ENL), as compared to an expected 0.3 log reduction in the unreactional group for the same MDT (multidrug therapy) period. However, no difference in the rate of BI reduction was noted at the end of MDT among ENL and unreactional lepromatous patients. Accordingly, although TNF-alpha (tumor necrosis factor) levels were enhanced in the sera of 70.6% of the ENL patients tested, no relationship was noted between circulating TNF-alpha levels and the decrease in BI detected at the onset of the reactional episode. Evaluation of bacterial viability of M. leprae isolated from the reactional lesions showed no growth in the mouse footpads. Only 20% of the patients demonstrated specific immune response to M. leprae during ENL. Moreover, high levels of soluble IL-2R (interleukin-2 receptor) were present in 78% of the patients. Circulating anti-neural (anti-ceramide and anti-galactocerebroside antibodies) and anti-mycobacterial antibodies were detected in ENL patients' sera as well, which were not related to the clinical course of disease. Our data suggest that bacterial killing is enhanced during reactions. Emergence of specific immune response to M. leprae and the effective role of TNF-alpha in mediating fragmentation of bacteria still need to be clarified.