Efeito de micobactéria ambiental (Mycobacterium avium) na imunidade induzida pela vacina gênica para tuberculose (pVAXhsp65)

Abstract

A eficacia da vacina BCG (Mycobacterium bovis atenuado) contra tuberculose pulmonar apresenta grande variabilidade em diferentes populacoes. Dentre as hipoteses sugeridas, prevalece a que atribui esta variacao as interacoes entre a vacina e micobacterias presentes no meio ambiente. Estudos tem demonstrado que antigenos expressos pelo BCG estao conservados no M. avium, corroborando com a hipotese de que a exposicao as micobacterias ambientais gera resposta imune cruzada que interfere com a eficacia do BCG. O objetivo deste estudo foi avaliar o efeito da exposicao previa de camundongos ao M. avium inativado na resposta imune e eficacia protetora induzidas pela vacina DNAhsp65 (pVAXhsp65) na tuberculose experimental murina. Na primeira etapa, grupos de camundongos BALB/c femeas foram injetados com diferentes doses (0,08x106, 4x106 e 200x106) de M. avium inativado termicamente, pela via subcutanea. Apos tres semanas, os animais foram imunizados com DNAhsp65 por via intramuscular. Os animais receberam tres doses de DNA (100ug/intervalos de 15 dias). Grupos controle receberam somente M. avium, vacina (pVAXhs65), vetor (pVAX) ou solucao salina esteril. A producao de citocinas e os niveis de anticorpos foram determinados por ELISA. Na segunda etapa, os animais foram inicialmente sensibilizados por via subcutanea com 200x106 Unidades Formadoras de Colonias (UFC) de M. avium inativado e depois imunizados com tres doses de pVAXhsp65 (100ug/intervalos de 15 dias) por via intramuscular. Os grupos controle foram injetados com salina, pVAX (4 doses), pVAXhsp65 (4 doses), M. avium, M. avium mais pVAX (3 doses) ou M. avium mais pVAXhsp65 (3 doses). Quinze dias apos a ultima dose de DNA os animais foram infectados com 1x104 UFC de M. tuberculosis, H37Rv, pela via intratraqueal. Trinta dias apos o desafio os animais foram sacrificados e a carga bacteriana determinada pelo numero de UFC nos pulmoes. Cortes histopatologicos dos pulmoes tambem foram analisados.

The efficacy of BCG vaccine (attenuated Mycobacterium bovis) against pulmonary tuberculosis varies enormously in different populations. The prevailing hypothesis attributes this variation to interactions between the vaccine and mycobacteria common in the environment. Studies have revealed that most protective antigens expressed by the antituberculous vaccine are conserved in M. avium, supporting the hypothesis that exposure to environmental mycobacteria generates a crossreactive immune response that interferes with BCG efficacy.The objective of this investigation was to evaluate the effect of a prior exposure to heat killed M. avium on both, the immune response and the protective efficacy induced by a DNAhsp65 vaccine (pVAXhsp65) in experimental murine tuberculosis. To evaluate the effect on the immune response, BALB/c female mice were, initially, injected with distinct doses (0,08x104, 4x106 and 200x106) of heat killed M. avium by subcutaneous route. Three weeks later the animals were immunized with 3 doses of DNAhsp65 by intramuscular route (100æg / 15 days apart). Control groups received only M. avium, vaccine (pVaxhsp65), vector (pVax) or saline solution. Cytokine production and antibody levels were determined by ELISA. To evaluate the effect on the protective efficacy, animals were initially, sensitized with 2x108 heat killed CFU of M. avium by subcutaneous route and then immunized with 3 doses of pVAXhsp65 (100æg/15 days apart) by intramuscular route. Control groups were injected with saline, pVAX (4 doses), pVAXhsp65 (4 doses), M. avium, M. avium plus pVAX (3 doses) or M. avium plus pVAXhsp65 (3 doses). Fifteen days after last DNA dose the animals were infected with 1x104 viable CFU of H37Rv M. tuberculosis by intratracheal route. Thirty days after challenge the animals were sacrificed and the bacterial burden was determined by the number of CFU in the lungs. Lung histological sections were also analysed.