Morphological analyses of caseous necrosis correlated to CD68+ and CD8+ cells in biopsies of tuberculous pleurisy / Análisis morfológico de la necrosis caseosa y su correlación con células CD68+ y CD8+ en biopsias de pleuritis tuberculosa

Tuberculosis is the most prevalent infectious disease in the world. Granuloma formation and caseous necrosis are hallmarks of M. tuberculosis infection and they represent the protective and inflammatory reactions in the infected tissues. The molecular mechanisms that mediate granuloma necrosis are still not well understood. Objectives: To immunolocalize and correlate the amounts of CD68+ macrophages and CD8+ lymphocytes to caseous necrosis extension in granulomas of tuberculous pleurisy. Methods: The study is a retrospective analysis of 30 pleural biopsies with histopathological diagnosis of chronic granulomatous pleurisy with caseous necrosis. These biopsies were classified according to necrosis intensity as minimal (N1), moderate (N2) and intense (N3). The number of granulomas was also observed and categorized as G1 (1 to 4 granulomas per section), G2 (5 to 8 granulomas per section), and G3 (more than 8 granulomas per section). Results: The means of CD68+ cells counts per mm² in N1, N2 and N3 categories of necrosis were 1,287 +/- 254, 1086 +/- 181 and 930 +/- 115 respectively. The means for CD8+ cells were 483.7 +/- 396, 366.3 +/- 43 and 558 +/- 53 cells per mm² in N1, N2 and N3 respectively. Conclusions: There were no significant statistical correlations between necrosis extension and cell counts. In analyzed biopsies, the number of CD68+ cells was significantly higher than the number of CD8+ cells.

La tuberculosis es una de las enfermedades más prevalentes en el mundo. La formación del granuloma junto con la necrosis caseosa son características propias de la infección por M. tuberculosis y representan reacciones inflamatorias y protectoras en los tejidos infectados. No se conocen bien los mecanismos moleculares que median la necrosis en el granuloma. Los objetivos fueron inmunolocalizar y correlacionar la cantidad de macrófagos CD68+ y linfocitos CD8+ con la extensión de la necrosis caseosa, en los granulomas de tuberculosis pleural. Análisis retrospectivo que incluyeron 30 biópsias con diagnóstico histopatológico de tuberculosis pleural granulomatosa crónica con necrosis caseosa. Estas biópsias fueron clasificadas según la intensidad de necrosis como mínima (N1), moderada (N2) e intensa (N3). También se determinó el número de granulomas, que fueron clasificados como G1 (1 a4 granulomas por sección), G2 (5 a 8 granulomas por sección), y G3 (más de 8 granulomas por sección). La cuantificación de células CD68+ por mm² en las categorías N1, N2 y N3 de necrosis fue de 1,287 +/- 254; 1086 +/-181 y 930 +/- 115, respectivamente. La cuantificación de las células CD68+ fue de 483,7 +/- 396; 366,3 +/- 43 y 558 +/- 53 células por mm² para N1, N2 y N3, respectivamente. No hubo correlación estadísticamente significativa entre la extensión de la necrosis y la cuantificación celular. El número de células CD68+ fue significativamente mayor que el número de células CD8+ en las biópsias analizadas.

Inhibition of caspase-8 activity reduces IFN-gamma expression by T cells from Leishmania major infection

Following infection with Leishmania major, T cell activation and apoptosis can be detected in draining lymph nodes of C57BL/6-infected mice. We investigated the mechanisms involved in apoptosis and cytokine expression following Tcellactivation. After two weeks of infection, apoptotic T cells were not detected in draining lymph nodes but activation with anti-CD3 induced apoptosis in both CD4 and CD8 T cells. Treatment with anti-FasLigand, caspase-8 or caspase- 9 inhibitors did not block activation-induced T-cell death. We also investigated whether the blockade of caspase-8 activity would affect the expression of type-1 or type-2 cytokines. At early stages of infection, both CD4 and CD8 T cells expressed IFN-gamma upon activation. Treatment with the caspase-8 inhibitor zIETD-fmk (benzyl-oxycarbonyl-Ile- Glu(OMe)-Thr-Asp(OMe)-fluoromethyl ketone) reduced the proportion of CD8 T cells and IFN-gamma expression in both CD4 and CD8T cells. We conclude that a non apoptotic role of caspase-8 activity may be required for T cell-mediated type-1 responses during L. major infection.

A ativação e a morte por apoptose de linfócitos T foram observadas em linfonodos drenantes de camundongos C57BL/6 infectados com Leishmania major. Investigamos os mecanismos envolvidos na apoptose e na expressão de citocinas após a ativação de linfócitos T. Após duas semanas de infecção, embora as células apoptóticas ainda não sejam detectadas em linfonodos drenantes, células T CD4 e CD8 sofrem apoptose após ativação com anti-CD3. O tratamento com anticorpo antagonista anti-Ligante de Fas, ou com inibidores das caspases-8 e 9, não bloqueou a morte induzida por ativação das células T. Investigamos também se a inibição da atividade da caspase-8 poderia afetar a expressão de citocinas tipo-1 ou tipo-2. Nos estágios iniciais da infecção, células T CD4 e CD8 de animais infectados com L. major expressaram IFN-gama após ativação. O tratamento com o inibidor de caspase-8 zIETD (benzoil-oxicarbonil-Ile-Glu(OMe)-Thr-Asp(OMe)-fluorometilcetona) durante a estimulação de células T reduziu a proporção de células T CD8 e a expressão de IFN-gama por células T CD4 e CD8. Concluimos que a atividade não apoptótica de caspase-8 pode ser necessária para o estabelecimento da imunidade mediada por células T durante a infecção por L. major.