In situ analysis of lung antigen-presenting cells during murine pulmonary infection with virulent Mycobacterium tuberculosis

Pulmonary tuberculosis, an infection caused by Mycobacterium tuberculosis has been declared a global health emergency by the World Health Organization (WHO). Tuberculosis is a re-emergent expanding disease, responsible for a great mortality and morbidity throughout the world. It has been estimated that one-third of the world's population is currently infected with M. tuberculosis, with 8 million new cases per year. Nevertheless, only 5–10% will develop the chronic pulmonary disease (Dye et al. 1999; WHO 2000). Most people will be able to control the infection through efficient immune responses, especially the cellular arm of immunity. However, the bacilli might not be eliminated completely and may persist dormant for years in a latent state, a phenomenon not well understood yet (Flynn & Ernest 2000; Hernandez-Pando et al. 2000; Manabe & Bishai 2000). M. tuberculosis spreads from a person with pulmonary tuberculosis through aerosols, entering the body via the respiratory tract, reaching the lungs where phagocytic cells become infected, especially the alveolar macrophages (AMs). Apparently, M. tuberculosis can survive within AM for long-lasting periods, causing either a latent infection or subsequently developing the pulmonary disease (Wiegeshaus et al. 1989; Andersen 1997). This bacillus has evolved several mechanisms to evade the host immune responses, including the inhibition of antimicrobial activity of macrophages (Schluger & Rom 1998). It is known that the first line of defence against M. tuberculosis is the AM, which interacts with the bacilli through different surface molecules like complement, mannose and Toll-like receptors (Arthur & Dannenberg 1991; Schluger & Rom 1998). Despite the fact that dendritic cells (DCs) are present throughout the respiratory tract, from the upper nasal mucosa to parenchymal lung tissue, little is known about lung DCs in vivo during tuberculosis, either in the naturally occurring disease or in experimental models of infection. Lung DCs are found within pleura and the alveolar septal walls; additionally they can be found, although in very small numbers, in the alveolar spaces (Holt & Schon–Hegrad 1987; McWilliam et al. 1995; Holt 2000). In vitro assays have shown that monocyte-derived DCs and macrophages internalize the mycobacteria, resulting in DC maturation and activation with a differential production of cytokines. Interleukin-12 (IL-12) was almost exclusively produced by DCs, while macrophages produced IL-10 and the pro-inflammatory cytokines tumour necrosis factor-α (TNF-α), IL-1 and IL-6 (Mohagheghpour et al. 2000; Giacomini et al. 2001). M. tuberculosis grew equally well within nonactivated DCs and macrophages; however, activation with IFN-γ and LPS inhibited the growth of this intracellular microorganism. While macrophages can kill the intracellular bacteria, DCs seem unable to do so and, apparently, only restrict the growth of this bacillus (Bodnar et al. 2001). Experiments conducted in vivo with the M. bovis (BCG) showed that, following intravenous administration, bacilli were detected in similar percentage in splenic macrophages and in splenic DCs. After 2 weeks, BCG bacilli were still viable without apparent growth within DCs (Jiao et al. 2001). In this study, using four different markers in situ, we describe the main lung antigen-presenting cells (APCs) in the susceptible BALB/c mice during experimental pulmonary tuberculosis, which is induced by intratracheal inoculation of the virulent strain of M. tuberculosis H37Rv. Pulmonary infection with M. tuberculosis induced an increase of parenchymal lung APC; these cells displayed differential tissue distribution and morphological features during the infection and, apparently, also a differential capacity to interact with bacilli in vivo.