Mycobacterium bovis-infected cervine alveolar macrophages secrete lymphoreactive lipid antigens

Tuberculosis is caused by intracellular bacteria belonging to the genus Myc obacterium, including M. tuberculosis and M. bovis. Alveolar macrophages (A Ms) are the primary host cell for inhaled mycobacteria. However, little is known about the mechanisms by which infected AMs can process and present my cobacterial antigens to primed lymphocytes and how these responses may affe ct ensuing protection in the host. In the present study, we sought to deter mine whether AMs from a naturally susceptible host for Mycobacterium bovis (red deer) could produce and secrete soluble immunoreactive antigens follow ing mycobacterial infection in vitro. Confluent monolayers of deer AMs were infected,vith either heat-killed or live virulent M. bovis or M. bovis BCG at a multiplicity of infection of 5:1 and cultured for 48 h. Culture super natants were collected, concentrated, and tested for the presence of mycoba cterial antigens in a lymphocyte proliferation assay by using peripheral bl ood mononuclear cells from M. bovis-sensitized or naive deer. Supernatants derived from macrophages which had been infected with live bacilli stimulat ed the proliferation of antigen-sensitized, but not naive, lymphocytes. Sup ernatants derived from uninoculated AMs or AMs inoculated with heat-killed bacilli failed to stimulate lymphocyte proliferation. The lymphoproliferati ve activity was retained following lipid extraction of the supernatants, wh ich were free of amino groups as determined by thin-layer chromatography. T hese results demonstrate that mycobacteria which are actively growing withi n AMs produce lipids which are secreted into the extracellular milieu and t hat these lipids are recognized by lymphocytes from mycobacterium-primed ho sts. We suggest that mycobacterial lipids are released from AMs following a erosol infection in vivo and that they play an important role in the early immune response to tuberculosis.