A common mechanism of necrosis is induction of mito- chondrial permeability transition (MPT), which is manifested as accelerated cell death with plasma membrane disintegration. It is thought that a pore opens in the inner mitochondrial membrane allowing water and other molecules to pass through. Opening of this permeability transition pore can be triggered by multiple stimuli and leads to dissipation of the mitochondrial inner membrane potential ( m) (9). Irreversible induction of MPT leads to mitochondrial damage associated with mito- chondrial swelling and subsequent necrosis of the cell. In vitro M infected with virulent H37Rv causes the catastrophic irre- versible MPT that commits the M to necrosis (10). The active nature of this necrosis induction by virulent Mtb is clearly revealed after infection of M with mutants of Mtb. Although host M support growth of virulent Mtb (3), innate immune mechanisms are activated that limit pathogen survival after infection with Mtb mutants having altered viru- lence. These responses include induction of apoptosis, a slow cell death modality which leaves the plasma membrane intact and is observed after infection of M with attenuated Mtb (11–13). This M death modality limits exploitation of the intracellular growth environment through direct microbici- dal effects and by sequestering bacilli in apoptotic bodies. Indeed, apoptosis both enhances antigen presentation by DC (14) and facilitates efficient pathogen killing (15–18). Two pathways are described that lead to this highly regu- lated form of cell death (19). First, the extrinsic apoptotic pathway implicates binding of the ligands TNF and FasL to their receptors that trigger apoptosis (20). Second, the in- trinsic apoptotic pathway involves the mitochondria, which release cytochrome c and other factors from the mitochondrial intermembrane space that promote apoptosis (21, 22). We demonstrated previously that mitochondria play an es- sential role to determine whether Mtb-infected M undergo cytolytic necrosis or apoptosis (8). Apoptosis induced by the mitochondrial pathway commences with Ca2+ release from the ER that leads to an increase of Ca2+ in the mitochondria as well as translocation of BAX into the mitochondria and BAK activation (23). Permeabilization of the mitochondrial outer membrane then proceeds via formation of a proteolipid pore and subsequent escape of proteins from the mitochondrial in- termembrane space into the cytosol. More specifically, upon mitochondrial outer membrane permeabilization proapoptotic factors, including cytochrome c, are released from the mito- chondrial intermembrane space into the cytosol. There, cyto- chrome c forms a complex with Apaf-1, leading to activation of caspase-9 that, in turn, activates executioner caspases such as caspase-3, -6, and -7, which are instrumental in the induction of apoptosis (19, 24). Infection with the attenuated Mtb H37Ra, which has a mutation in PhoP which inhibits ESX-1 function (25), pre- dominantly prevents necrosis and leads to sequestration and decimation of the intracellular bacteria (8, 26). Mtb-induced apoptosis and antimycobacterial activity of human M requires the activity of cPLA2- , a group IV cyto- solic PLA2 (cPLA2) (27) which catalyzes the release of arachidonic 2792 acid (AA) from the sn-2 position of membrane phospho- lipids (28). AA and its functionally diverse and biologically active eicosanoid products have been implicated in the regula- tion of programmed cell death in several cell types (29, 30). The lipoxins are AA metabolites generated by 5- and 15-lipoxy- genases (5- and 15-LO) (31). Lipoxins modulate chemokine and cytokine expression, stimulate monocyte trafficking, and en- hance M engulfment of apoptotic leucocytes (32). However the role of 5-LO products of AA such as lipoxins and leukotri- ene B4, which amplifies PMN chemotaxis and production of granule products (33), has not been elucidated in modulating the death modality of M . Prostanoids are lipid mediators generated from AA by the enzymatic action of the cyclooxygenases (COX) 1 and 2 to form the intermediate PGH2, with subsequent metabolism to specific prostanoid species (e.g., PGD2, PGE2, PGF2 , PGI2, and thromboxane) by prostaglandin synthases (34, 35). The functions of these prostanoids are defined by an array of spe- cific receptors. In the case of PGE2, differential expression of four distinct EP receptors defines the intracellular pathways that are activated by PGE2 and results in specific functions that might either promote or inhibit inflammation. For example, in the lung, PGE2 either promotes vasodilatation and increases vascular permeability or increases bronchodilatation (36). A recent study describes a noninflammatory function of PGE2, up- or down-regulation of programmed cell death, which in- dicates that the mechanisms of PGE2 activation are complex and context dependent (37). Mice with a deleted 5-LO gene have increased IL-12, IFN- , and NO synthase 2 levels compared with WT mice after pulmonary Mtb infection. 5-LO / mice also have sig- nificantly lower Mtb burdens and survive longer than WT animals after respiratory Mtb infection (38). Importantly, ad- ministration of a stable LXA4 analogue was sufficient to re- verse the increased resistance of 5-LO / mice to Mtb infection. The finding that Mtb infection significantly increases AA pro- duction via activation of cPLA2- (27) and that 5 LO / mice are significantly more resistant to Mtb infections (38), raised the question of whether eicosanoid production is in- volved in macrophage necrosis and inhibition of apoptosis induced by virulent Mtb. We hypothesize that stimulation of lipoxin by virulent Mtb inhibits macrophage apoptosis, pro- motes necrosis, and represents a mechanism that allows Mtb to evade elimination by the innate immune system. We there- fore investigated how lipoxins and prostanoids affect the out- come of Mtb-driven M death with respect to apoptosis and necrosis as a novel pathogenic mechanism that regulates the innate immune response in Mtb infection. RESULTS Virulent Mtb trigger production of LXA4 in M We demonstrated that inhibition of cPLA2 abrogates apopto- sis in Mtb-infected M , which could be restored by reconsti- tution of AA, the predominant product of cPLA2 (27). These findings suggest that products synthesized from AA, namely the eicosanoids, regulate programmed M death induced LIPID MEDIATORS IN MTB INFECTION | Chen et al.