Mycobacterium tuberculosis Uses Host Triacylglycerol to Accumulate Lipid Droplets and Acquires a Dormancy-Like Phenotype in Lipid-Loaded Macrophages

Abstract

Two billion people are latently infected with Mycobacterium tuberculosis (Mtb). Mtb-infected macrophages are likely to be sequestered inside the hypoxic environments of the granuloma and differentiate into lipid-loaded macrophages that contain triacylglycerol (TAG)-filled lipid droplets which may provide a fatty acid-rich host environment for Mtb. We report here that human peripheral blood monocyte-derived macrophages and THP-1 derived macrophages incubated under hypoxia accumulate Oil Red O-staining lipid droplets containing TAG. Inside such hypoxic, lipid-loaded macrophages, nearly half the Mtb population developed phenotypic tolerance to isoniazid, lost acid-fast staining and accumulated intracellular lipid droplets. Dual-isotope labeling of macrophage TAG revealed that Mtb inside the lipid-loaded macrophages imports fatty acids derived from host TAG and incorporates them intact into Mtb TAG. The fatty acid composition of host and Mtb TAG were nearly identical suggesting that Mtb utilizes host TAG to accumulate intracellular TAG. Utilization of host TAG by Mtb for lipid droplet synthesis was confirmed when fluorescent fatty acid-labeled host TAG was utilized to accumulate fluorescent lipid droplets inside the pathogen. Deletion of the Mtb triacylglycerol synthase 1 (tgs1) gene resulted in a drastic decrease but not a complete loss in both radiolabeled and fluorescent TAG accumulation by Mtb suggesting that the TAG that accumulates within Mtb is generated mainly by the incorporation of fatty acids released from host TAG. We show direct evidence for the utilization of the fatty acids from host TAG for lipid metabolism inside Mtb. Taqman real-time PCR measurements revealed that the mycobacterial genes dosR, hspX, icl1, tgs1 and lipY were up-regulated in Mtb within hypoxic lipid loaded macrophages along with other Mtb genes known to be associated with dormancy and lipid metabolism.

Protein kinase C-delta (PKCδ), a marker of inflammation and tuberculosis disease progression in humans, is important for optimal macrophage killing effector functions and survival in mice

“…We infer from these findings where PKCd ablation in mice renders them hypersusceptible to TB, and therefore it is likely that increased expression (blood) and abundance (granulomas) of this kinase reflect its role as a host factor to contain ongoing inflammation in humans. Host lipid bodies provide an excellent nutrient source for the survival of intracellular Mtb, 36 thereby decreasing its own metabolism 37 that allows pathogenesis 38 and persistence 12,39,40 of the bacterium. The necessity of triacylglycerol for slow replication was demonstrated in a recent study where deposition of excessive triacylglycerol in Mtb bacterium slowed down its growth as opposed to a Mtb strain capable of pumping out lipids.…”

Role of bioactive lipid mediators in obese adipose tissue inflammation and endocrine dysfunction

“…In this regard, the number of LD has been shown to increase during the interaction of macrophages with pathogens, such as parasites (Melo et al, 2003), bacteria (Peyron et al, 2008;Daniel et al, 2011;Cardona et al, 2000;Tanigawa et al, 2008;Cao et al, 2007) and viruses (Barba et al, 1997;Samsa et al, 2009). Moreover, during infection, LD are mobilized into phagosomes, so the pathogen can use them as energy sources and for increased production of inflammatory mediators.…”

Interplay of human macrophages and Mycobacterium tuberculosis phenotypes

“…Macrophages accumulated lipid bodies and Mtb synthesized lipid droplets from host-derived fatty acids, and similarly to the multiplestress in vitro model developed by the same group and described above, intracellular Mtb experienced loss of acid fastness [306]. Controlling Mtb replication for a few days only, it is not clear if upregulation of dosR and other dormancy-related genes and tolerance to INH was mediated by macrophage mechanisms, by the hypoxic cultivation conditions or by a combination of both [306]. Other studies on foamy macrophages confirmed the growth arrest, induction of dormancy-related genes as well as Mtb 'feeding' on host lipids [105,288].…”

“…Tolerance to INH could be recapitulated in a THP-1 macrophage-based model of Mtb infection when cells were cultivated at a reduced oxygen tension of 1%. Macrophages accumulated lipid bodies and Mtb synthesized lipid droplets from host-derived fatty acids, and similarly to the multiplestress in vitro model developed by the same group and described above, intracellular Mtb experienced loss of acid fastness [306]. Controlling Mtb replication for a few days only, it is not clear if upregulation of dosR and other dormancy-related genes and tolerance to INH was mediated by macrophage mechanisms, by the hypoxic cultivation conditions or by a combination of both [306].…”

“…Cell wall alterations are believed to be a consequence of Mtb residing in lung tissue [233] or inside macrophages [304] and were substantiated by transcriptomic findings of altered expression of cell wall synthesis genes in host cells [244,305]. These alterations lead to a decrease in acid-fastness, and coincide with the accumulation of TAG-containing lipid droplets in the cytoplasm of Mtb, probably derived from host cell lipids [288,306]. As demonstrated with a non-acid fast Mtb mutant, loss of acid fastness could be due to the synthesis of shorter mycolic acids, which further abolished cording of Mtb and produced an overall attenuated phenotype [307].…”