Porphyromonas gingivalis sphingolipids affect early responses of THP-1 macrophages to outer membrane vesicles

Porphyromonas gingivalis is an oral bacterium commonly associated with periodontitis. P. gingivalis synthesizes sphingolipids (SLs), and recently, we reported that P. gingivalis SLs packaged into outer membrane vesicles (OMVs) can regulate THP-1 macrophage inflammatory responses to P. gingivalis OMVs. The contribution that P. gingivalis SLs have on host-pathogen interactions remains poorly understood, especially in the context of OMVs. Here, we demonstrate that P. gingivalis SLs significantly reduce the uptake of OMVs isolated from SL-containing wild-type (WT) P. gingivalis compared to OMVs from an SL-null mutant P. gingivalis strain, and that the loss of SLs drives uptake of P. gingivalis OMVs via lipid rafts in THP-1 macrophages. Intriguingly, we found that the sensing of P. gingivalis OMVs via TLR2 and MyD88 is attenuated due to the presence of SLs. Lastly, transcriptomic analysis of THP-1 macrophages co-cultured with either WT or SL-null P. gingivalis OMVs for 2 h revealed an array of differentially expressed genes. Interestingly, at 2 h, WT P. gingivalis OMVs promoted an overall trend of gene downregulation in THP-1 macrophages when compared to unchallenged, whereas SL-null OMVs strongly promoted upregulated gene expression. These findings provide early-phase characterization of the role of SLs in initial host cellular responses to P. gingivalis OMVs and expand our knowledge of interactions between SL-containing P. gingivalis OMVs and the host.