Staphylococcus aureus α-Toxin Induces Inflammatory Cytokines via Lysosomal Acid Sphingomyelinase and Ceramides

Abstract

Background/Aims: Staphylococcus aureus (S. aureus) infections are a major clinical problem and range from mild skin and soft-tissue infections to severe and even lethal infections such as pneumonia, endocarditis, sepsis, osteomyelitis, and toxic shock syndrome. Toxins that are released from S. aureus mediate many of these effects. Here, we aimed to identify molecular mechanisms how α-toxin, a major S. aureus toxin, induces inflammation. Methods: Macrophages were isolated from the bone marrow of wildtype and acid sphingomyelinase-deficient mice, stimulated with S. aureus α-toxin and activation of the acid sphingomyelinase was quantified. The subcellular formation of ceramides was determined by confocal microscopy. Release of cathepsins from lysosomes, activation of inflammasome proteins and formation of Interleukin-1β (IL-1β) and Tumor Necrosis Factor-α (TNF-α) were analyzed by western blotting, confocal microscopy and ELISA. Results: We demonstrate that S. aureus α-toxin activates the acid sphingomyelinase in ex vivo macrophages and triggers a release of ceramides. Ceramides induced by S. aureus α-toxin localize to lysosomes and mediate a release of cathepsin B and D from lysosomes into the cytoplasm. Cytosolic cathepsin B forms a complex with Nlrc4. Treatment of macrophages with α-toxin induces the formation of IL-1β and TNF-α. These events are reduced or abrogated, respectively, in cells lacking the acid sphingomyelinase and upon treatment of macrophages with amitriptyline, a functional inhibitor of acid sphingomyelinase. Pharmacological inhibition of cathepsin B prevented activation of the inflammasome measured as release of IL-1β, while the formation of TNF-α was independent of cathepsin B. Conclusion: We demonstrate a novel mechanism how bacterial toxins activate the inflammasome and mediate the formation and release of cytokines: S. aureus α-toxin triggers an activation of the acid sphingomyelinase and a release of ceramides resulting in the release of lysosomal cathepsin B and formation of pro-inflammatory cytokines.

Sulforaphane reduces intracellular survival of Staphylococcus aureus in macrophages through inhibition of JNK and p38 MAPK‑induced inflammation

“…In patients with atopic dermatitis, S. aureus recognition mediated an increase in the production of inflammatory cytokines IL-1β or IFN-γ and promoted the cytokine-dependent growth enhancement of S. aureus, increasing the severity of the skin lesion (33). A recent study suggested that S. aureus-alpha toxin may activate the inflammasome through activation of the acid sphingomyelinase, which resulted in the rapid release of cathepsins and the production of IL-1β and TNF-α in BMdMs (34). The present study demonstrated that S. aureus infection of macrophages triggered a robust increase in the transcriptional expression levels of proinflammatory genes, which was associated with increased expression levels of M1 marker genes.…”

Crosstalk Between Acid Sphingomyelinase and Inflammasome Signaling and Their Emerging Roles in Tissue Injury and Fibrosis

“…Recently, a further study investigated the role of ASM in inflammasome signaling (Ma et al, 2017). Staphylococcus aureus alpha-toxin (α-toxin) is a pore-forming toxin that triggers membrane permeabilization and induces inflammation (Cohen et al, 2016).…”

“…Release of cathepsins caused by lysosomal rupture is crucial for inflammasome activation (Hornung et al, 2008;Liu et al, 2016). The pharmacological inhibitor CA-074Me prevents the inflammasome signaling activating and the IL-1β production upon exposure to α-toxin (Ma et al, 2017). Besides, the mature form of cathepsin D follows the ceramide production pattern (Spengler et al, 2018).…”

“…The properties of tricyclic antidepressants and analogs lead them trapped and accumulated into the lysosome thereby detaching the ASM for lysosomal degradation, which makes these drugs as functional ASM inhibitor (Beckmann et al, 2014). The application of amitriptyline (Ma et al, 2017;Hong et al, 2019), desipramine (Chung et al, 2017), or imipramine (Biswas et al, 2017) normalizes ceramide levels and attenuates tissue injury and fibrosis. Importantly, a clinical trial shows systematic application of amitriptyline to CF patient results in beneficial effects on the lung and an increase in lung function (Nahrlich et al, 2013).…”

Infection of Primary Human Alveolar Macrophages Alters Staphylococcus aureus Toxin Production and Activity

“…We next determined the nature of SaCPs, as reports suggest that some S. aureus strains survive in acidic compartments containing active lysosomal enzymes (15,17,23). S. aureus-infected hAMs were investigated using fluorescence microscopy to determine bacterial association with the lysosomal aspartyl protease cathepsin D, the most definitive marker of a degradative lysosome.…”