Streptococcus anginosus activates the NLRP3 inflammasome to promote inflammatory responses from macrophages

Chronic inflammation and oral dysbiosis are common features of oral squamous cell carcinoma (OSCC). The commensal bacterium Streptococcus anginosus is increased in oral diseases, including OSCC. Our previous work revealed that S. anginosus promotes inflammatory responses from macrophage cell lines; however, the molecular mechanism by which S. anginosus interacts with macrophages to instigate this response remains to be investigated. Here, we found S. anginosus activated primary bone marrow-derived macrophages (BMMs), which presented increased NF-κB activation and downstream inflammatory cytokines TNF-α, IL-6, and IL-1β at 24 h after infection. S. anginosus viability, TLR2, TLR4, and MyD88 were dispensable for NF-κB activation, but each promoted the induction of distinct downstream inflammatory mediators, with only MyD88 being necessary for NF-κB activation in response to heat-killed S. anginosus. S. anginosus replicated intracellularly within BMMs without causing cell death and induced expression of inflammasome sensors AIM2, NLRC4, and NLRP3. S. anginosus-infected BMMs lacking the inflammasome adapter protein ASC (Asc-/-) or caspase-1 (Caspase1-/-) had significantly diminished IL-1β production compared with wild-type (WT) BMMs, indicating that S. anginosus activated the inflammasome. S. anginosus primarily triggered the inflammasome through NLRP3: S. anginosus-infected Nlrp3-/- BMMs and NLRP3 inhibitor (MCC950)-treated WT BMMs displayed diminished IL-1β production compared with WT controls. Lastly, S. anginosus-infected Asc-/-, and to a lesser extent Nlrp3-/-, mice displayed reduced weight loss, reduced inflammatory cytokines, and increased bacterial burden compared with C57BL/6 mice. These findings indicate S. anginosus replicates within macrophages and promotes a proinflammatory response in part through activating the NLRP3 inflammasome.