Saikosaponin A Targets HDAC6 to Inhibit Mycobacterium tuberculosis-Induced Macrophage Pyroptosis via Autophagy-Mediated NLRP3 Inflammasome Inactivation

Mycobacterium tuberculosis (Mtb) is among the oldest and most resilient human pathogens, remaining a major global public health threat. Its characteristic pathological features include granuloma formation and a systemic inflammatory response, primarily resulting from dysregulated host immune reactions. Therefore, host-directed therapy (HDT) is considered an important complement to conventional anti-TB treatment. This study sought to examine the inhibitory effects of Saikosaponin A (SSA), an active compound extracted from Bupleurum, on Mtb-induced macrophage pyroptosis, as well as the underlying molecular mechanisms. The effects of SSA on key molecules involved in pyroptosis and autophagy were examined in an in vitro model of Mtb-infected macrophages using Western blotting, ELISA, co-immunoprecipitation, and immunofluorescence assays. The function of histone deacetylase 6 (HDAC6) in modulating autophagy and pyroptosis in Mtb-infected macrophages was elucidated using gene silencing techniques. The SSA-HDAC6 interaction was validated using drug target identification methods such as molecular docking and site-directed mutagenesis. Furthermore, we established an in vivo model of lipopolysaccharide-induced pulmonary inflammation via intraperitoneal injection to assess whether SSA exerts a protective effect by inhibiting pyroptosis. In vitro experiments demonstrated that SSA enhanced autophagy to inactivate the NLRP3 inflammasome, thereby inhibiting Mtb-induced pyroptosis. Mechanistically, SSA interacted with HDAC6 and effectively suppressed its enzymatic activity. This interaction enabled SSA to target HDAC6, thereby modulating autophagy via the AMPK/mTOR/ULK1 axis, ultimately attenuating Mtb-induced pyroptosis in macrophages. Furthermore, in vivo experiments revealed that SSA regulated the acetylation of α-tubulin (Lys40), alleviating inflammatory lung injury in mice. SSA targets HDAC6 and exerts an immunomodulatory effect, highlighting its potential as a promising novel host-directed anti-tuberculosis agent.