GABA B R activation alleviates Streptococcus equi subsp. zooepidemicus -induced acute lung injury by suppressing inflammation and oxidative stress in pulmonary macrophages via IRAK3

ABSTRACT Streptococcus equi subsp. zooepidemicus (SEZ) infection poses a significant threat to animals and humans, causing acute lung injury driven by excessive inflammation and tissue destruction. The gamma-aminobutyric acid type B receptor (GABA B R) has been implicated in modulating inflammatory responses, but its specific role in SEZ-induced acute lung injury remains unexplored. This study aimed to investigate the protective effects of pharmacological GABA B R activation against SEZ-triggered pulmonary damage. Employing a mouse model of SEZ infection and MH-S alveolar macrophages, we demonstrated that pretreatment with baclofen downregulated a spectrum of pro-inflammatory mediators (CXCL9, CXCL10, GM-CSF, IL-1β, IL-6, IL-17A, and MCP-1) and counteracted oxidative stress by enhancing the NRF2/HO-1 antioxidant pathway and its downstream effectors (GPX4 and SOD) in both lung tissues and macrophages. Mechanistically, baclofen exerted its effects by elevating the expression of the negative regulator interleukin-1 receptor-associated kinase 3 (IRAK3), which in turn suppressed the NLRP3/caspase-1/gasdermin D (GSDMD) inflammatory axis. Due to the suppression of inflammation and oxidative stress, the pulmonary epithelial barrier was restored in infected mice, as evidenced by the upregulation of tight junction proteins (Claudin-1, Claudin-2, Occludin, and ZO-1) in the lung. These effects collectively markedly attenuated lung histopathological injury, reduced inflammatory cell infiltration, and restored goblet cell morphology and mucin secretion. Conversely, IRAK3 deficiency exacerbated SEZ-induced lung injury, barrier disruption, oxidative stress, and NLRP3/caspase-1/GSDMD pathway activation. Collectively, our findings establish that GABA B R activation protects against SEZ-induced acute lung injury by enhancing barrier integrity, reprogramming macrophage antioxidant responses, and inhibiting inflammatory signaling through an IRAK3-dependent mechanism, identifying GABA B R as a promising therapeutic target for bacterial pneumonia. IMPORTANCE Streptococcus equi subsp. zooepidemicus (SEZ) is a major zoonotic pathogen responsible for severe animal outbreaks and occasional life-threatening human infections, creating an urgent need for effective interventions. Gamma-aminobutyric acid type B receptor (GABA B R) represents a promising immunomodulatory target, as it is known to help limit inflammation during bacterial infections. In this study, we demonstrate that activating GABA B R pharmacologically protects against SEZ-induced acute lung injury by increasing interleukin-1 receptor-associated kinase 3 (IRAK3) expression in macrophages. This elevation of IRAK3 serves a dual function, suppressing the NLRP3/caspase-1/gasdermin D pathway while also strengthening antioxidant defenses and epithelial barrier integrity. Further support for this mechanism comes from the observation that IRAK3 deletion leads to exacerbated tissue injury. Taken together, these findings reveal a previously unrecognized GABA B R-IRAK3 regulatory axis that plays a central role in SEZ pathogenesis, highlighting a promising host-directed target for therapeutic development.