Group A streptococcus (GAS) imposes a great burden on humans. Efforts to minimize the associated morbidity and mortality represent a critical issue. Glycogen synthase kinase-3 β (GSK-3 β) is known to regulate inflammatory response in infectious diseases. However, the regulation of GSK-3 β in GAS infection is still unknown. The present study investigates the interaction between GSK-3 β , NF- κ B, and possible related inflammatory mediators in vitro and in a mouse model. The results revealed that GAS could activate NF- κ B, followed by an increased expression of inducible nitric oxide synthase (iNOS) and NO production in a murine macrophage cell line. Activation of GSK-3 β occurred after GAS infection, and inhibition of GSK-3 β reduced iNOS expression and NO production. Furthermore, GSK-3 β inhibitors reduced NF- κ B activation and subsequent TNF- α production, which indicates that GSK-3 β acts upstream of NF- κ B in GAS-infected macrophages. Similar to the in vitro findings, administration of GSK-3 β inhibitor in an air pouch GAS infection mouse model significantly reduced the level of serum TNF- α and improved the survival rate. The inhibition of GSK-3 β to moderate the inflammatory effect might be an alternative therapeutic strategy against GAS infection.