Protective effect of ginsenoside Rh2 against Toxoplasma gondii infection-induced neuronal injury through binding TgCDPK1 and NLRP3 to inhibit microglial NLRP3 inflammasome signaling pathway

Toxoplasma gondii (T. gondii) is a neurotropic obligate intracellular parasite that can activate microglial and promote neuronal apoptosis, leading to central nervous system diseases. The NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome signaling complex plays a key role in inducing neuroinflammation. Our previous studies have found that ginsenoside Rh2 (GRh2) inhibits T. gondii infection-induced microglial activation and neuroinflammation by downregulating the Toll-like receptor 4/nuclear factor-kappa B signaling pathway. However, whether GRh2 reduces T. gondii infection-induced neuronal injury through actions on microglial NLRP3 inflammasome signaling has not yet been clarified.In this study, we employed T. gondii RH strain to establish in vitro and in vivo infection models in BV2 microglia cell line and BALB/c mice. Molecular docking, localized surface plasmon resonance assay, quantitative competitive-PCR, ELISA, western blotting, flow cytometric analysis, and immunofluorescence were performed.Our results showed that GRh2 alleviated neuropathological damage and neuronal apoptosis in cortical tissue of T. gondii-infected mice. GRh2 and CY-09 (an inhibitor of NLRP3) exhibited potent anti-T. gondii effects through binding T. gondii calcium-dependent protein kinase 1 (TgCDPK1). GRh2 decreased Iba-1 (a specific microglial marker) and NLRP3 inflammasome signaling pathway-related protein expression by binding NLRP3. Co-culture of microglia/primary cortical neurons revealed that T. gondii-induced microglial activation caused neuronal apoptosis, but GRh2 reduced this effect, consistent with the effects of CY-09.Taken together, our results show that GRh2 has a protective effect against T. gondii infection-induced neuronal injury by binding TgCDPK1 and NLRP3 to inhibit NLRP3 inflammasome signaling pathway in microglia.