Lactoferrin-modified nanoemulsions enhance brain-targeting and therapeutic efficacy of arctigenin against Toxoplasma gondii-induced neuronal injury

Toxoplasma gondii, a neurotropic protozoan parasite, affects the central nervous system and causes various neurological disorders. Previous studies have demonstrated that Arctigenin (AG) exhibits anti-T. gondii activity and reduces depression-like behaviors induced by T. gondii infection. This study aimed to enhance AG's brain-targeting and therapeutic efficacy by developing lactoferrin-modified nanoemulsions loaded with AG (Lf-AG-NEs). Lf-modified nanoemulsions were prepared and assessed using in vivo and in vitro infection models with the T. gondii RH strain, and a co-culture system of BV2 microglia and primary neuron cells. The effects of Lf-AG-NEs on T. gondii-induced neuronal injury were examined, and potential molecular mechanisms were elucidated through real-time quantitative PCR, western blotting, immunofluorescence, flow cytometry, immunohistochemistry, and Nissl staining. In vitro assessments showed significant increases in cellular uptake and blood-brain barrier penetration by Lf-AG-NEs. These nanoemulsions notably inhibited T. gondii proliferation in brain tissue and BV2 cells, surpassing the effects of free AG or AG-NEs alone. Additionally, Lf-AG-NEs substantially alleviated neuropathological changes and reduced microglial activation and neuroinflammation by downregulating the TLR4/NF-κB and TNFR1/NF-κB signaling pathways. Co-culturing BV2 cells with primary cortical neurons indicated that Lf-AG-NEs, similarly to CLI-095 and R7050, attenuated T. gondii-induced microglial activation and subsequent neuronal injury. In conclusion, the successfully prepared Lf-AG-NEs not only enhanced the anti-T. gondii effect but also strengthened the protective impact against neuronal injury induced by T. gondii, through the modulation of microglial signaling pathways.