Zinc oxide nanoparticles have biphasic roles on Mycobacterium-induced inflammation by activating autophagy and ferroptosis mechanisms in infected macrophages

The ability of zinc oxide nanoparticles (ZnONPs) to induce bacteriostasis in Mycobacterium tuberculosis (M. tb) and their roles in regulating the pathogenic activities of immune cells have been reported previously, but the specific mechanisms underlying these regulatory functions remain unclear. This work aimed to determine how ZnONPs play the antibacterial role against M. tb. In vitro activity assays were employed to determine the minimum inhibitory concentrations (MICs) of the ZnONPs against various strains of M. tb (BCG, H37Rv, and clinical susceptible MDR and XDR strains). The ZnONPs had MICs of 0.5-2 mg/L against all tested isolates. In addition, changes in the expression levels of autophagy and ferroptosis-related markers in BCG-infected macrophages exposed to ZnONPs were measured. BCG-infected mice that were administered ZnONPs were used to determine the ZnONPs functions in vivo. ZnONPs decreased the number of bacteria engulfed by the macrophages in a dosedependent manner, while different doses of ZnONPs also affected inflammation in different directions. Although ZnONPs enhanced the BCG-induced autophagy of macrophages in a dose-dependent manner, only low doses of ZnONPs activated autophagy mechanisms by increasing the levels of pro-inflammatory factors. The ZnONPs also enhanced BCG-induced ferroptosis of macrophages at high doses. Co-administration of a ferroptosis inhibitor with the ZnONPs improved the anti-Mycobacterium activity of ZnONPs in an in vivo mouse model and alleviated acute lung injury caused by ZnONPs. Based on the above findings, we conclude that ZnONPs may act as potential antibacterial agents in future animal and clinical studies.

Automated summary

This study aimed to investigate the antibacterial mechanisms of zinc oxide nanoparticles (ZnONPs) against Mycobacterium tuberculosis (M. tb). In vitro activity assays showed that ZnONPs had minimum inhibitory concentrations (MICs) of 0.5-2 mg/L against all tested M. tb strains. The expression levels of autophagy and ferroptosis-related markers were measured in BCG-infected macrophages exposed to ZnONPs, and BCG-infected mice were used to determine the in vivo functions of ZnONPs. ZnONPs reduced the number of bacteria engulfed by macrophages in a dose-dependent manner and had varying effects on inflammation. Low doses of ZnONPs activated autophagy mechanisms, while high doses enhanced ferroptosis. Co-administration of a ferroptosis inhibitor improved the anti-Mycobacterium activity of ZnONPs in an in vivo mouse model. Based on these findings, ZnONPs have potential as antibacterial agents in future animal and clinical studies.