Potential Immunomodulatory Effects of Nb2C and Ti3C2 MXenes on Human Macrophages

Macrophages are essential components of innate immunity, playing a crucial role in the defence against pathogens and foreign materials. Polarisation of these versatile cells affects modulating inflammation and repair, and also impacts adaptive immunity reactions. MXenes, a class of two-dimensional transition metal carbides, nitrides or carbonitrides, have gained significant attention due to their unique properties and potential applications in various fields, including biomedicine. While the interaction of macrophages with MXenes has not been addressed yet, it can provide insights into the potential immunomodulatory effects of these nanoparticles. Therefore, this study aimed to investigate the interaction of MXenes, specifically Nb2C and Ti3C2, with human macrophages in vitro. 

For defining macrophage phenotype and activity, expression of CD68, CD163 and COX-2 was assessed after lipopolysaccharide (LPS) and MXene exposure. Peripheral blood mononuclear cells (PBMC) were isolated from human donor blood using density gradient centrifugation and 100,000 cells/cm2 were plated on glass slides in 24x well plates in standard complete cell culture medium. After 10-14 days in culture, part of PBMCs attached and differentiated to ~1000 macrophages/cm2, while the rest of the cells did not adhere and was washed away. Six experimental groups were established: a control untreated group; cells incubated with Nb2C MXenes for 1 and 3 days; cells incubated with Ti3C2 MXenes for 1 and 3 days; and cells stimulated with LPS (100 ng/ml) for 24 h. MXenes were used at 25 μg/ml. After incubation, cells were fixed in 4% formaldehyde for 10 min in PBS and subjected to immunocytochemistry analysis.

Morphological analysis revealed that Nb2C and LPS treated cells appeared larger in size compared to the control group, whereas the size of the Ti3C2 treated cells did not change. In the Nb2C MXene group, COX-2 expression was significantly higher compared to the control group, CD68 expression did not differ between groups, while CD163 expression was elevated compared to the control group. The Ti3C2 MXene group showed no differences in COX-2, CD68, and CD163 expression as compared to the control group. However, the analysis was hindered by the presence of an intensive black background, likely caused by the phagocytosed MXenes, making accurate assessment challenging. In the LPS group, COX-2 expression was higher than the control group, CD68 expression was elevated, and CD163 expression was significantly higher than the control group.

Consequently, this study demonstrated that MXene can modulate macrophage polarisation in vitro. Various types of MXenes differ in their effect on macrophage phenotype, demonstrating specific properties. Further investigations are required to investigate doze-dependent effect and decipher the mechanisms of various MXene influence on macrophage polarization and functional activity.