## 2 Discussion
### Limitations, Safety and Challenges  
Cell viability was examined in the majority of studies, using a variety of exposure checkpoints and concentrations. [PERSON] et al. reported no statistically significant change in cell viability and function 24 h post-exposure to 10, 20, 100, or 1000 uL of MNPs in vitro compared to controls [80]. Similarly, [PERSON] et al. investigated cell viability in immortalized and primary HCECs after 48 and 72 h of exposure to magnetic micro and nanoparticles (average size 100-1000 nm) [65]. They found that exposure to low concentrations (up to 20 ug/mL) over 48 and 72 h did not significantly affect viability, while higher concentrations (40 ug/mL and 80 ug/mL) reduced viability in immortalized HCECs; primary HCECs showed reduced viability only at the highest concentration of 80 ug/mL after 48 h. In a comparative study where bovine CEC viability was assessed in vitro, [PERSON] et al. found that cell viability remains unaffected until the concentration of iron oxide MNPs per cell reaches 100 \(\times\) 106; moreover, the study showed that the cytoskeletal integrity, measured by actin filament structure, was unaffected in the course of the study [79]. Although differences between human and animal CECs hinder direct comparisons, these results may indicate a dose-dependent toxic effect on cell viability. [PERSON] et al. monitored cell viability over a 7-day period and found no significant differences in viability after 1 day regardless of HA, MNP concentration, or magnet use [83]. By day 4, magnets adversely affected cell viability in non-HA controls at higher MNP concentrations (around 50 and 100 ug/mL), possibly due to increased MNP retention via endocytosis [80; 92]. Conversely, HA groups showed minimal impact on cell viability at 100 ug/mL and no significant effects at other concentrations; this protective effect became more pronounced with longer cell culture. By day 7, magnets began to detrimentally affect cell viability at concentrations as low as 25 ug/mL in the non-HA group, with effects intensifying at higher MNP concentrations. In contrast, cell viability in HA groups remained stable at 25 and 50 ug/mL, with significant impact only observed at 100 ug/mL, albeit to a lesser extent compared to the non-HA control. Furthermore, at the 7-day mark, cells cultured with 25 ug/mL MNPs in the HA group exhibited expression of tight junction marker ZO-1 and maintained F-actin structure between cells, indicatingthat cellular morphology of CECs remained unaffected by external magnetic fields and increasing MNP concentrations, consistent with earlier findings [65; 79; 80; 81]. Concerns over iron toxicity have been explored in [PERSON] et al. in vitro [82]. Concentrations of iron powder of 10 \(\upmu\)M or above decreased viability of rabbit CECs by over 50%. Therefore, the concentration was kept at a maximum of 5 \(\upmu\)M for in vivo experiments. Toxicity of magnetic micro and nanoparticles in rats was studied by [PERSON] et al., who found 50 nm-sized nanoparticles containing 55-59% of iron oxide had no statistically significant toxic effect on CECs; however, injection of 4 \(\upmu\)m-sized microparticles caused a reduction in CEC density after 1 week and 5 months [93]. These data introduced the consideration of long-term toxicity. When circulating systemically, nanoparticles have demonstrated potential toxic effects, such as triggering inflammatory responses and oxidative stress [94]. For instance, iron oxide nanoparticles induced cardiac oxidative stress and DNA damage in mice at remarkably low doses ranging from 0.4 to 10 \(\upmu\)g/kg [95]. Similarly, nickel nanoparticles induced cardiac toxicity and organ damage in rats at a dose of 20 mg/kg, resulting in arrhythmias and damage to the liver, spleen, and lungs [96]. However, it is noteworthy that the utilisation of nanoparticles within ocular contexts has not been associated with identifiable toxic effects [97].