A new study published in the Investigative Ophthalmology & Visual Science investigated the anti-Acanthamoeba effect of exogenous nitric oxide (NO). In this study, Acanthamoeba castellanii was axenically cultured and exposed to various concentrations of NO sources, such as sodium nitrite, sodium nitroprusside (SNP), and NO-releasing silica nanoparticles, for 1 to 7 days. Human corneal epithelial cells (HCECs) were cultured and exposed to sodium nitrite, SNP (0, 0.1, 1, 10, 100, and 1000 μM), and silica nanoparticles for 1, 2, and 3 days. A dose-dependent inhibitory effect of sodium nitrite and SNP was noted on A. castellanii viability. Meanwhile, a more prominent inhibitory effect was observed with SNP compared to sodium nitrite. However, SNP was found to be more cytotoxic for HCEC. The findings revealed that NO-releasing silica nanoparticles were successfully internalized into the amoebic cytoplasm and accumulated in large vacuoles. Although blank silica nanoparticles had no inhibitory effect on A. castellanii viability, NO-releasing silica nanoparticles showed a dose-dependent amoebicidal effect. Additionally, no cystic transformation of A. castellanii was observed under a phase contrast microscope or transmission electron microscope after an exogenous NO treatment. Hence, the results confirmed the anti-Acanthamoeba effect of exogenous NO. Thus, it was stated that NO-releasing drug-platforms, including nano-carriers, can be a promising therapeutic strategy for Acanthamoeba keratitis.