| Summary: | Acanthamoeba keratitis (AK) is a severe corneal infection that poses a public health problem. Treatment of AK remains challenging because it relies on prolonged therapeutic regimens often utilising drug combinations. The considerable damage to human ocular tissue of patients with AK is partially mediated by soluble factors present in the protozoan Acanthamoeba castellanii’s secretome. The two aims of this thesis were to characterise the structural, functional and metabolomic changes occurring in human corneal cells caused by A. castellanii secretome and to investigate the potential amelioration of infection using cerium oxide nanoparticles (CeO2NPs).
Human SV40 immortalised corneal epithelial cells (ihCECs) and corneal stromal cells (CSCs) were treated with various concentrations of A. castellanii-conditioned medium (ACCM) for different exposure times. ACCM significantly decreased cell viability, triggered apoptosis, disrupted the cell actin cytoskeleton and altered the ultrastructural properties of corneal cells, in a time- and concentration-dependant manner. These ACCM-related cellular alterations were mediated by A. castellanii proteases. Amino acid analysis of ACCM showed that serine and cysteine proteases are secreted by A. castellanii. ACCM also altered the levels of elements within CSCs, including changes in the abundance of potassium, sodium and calcium. However, ACCM exposure did not affect the DNA concentration of CSC cells. Liquid chromatography/mass spectrometry-based metabolomic analysis showed that ACCM induced alterations in nucleotide and amino acid contents in CSCs.
CeO2NPs adversely compromised the proliferation and structural integrity, and encystation ability of A. castellanii in a dose-dependent manner, they had no significant effect on corneal cells when utilised at doses up to 200 μg/mL. These results reaffirm previous findings and provide new insight into the adverse impact caused by A. castellanii secretome on corneal cells. The promising anti-A. castellanii effect of CeO2NPs warrants further validation including testing their cytotoxic potential against human ocular tissue.
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