Chowdhury, Lamia Noushin Sadeque
(2024)
Investigating the size-dependent toxicological effects of zinc oxide nanoparticles on Caenorhabditis elegans and the inhibitory potential of Cytochrome P450 isozymes.
MPhil thesis, University of Nottingham.
Abstract
Zinc oxide nanoparticles (ZnONPs) are used in several industries including the cosmetics, textiles, and therapeutics. Wild-type Caenorhabditis elegans (N2) is commonly used as an ecotoxicological model, and as an in vivo model to understand the toxicity of xenobiotics in humans. Cytochrome P540 (CYP) isozymes are responsible for drug metabolism in humans, and its inhibition may lead to adverse drug reactions due to the decreased metabolism and subsequent delayed clearance of co-administered drugs. However, the size-dependent toxicological potential of ZnO particles towards N2 and inhibitory potential towards CYP isozymes are not well known. Firstly, this project aimed to investigate the size-dependent toxicological potential of ZnONP50 (zinc oxide nanoparticles <50 nm), ZnONP100 (zinc oxide nanoparticles <100 nm), and ZnOBP (zinc oxide bulk particle) towards the reproductive capacity, pharyngeal pumping rate, and lifespan of N2. Additionally, gene expression studies were carried to out to assess the variation of the nematode’s cep-1 and pmk-1 genes. Secondly, this project aimed to elucidate the size-dependent in vitro inhibitory effect and mode of inhibition of the ZnO particles towards CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2E1, CYP2J2, and CYP3A5.
Behavioural assays were conducted using N2 to investigate the size-dependent toxicological effect of ZnONP50, ZnONP100, and ZnOBP on the nematode’s reproductive capacity, pharyngeal pumping rate, and lifespan. A gene expression study using conventional PCR was performed to assess the variation in the expression of the cep-1 and pmk-1 genes to elicit the plausible mechanism of ZnO particles’ toxicity. Inhibitory potential of ZnONP50, ZnONP100, and ZnOBP on CYP isozymes was investigated using in vitro fluorescence enzymatic assays. Half maximal inhibitory concentration (IC50), IC50-shift, inhibitor constant (Ki), and the mode of inhibition were determined.
ZnONP50 exhibited the strongest toxicological effect in terms of N2’s lifespan, while the nematode’s reproduction capacity and pharyngeal pumping rate was negatively altered the most by ZnONP100. Based on the gene expression study, ZnONP50 upregulated the expression of cep-1 and pmk-1 when compared to ZnONP100 and ZnOBP. ZnO particles were able to inhibit the CYP isozymes with various potencies in various reversible inhibition modes. ZnONP50 was able to significantly inhibit CYP2B6 (Ki:20.33 g/mL, mixed-mode), CYP2C8 (Ki:43.33 g/mL, competitive-mode), CYP2E1 (Ki: 16.63 g/mL, mixed-mode), CYP2J2 (Ki: 8.18 g/mL, non-competitive-mode), and CYP3A5 (Ki: 2.54 g/mL, non-competitive mode). ZnONP100 inhibited CYP2B6 (Ki: 6.67 g/mL, mixed-mode), CYP2E1 (Ki: 40.1 g/mL, mixed-mode), CYP2J2 (Ki: 20.0 g/mL, mixed/non-competitive-mode), and CYP3A5’s (Ki: 0.5 g/mL, non-competitive-mode) enzymatic activity. ZnOBP inhibited the enzymatic activity of CYP2E1 (Ki: 28.3 g/mL, competitive-mode). Size-dependent toxicity demonstrated by ZnO particles towards N2, and the targeted genes were investigated. Based on the gene expression study results, it is suggested that ZnONPs likely elicited its toxicological effects through the production of ROS and the subsequently generated oxidative stress.
Size-dependent inhibition was exhibited towards CYP2B6, CYP2J2, and CYP2E1. Based on the results of this project, future studies are recommended to quantify the production of ROS and the subsequent oxidative stress production to improve the understanding of ZnONPs’ toxicological impact in N2 and in humans. Additionally, in silico studies using CYP isozymes are required to improve the understanding of the mechanism of ZnONPs’ inhibitory potential.
Actions (Archive Staff Only)
 |
Edit View |
Tools
Tools
