Elucidating mechanisms of sulforaphane induced cell stress and death in the human lens

Huynh, Thao (2021) Elucidating mechanisms of sulforaphane induced cell stress and death in the human lens. Doctoral thesis, University of East Anglia.

[thumbnail of 2022HuynhNPTPhD.pdf]
Download (51MB) | Preview


Purpose: Sulforaphane (SFN), derived from cruciferous vegetables, is an emerging therapeutic agent for many health conditions. Recently, SFN-induced cytotoxicity has been shown to have promise in preventing posterior capsule opacification (PCO) - a common lens disorder arising from undesirable cell proliferation and migration. However, no study in any models has elucidated the complete interplay and sequence of events contributing to the cytotoxicity. To address the gap in knowledge, this thesis, therefore, aimed to elucidate key processes and mechanisms linking SFN treatment to cell death in the human lens.

Method: The human lens epithelial cell line FHL124 and central anterior epithelium were used as experimental models. Cell death was assessed by microscopic observation and cell damage/viability assays. Gene or protein levels were assessed by qRT-PCR or Western blot. Mitochondrial networks and DNA damage were assessed by immunofluorescence. Mitochondrial membrane potential, ATF6 activity, GSH/GSSG ratio and glutathione reductase (GR) activity were measured using different light reporter assays. SFN metabolites were analysed by LC-MS/MS.

Results: High concentrations of SFN triggered mitochondrial dysfunction, endoplasmic reticulum stress, DNA damage, autophagy and cell death in both models. The treatment of a ROS scavenger prevented all of these stress responses. SFN also significantly depleted glutathione (GSH), the major antioxidant in the eye, via conjugation since SFN-GSH was detected as the most abundant intracellular SFN metabolite. SFN also reduced GR activity. The supplementation of GSH protected lens cells from all SFN-induced stress responses and death.

Conclusions: SFN depletes GSH levels in human lens epithelial cells through conjugation with GSH and inhibition of GR activity. This leads to ROS generation and oxidative stress that triggers mitochondrial dysfunction, ERS, autophagy and DNA damage leading to cell death. In summary, the work presented provides mechanistic understanding to support the therapeutic application of SFN for PCO and other conditions associated with GSH imbalance.

Item Type: Thesis (Doctoral)
Faculty \ School: Faculty of Science > School of Biological Sciences
Depositing User: Chris White
Date Deposited: 06 Apr 2022 14:54
Last Modified: 06 Apr 2022 14:54
URI: https://ueaeprints.uea.ac.uk/id/eprint/84502


Downloads per month over past year

Actions (login required)

View Item View Item