Inflammation and oxidative stress induced by lipid peroxidation metabolite 4-hydroxynonenal in human corneal epithelial cells
- 22.05.2020
- Basic Science
Erschienen in:
- Verfasst von
- Hui Liu
- Frank Gambino Jr
- Crystal S. Algenio
- Charles Wu
- Yichen Gao
- Charles S. Bouchard
- Liang Qiao
- Ping Bu
- Shaozhen Zhao
Abstract
Purpose
Oxidative stress is widely known to be a major contributor in the pathogenesis of dry eye disease (DED). 4-Hydroxynonenal (4-HNE), a well-known byproduct frequently measured as an indicator of oxidative stress-induced lipid peroxidation, has been shown to be elevated in both human and murine corneal DED samples. This study aims to investigate if 4-HNE is responsible for the oxidative stress in human corneal epithelial cells (HCECs) and explores the underlying mechanism by which it confers its effects.
Methods
SV40-immortalized HCECs were cultured in minimum essential media (MEM) with 1% penicillin/streptomycin and 10% fetal bovine serum. HCECs were exposed to media with or without 4-HNE and cell culture supernatants were collected at 4 and 24 h. Cellular reactive oxygen species (ROS) measurement was performed using a 2′,7′-dichlorofluorescein diacetate (DCFDA) assay kit according to the manufacturer’s instructions. Protein levels of antioxidant enzymes copper/zinc superoxide dismutase 1 (SOD1) and NAD(P)H quinone dehydrogenase 1 (NQO1) were analyzed by Western blot. NF-κB activation and expression of IL-6 and IL-8 were measured using an NF-κB p65 Total SimpleStep ELISA Kit and Proteome Profiler Human Cytokine Array Kit. Cell viability was evaluated by LDH cytotoxicity assay.
Results
Treatment with 4-HNE decreased cell viability of HCECs. Band intensities corresponding to levels of ROS production showed a significant increase in ROS generation after treatment with 4-HNE. 4-HNE decreased SOD1 levels and upregulated NQO1 expression in HCECs. A significant increase in activation of NF-κB and production of pro-inflammatory cytokines IL-6 and IL-8 was observed after treatment with 4-HNE. Exposure to N-acetylcysteine (NAC), an antioxidant and ROS scavenger, antagonized the oxidative effects of 4-HNE on HCECs.
Conclusion
4-HNE induces oxidative stress in corneal epithelial cells by increasing levels of ROS generation and modifying the expression of antioxidant enzyme levels, decreasing cell viability of HCECs in vitro. This study demonstrates a potential pathway by which 4-HNE functions to confer its detrimental effects and provides a new therapeutic target for the treatment of DED.
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- Titel
- Inflammation and oxidative stress induced by lipid peroxidation metabolite 4-hydroxynonenal in human corneal epithelial cells
- Verfasst von
-
Hui Liu
Frank Gambino Jr
Crystal S. Algenio
Charles Wu
Yichen Gao
Charles S. Bouchard
Liang Qiao
Ping Bu
Shaozhen Zhao
- Publikationsdatum
- 22.05.2020
- Verlag
- Springer Berlin Heidelberg
- Erschienen in
-
Graefe's Archive for Clinical and Experimental Ophthalmology / Ausgabe 8/2020
Print ISSN: 0721-832X
Elektronische ISSN: 1435-702X - DOI
- https://doi.org/10.1007/s00417-020-04647-2
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