Oxidative stress leads to the formation of esterified erythro- and threo-dihydroxy-fatty acids in HepG2 cells
Oxidative stress plays a central role in pathophysiology. To assess oxidative stress, sensitive methods are required to monitor the cellular damage caused by reactive oxygen species. Phospholipid-bound isoprostanes, such as 5-iPF2α-VI determined by liquid chromatography-tandem mass spectrometry (LC-...
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Elsevier
2025-05-01
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| Series: | Redox Biology |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2213231725001028 |
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| author | Lilli Scholz Luca M. Wende Michel A. Chromik Nadja Kampschulte Nils Helge Schebb |
| author_facet | Lilli Scholz Luca M. Wende Michel A. Chromik Nadja Kampschulte Nils Helge Schebb |
| author_sort | Lilli Scholz |
| collection | DOAJ |
| description | Oxidative stress plays a central role in pathophysiology. To assess oxidative stress, sensitive methods are required to monitor the cellular damage caused by reactive oxygen species. Phospholipid-bound isoprostanes, such as 5-iPF2α-VI determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS) are currently the best markers for oxidative stress. Here, we describe erythro- and threo-dihydroxy-polyunsaturated fatty acids (PUFA), the hydrolysis products of trans- and cis-epoxy-PUFA, as new biomarkers of oxidative stress in cells. This is demonstrated in four oxidative stress models in HepG2 cells using radical-forming tert-butyl hydroperoxide, glutathione peroxidase 4 inhibiting RSL-3, the redox cycling agent paraquat or rotenone blocking the electron transport chain. LC-MS/MS analysis following the liberation of esterified oxylipins by saponification unveiled in all oxidative stress models a strong formation of erythro- and threo-dihydroxy-PUFA. The levels increased concentration-dependently and correlated to isoprostane 5-iPF2α-VI formation. Among the positional isomers derived from linoleic acid, arachidonic acid and docosahexaenoic acid, those bearing the dihydroxy-group closest to the carboxy terminus were predominantly formed. Thus, the highest concentrations were found of erythro-5,6-DiHETrE and erythro-4,5-DiHDPE, which allowed a more sensitive detection of oxidative stress compared to 5-iPF2α-VI levels. The (erythro-) dihydroxy-PUFA are a new set of markers, which enable a more comprehensive analysis of oxidative stress, particularly when combined with simultaneous LC-MS/MS quantification of trans-epoxy-PUFA and isoprostanes. |
| format | Article |
| id | doaj-art-8fbf64b98227480dae0da0353f2afc46 |
| institution | DOAJ |
| issn | 2213-2317 |
| language | English |
| publishDate | 2025-05-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Redox Biology |
| spelling | doaj-art-8fbf64b98227480dae0da0353f2afc462025-08-20T03:14:52ZengElsevierRedox Biology2213-23172025-05-018210358910.1016/j.redox.2025.103589Oxidative stress leads to the formation of esterified erythro- and threo-dihydroxy-fatty acids in HepG2 cellsLilli Scholz0Luca M. Wende1Michel A. Chromik2Nadja Kampschulte3Nils Helge Schebb4Chair of Food Chemistry, Faculty of Mathematics and Natural Sciences, University of Wuppertal, Gaussstrasse 20, 42119, Wuppertal, GermanyChair of Food Chemistry, Faculty of Mathematics and Natural Sciences, University of Wuppertal, Gaussstrasse 20, 42119, Wuppertal, GermanyChair of Food Chemistry, Faculty of Mathematics and Natural Sciences, University of Wuppertal, Gaussstrasse 20, 42119, Wuppertal, GermanyChair of Food Chemistry, Faculty of Mathematics and Natural Sciences, University of Wuppertal, Gaussstrasse 20, 42119, Wuppertal, GermanyCorresponding author.; Chair of Food Chemistry, Faculty of Mathematics and Natural Sciences, University of Wuppertal, Gaussstrasse 20, 42119, Wuppertal, GermanyOxidative stress plays a central role in pathophysiology. To assess oxidative stress, sensitive methods are required to monitor the cellular damage caused by reactive oxygen species. Phospholipid-bound isoprostanes, such as 5-iPF2α-VI determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS) are currently the best markers for oxidative stress. Here, we describe erythro- and threo-dihydroxy-polyunsaturated fatty acids (PUFA), the hydrolysis products of trans- and cis-epoxy-PUFA, as new biomarkers of oxidative stress in cells. This is demonstrated in four oxidative stress models in HepG2 cells using radical-forming tert-butyl hydroperoxide, glutathione peroxidase 4 inhibiting RSL-3, the redox cycling agent paraquat or rotenone blocking the electron transport chain. LC-MS/MS analysis following the liberation of esterified oxylipins by saponification unveiled in all oxidative stress models a strong formation of erythro- and threo-dihydroxy-PUFA. The levels increased concentration-dependently and correlated to isoprostane 5-iPF2α-VI formation. Among the positional isomers derived from linoleic acid, arachidonic acid and docosahexaenoic acid, those bearing the dihydroxy-group closest to the carboxy terminus were predominantly formed. Thus, the highest concentrations were found of erythro-5,6-DiHETrE and erythro-4,5-DiHDPE, which allowed a more sensitive detection of oxidative stress compared to 5-iPF2α-VI levels. The (erythro-) dihydroxy-PUFA are a new set of markers, which enable a more comprehensive analysis of oxidative stress, particularly when combined with simultaneous LC-MS/MS quantification of trans-epoxy-PUFA and isoprostanes.http://www.sciencedirect.com/science/article/pii/S2213231725001028Liquid chromatographyTandem mass spectrometryOxylipinsIsoprostaneEicosanoids |
| spellingShingle | Lilli Scholz Luca M. Wende Michel A. Chromik Nadja Kampschulte Nils Helge Schebb Oxidative stress leads to the formation of esterified erythro- and threo-dihydroxy-fatty acids in HepG2 cells Redox Biology Liquid chromatography Tandem mass spectrometry Oxylipins Isoprostane Eicosanoids |
| title | Oxidative stress leads to the formation of esterified erythro- and threo-dihydroxy-fatty acids in HepG2 cells |
| title_full | Oxidative stress leads to the formation of esterified erythro- and threo-dihydroxy-fatty acids in HepG2 cells |
| title_fullStr | Oxidative stress leads to the formation of esterified erythro- and threo-dihydroxy-fatty acids in HepG2 cells |
| title_full_unstemmed | Oxidative stress leads to the formation of esterified erythro- and threo-dihydroxy-fatty acids in HepG2 cells |
| title_short | Oxidative stress leads to the formation of esterified erythro- and threo-dihydroxy-fatty acids in HepG2 cells |
| title_sort | oxidative stress leads to the formation of esterified erythro and threo dihydroxy fatty acids in hepg2 cells |
| topic | Liquid chromatography Tandem mass spectrometry Oxylipins Isoprostane Eicosanoids |
| url | http://www.sciencedirect.com/science/article/pii/S2213231725001028 |
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