Single-cell redox states analyzed by fluorescence lifetime metrics and tryptophan FRET interaction with NAD(P)H.
Cytosol
/ drug effects
Doxorubicin
/ pharmacology
Energy Metabolism
/ drug effects
Flavin-Adenine Dinucleotide
/ analysis
Fluorescence
Fluorescence Resonance Energy Transfer
/ methods
HeLa Cells
Humans
Microscopy, Fluorescence, Multiphoton
/ methods
Mitochondria
/ drug effects
NAD
/ analysis
NADP
/ analysis
Optical Imaging
Oxidation-Reduction
Oxidative Phosphorylation
/ drug effects
Reactive Oxygen Species
/ metabolism
Single-Cell Analysis
/ methods
Tryptophan
/ chemistry
FAD
Fluorescence Lifetime Imaging Microscopy (FLIM)
NAD(P)H
NAD(P)H-a2%, redox: reduction/oxidation states
NADPH/NADH ratio
fluorescence lifetime redox ratio (FLIRR)
single-cell analysis
Journal
Cytometry. Part A : the journal of the International Society for Analytical Cytology
ISSN: 1552-4930
Titre abrégé: Cytometry A
Pays: United States
ID NLM: 101235694
Informations de publication
Date de publication:
01 2019
01 2019
Historique:
received:
23
10
2018
revised:
12
12
2018
accepted:
18
12
2018
pubmed:
4
1
2019
medline:
13
3
2020
entrez:
4
1
2019
Statut:
ppublish
Résumé
Redox changes in live HeLa cervical cancer cells after doxorubicin treatment can either be analyzed by a novel fluorescence lifetime microscopy (FLIM)-based redox ratio NAD(P)H-a2%/FAD-a1%, called fluorescence lifetime redox ratio or one of its components (NAD(P)H-a2%), which is actually driving that ratio and offering a simpler and alternative metric and are both compared. Auto-fluorescent NAD(P)H, FAD lifetime is acquired by 2- photon excitation and Tryptophan by 3-photon, at 4 time points after treatment up to 60 min demonstrating early drug response to doxorubicin. Identical Fields-of-view (FoV) at each interval allows single-cell analysis, showing heterogeneous responses to treatment, largely based on their initial control redox state. Based on a discrete ROI selection method, mitochondrial OXPHOS and cytosolic glycolysis are discriminated. Furthermore, putative FRET interaction and energy transfer between tryptophan residue carrying enzymes and NAD(P)H correlate with NAD(P)H-a2%, as does the NADPH/NADH ratio, highlighting a multi-parametric assay to track metabolic changes in live specimens. © 2019 International Society for Advancement of Cytometry.
Identifiants
pubmed: 30604477
doi: 10.1002/cyto.a.23711
doi:
Substances chimiques
Reactive Oxygen Species
0
NAD
0U46U6E8UK
Flavin-Adenine Dinucleotide
146-14-5
NADP
53-59-8
Doxorubicin
80168379AG
Tryptophan
8DUH1N11BX
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
110-121Informations de copyright
© 2019 International Society for Advancement of Cytometry.