The intracellular oxidation of 2′,7′-dichlorofluorescin in murine T lymphocytes

Van Reykl, DM; King, NJC; Dinauer, MC; Hunt, NH

The intracellular oxidation of 2′,7′-dichlorofluorescin in murine T lymphocytes

Van Reykl, DM

King, NJC Dinauer, MC Hunt, NH

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Publication Type:: Journal Article
Citation:: Free Radical Biology and Medicine, 2001, 30 (1), pp. 82 - 88
Issue Date:: 2001-01-01

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Field	Value	Language
dc.contributor.author	Van Reykl, DM https://orcid.org/0000-0002-7768-8662	en_US
dc.contributor.author	King, NJC	en_US
dc.contributor.author	Dinauer, MC	en_US
dc.contributor.author	Hunt, NH	en_US
dc.date.issued	2001-01-01	en_US
dc.identifier.citation	Free Radical Biology and Medicine, 2001, 30 (1), pp. 82 - 88	en_US
dc.identifier.issn	0891-5849	en_US
dc.identifier.uri	http://hdl.handle.net/10453/3218
dc.description.abstract	Intracellular reactive oxygen species (ROS) production by activated murine T lymphocytes was investigated by analyzing intracellular dichlorofluorescin (DCFH2) oxidation in lymph node cells (LNC). An increase in DCFH2 oxidation in LNC induced by phorbol myristate acetate (PMA) was detected by flow cytometry. It was confirmed that this increase was present in Thy1+ LNC. We examined the contribution to intracellular DCFH2 oxidation of ROS released by leukocytes other than T cells present in the LNC suspension. Superoxide dismutase, catalase, and glutathione/glutathione peroxidase inhibited the PMA-induced increase in intracellular DCFH2 oxidation. Furthermore, PMA failed to elicit DCFH2 oxidation in LNC isolated from mice lacking a functional NADPH oxidase (gp91phox gene knockout mice), but this response could be restored in these cells by the addition of T cell-depleted LNC from wild-type litter mates. This study highlights the necessity for caution in using the DCFH2 assay to demonstrate specific intracellular ROS production in heterogeneous cell populations. It also suggests that cells other than T cells in lymph node populations may, through production of ROS, influence the intracellular redox state of T lymphocytes. © 2000 Elsevier Science Inc.	en_US
dc.relation.ispartof	Free Radical Biology and Medicine	en_US
dc.relation.isbasedon	10.1016/S0891-5849(00)00449-4	en_US
dc.subject.classification	Biochemistry & Molecular Biology	en_US
dc.subject.mesh	Lymph Nodes	en_US
dc.subject.mesh	T-Lymphocytes	en_US
dc.subject.mesh	Animals	en_US
dc.subject.mesh	Mice, Inbred BALB C	en_US
dc.subject.mesh	Mice, Inbred C57BL	en_US
dc.subject.mesh	Mice, Knockout	en_US
dc.subject.mesh	Mice	en_US
dc.subject.mesh	Reactive Oxygen Species	en_US
dc.subject.mesh	Tetradecanoylphorbol Acetate	en_US
dc.subject.mesh	Fluoresceins	en_US
dc.subject.mesh	Catalase	en_US
dc.subject.mesh	Glutathione	en_US
dc.subject.mesh	Glutathione Peroxidase	en_US
dc.subject.mesh	NADPH Oxidases	en_US
dc.subject.mesh	Oxidation-Reduction	en_US
dc.subject.mesh	Superoxide Dismutase	en_US
dc.title	The intracellular oxidation of 2′,7′-dichlorofluorescin in murine T lymphocytes	en_US
dc.type	Journal Article
utslib.citation.volume	1	en_US
utslib.citation.volume	30	en_US
utslib.for	110106 Medical Biochemistry: Proteins and Peptides (incl. Medical Proteomics)	en_US
utslib.for	0304 Medicinal and Biomolecular Chemistry	en_US
utslib.for	0601 Biochemistry and Cell Biology	en_US
utslib.for	1101 Medical Biochemistry and Metabolomics	en_US
pubs.embargo.period	Not known	en_US
pubs.organisational-group	/University of Technology Sydney
pubs.organisational-group	/University of Technology Sydney/Faculty of Science
pubs.organisational-group	/University of Technology Sydney/Faculty of Science/School of Life Sciences
utslib.copyright.status	closed_access
pubs.issue	1	en_US
pubs.publication-status	Published	en_US
pubs.volume	30	en_US

Abstract:

Intracellular reactive oxygen species (ROS) production by activated murine T lymphocytes was investigated by analyzing intracellular dichlorofluorescin (DCFH2) oxidation in lymph node cells (LNC). An increase in DCFH2 oxidation in LNC induced by phorbol myristate acetate (PMA) was detected by flow cytometry. It was confirmed that this increase was present in Thy1+ LNC. We examined the contribution to intracellular DCFH2 oxidation of ROS released by leukocytes other than T cells present in the LNC suspension. Superoxide dismutase, catalase, and glutathione/glutathione peroxidase inhibited the PMA-induced increase in intracellular DCFH2 oxidation. Furthermore, PMA failed to elicit DCFH2 oxidation in LNC isolated from mice lacking a functional NADPH oxidase (gp91phox gene knockout mice), but this response could be restored in these cells by the addition of T cell-depleted LNC from wild-type litter mates. This study highlights the necessity for caution in using the DCFH2 assay to demonstrate specific intracellular ROS production in heterogeneous cell populations. It also suggests that cells other than T cells in lymph node populations may, through production of ROS, influence the intracellular redox state of T lymphocytes. © 2000 Elsevier Science Inc.

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http://hdl.handle.net/10453/3218