Abstract
Fyn kinase has emerged as a regulator of diverse pathological processes. However, therapeutic Fyn inhibitors are not available. This study investigated the potential of a series of cycloalkane-fused dithiolethiones (CDTs) or other congeners to increase antioxidant capacity in association with Fyn inhibition, as well as the molecular basis for this effect. Treatment of HepG2 cells with each agent protected the mitochondria from oxidative injury elicited by arachidonic acid and iron, which increased cell viability; 4,5,6,7-tetrahydrobenzo-1,2-dithiole-3-thione (SNU1A) and 5,6-dihydro-4H-cyclopenta-1,2-dithiole-3-thione (SNU2A) were the most effective, whereas 5-methyl-1,2-dithiole-3-thione (SNU3A) was less active. 5-(Quinolin-2-yl)-1,2-dithiole-3-thione (SNU3E) had a minimal effect. SNU1A treatment decreased mitochondrial superoxide production and enabled cells to restore mitochondrial membrane permeability. Oxidative injury caused by arachidonic acid and iron enhanced Fyn phosphorylation at a tyrosine residue, which was decreased by SNU1A treatment. 2,3-Dihydro-N,N-dimethyl-2-oxo-3-[(4,5,6,7-tetrahydro-1H-indol-2-yl)methylene]-1H-indole-5-sulfonamide (SU6656), a known Fyn inhibitor, had a similar effect. Fyn inhibition contributed to protecting mitochondria from injury through AMP-activated protein kinase (AMPK), as supported by reversal of this effect with Fyn overexpression. Consistently, Fyn overexpression attenuated AMPK activation by SNU1A, which strengthens the inhibitory role of Fyn in AMPK activity. CDTs had antioxidant effects, as shown by increases in GSH contents and inhibition of H2O2 production. They also had the ability to activate nuclear factor E2–related factor 2 (Nrf2), a key antioxidant transcription factor. Fyn overexpression decreased the Nrf2 activation induced by SNU1A. Our results demonstrate that CDTs exert cytoprotective effects by protecting mitochondria and increasing the cellular antioxidant capacity, which may result not only from Fyn inhibition leading to AMPK activation but also from Nrf2 activation.
Footnotes
This work was supported by the National Research Foundation of Korea, with grants funded by the Ministry of Education, Science, and Technology [Grant 2011-0001204] and by the World Class University Project [Grant R32-2011-000-10098-0].
Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org.
ABBREVIATIONS:
- ROS
- reactive oxygen species
- AA
- arachidonic acid
- Ad
- adenoviral
- ACC
- acetyl-CoA carboxylase
- AMPK
- AMP-activated protein kinase
- CDT
- cycloalkane-fused dithiolethione
- DCFH-DA
- 2′,7′-dichlorofluorescein diacetate
- FACS
- fluorescence-activated cell sorting
- GSK3β
- glycogen synthase kinase 3β
- LKB1
- liver kinase B1
- MMP
- mitochondrial membrane potential
- MTT
- 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide
- Nrf2
- nuclear factor E2–related factor 2
- Rh123
- Rhodamine 123
- ARE
- antioxidant-response element
- SNU1A
- 4,5,6,7-tetrahydrobenzo-1,2-dithiole-3-thione
- SNU2A
- 5,6-dihydro-4H-cyclopenta-1,2-dithiole-3-thione
- SNU3A
- 5-methyl-1,2-dithiole-3-thione
- SNU3E
- 5-(quinolin-2-yl)-1,2-dithiole-3-thione
- DMEM
- Dulbecco's modified Eagle's medium
- FBS
- fetal bovine serum
- DN
- dominant-negative
- SU6656
- 2,3-dihydro-N,N-dimethyl-2-oxo-3-[(4,5,6,7-tetrahydro-1H-indol-2-yl)methylene]-1H-indole-5-sulfonamide.
- Received December 8, 2011.
- Accepted April 2, 2012.
- Copyright © 2012 The American Society for Pharmacology and Experimental Therapeutics
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