Canna~Fangled Abstracts

Delta9-tetrahydrocannabinol increases C6 glioma cell death produced by oxidative stress.

By October 31, 2013No Comments

pm2Delta9-tetrahydrocannabinol increases C6 glioma cell death produced by oxidative stress.

Source

Department of Neurobiology, Weizmann Institute of Science, Herzel Street, Rehovot 76100, Israel. igor.goncharov@weizmann.ac.il

Abstract

(-)Delta9-tetrahydrocannabinol is a scavenger of free radicals. However, the activation of the CB1 receptor in cultured C6 glioma cells by (-)delta9-tetrahydrocannabinol in the presence of reagents generating reactive oxygen species leads to amplification of the cellular damage from oxidative stress. This was evident by increased loss of cell wall integrity, impaired mitochondrial function and reduction of glucose uptake. In addition, (-)delta9-tetrahydrocannabinol treatment was also found to be deleterious to the cells under conditions of glucose starvation. Free radicals have been implicated in various conditions leading to cell death and, as a routine, the Fenton reaction is utilized for modeling reactive oxygen species production. Our study was performed using a cell permeating Fe(III) chelating quinone that provides more physiological conditions for mimicking the naturally occurring oxidative stress within the cell and thus serves as a better model for natural reactive oxygen species formation.
PMID:

 15975726
[PubMed – indexed for MEDLINE]

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Abbreviations

  • CHO, Chinese hamster ovary;
  • DMPO, 5′,5′-dimethyl-1-pyroline N-oxide;
  • ESR, electron spin resonance;
  • LDH, lactate dehydrogenase;
  • MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide;
  • Qcb, 2-phenyl-4-(butylamino)-naphtholquinoline-7,12-dione;
  • Qn, 2-phenyl-5-nitronaphtho(2,3-g)indole-6,11-dione;
  • ROS, reactive oxygen species;
  • THC, (−)Δ9-tetrahydrocannabinol

Figures and tables from this article:

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Fig. 1.

Effect of short THC treatment on C6 cell viability following oxidative stress. Following 10 min of pretreatment with either 0.5 or 2 μM THC, synthetic quinone Qcb (50 nM) was added and the cells further incubated for 1 h. The cannabinoid antagonists SR141716 and SR144528 were added (where indicated) at final concentration of 2 μM 1 h prior to THC treatment. (A) Medium aliquots were taken every 30 min and assayed for LDH release. (B) MTT viability assay; cells were kept for additional 12 h and assayed for formazan formation. Cell viability in control cells (without THC) is taken as 100%. Values represent the average of three independent experiments ±S.E.M. Levels of significance ranged from **P<0.01 to * P<0.05 compared with control.

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Fig. 2.

Effect of extended THC treatment on C6 cell viability following oxidative stress. Following 72 h with THC at the indicated concentrations, cells were treated with quinone for 1 h and analyzed for (A). LDH release and (B). MTT viability assay (formazan formation). Values represent the average of three independent experiments ±S.E.M. Levels of significance ranged from *** P<0.001 to ** P<0.01 compared with control.

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Fig. 3.

Effect of THC treatment and glucose starvation on C6 cells. Cells were transferred to serum- and glucose-free medium and treated with 0.5 or 2 μM THC. After 36 h the cells were analyzed for: (A). LDH release and (B). MTT assay. No quinone was used in this experiment. Values represent the average of three independent experiments ±S.E.M. Levels of significance ranged from ** P<0.01 to * P<0.05 compared with control.

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Fig. 4.

Effect of THC treatment on LDH release following glucose starvation combined with oxidative stress. Following 36 h of glucose starvation in the presence of THC, at the indicated concentrations, cells were treated with quinone and analyzed for LDH release. Values represent the average of three independent experiments ±S.E.M. Levels of significance ranged from ** P<0.01 to * P<0.05 compared with control.

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Fig. 5.

Effect of THC on glucose uptake in C6 cells. C6 cells were incubated for 24 or 48 h with either 0.5 or 2 μM THC. Untreated cells served as control. The uptake of 3Hdeoxy-glucose is given as percentage of the uptake by control cells and is the mean of three experiments. Values represent the average of three independent experiments ±S.E.M. Levels of significance ranged from ** P<0.01 to * P<0.05 compared with control.

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Fig. 6.

Effect of THC on the interaction of DMPO with ·OH radicals. Generation of ROS was induced by photolysis of H2O2(3%) with white light in the presence of DMPO (0.1 M) and the indicated concentrations of THC. The rate of ROS formation was measured by testing the peak intensity of the second component of DMPO spin adduct. Inset: Representative example of DMPO spin adduct ESR spectrum (obtained after 5 min of H2O2 photolysis). Values represent the average of three independent experiments ±S.E.M. Levels of significance ranged from *** P<0.001 to **P<0.01 compared with control.

Corresponding author contact information
Correspondence to: I. Goncharov, Department of Biological Chemistry, Weizmann Institute of Science, Herzel Street, Rehovot 76100, Israel. Tel: +972-545-767735.

Copyright © 2005 IBRO. Published by Elsevier Ltd. All rights reserved.

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