Abstract
The present study was designed to investigate ex vivo the protective mechanisms of heat-shock response against H2O2-induced oxidative stress in peripheral blood mononuclear cells (PBMCs) of rats. Twenty-four hours later, heat-shock treatment was executed in vivo; rat PBMCs were collected and treated with H2O2. The accumulation of reactive oxygen species and the mitochondrial membrane potential were evaluated by intracellular fluorescent dHE and JC-1 dye staining, respectively, and expression of HSP72 and cytochrome c was detected by Western blot analysis. Cellular apoptosis was assayed by TUNEL staining and double staining of Annexin V and PI. The results showed that H2O2-induced oxidative stress leads to intracellular superoxide accumulation and collapse of the mitochondrial membrane potential in rat PBMCs. Moreover, cellular apoptosis was detected after H2O2 treatment, and the release of mitochondrial cytochrome c from mitochondria to cytosol was significantly enhanced. Heat-shock pretreatment decreases the accumulation of intracellular superoxide in PBMCs during H2O2-induced oxidative stress. Moreover, heat-shock treatment prevents the collapse of the mitochondrial membrane potential and cytochrome c release from mitochondria during H2O2-induced oxidative stress. In conclusion, mitochondria are critical organelles of the protective effects of heat-shock treatment. Cellular apoptosis during H2O2-induced oxidative stress is decreased by heat-shock treatment through a decrease in superoxide induction and preservation of the mitochondrial membrane potential.
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Abbreviations
- DAB:
-
diaminobenzidine
- dHE:
-
dihydroethidium
- HSP:
-
heat-shock protein
- Δψ:
-
mitochondrial membrane potential
- PBMC:
-
peripheral blood mononuclear cell
- ROS:
-
Reactive oxygen species
- PI:
-
propidium iodium
- TUNEL:
-
TdT-mediated dUTP nick end-labeling assay
- TdT:
-
terminal deoxynucleotidyl transferase
- JC-1:
-
5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazol-carbocyanine iodide
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Chiu, HY., Tsao, LY. & Yang, RC. Heat-shock response protects peripheral blood mononuclear cells (PBMCs) from hydrogen peroxide-induced mitochondrial disturbance. Cell Stress and Chaperones 14, 207–217 (2009). https://doi.org/10.1007/s12192-008-0075-8
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DOI: https://doi.org/10.1007/s12192-008-0075-8