Abstract
Long-term heat acclimation (AC, 30d/34°C) is a phenotypic adaptation leading to increased thermotolerance during heat stress (HS, 2 h 41°C). AC also renders protection against ischemic/reperfusion (I/R, 30′ global ischemia/40′ reperfusion) insult via cross-tolerance mechanisms. In contrast to the protected AC phenotype, the onset of acclimation (34°C, AC2d) is characterized by cellular perturbations, suggesting increased susceptibility to HS and I/R insults. In this investigation, we tested the hypothesis that apoptosis resistance is part of the AC repertoire and that, at the initial phase of acclimation (AC2d), cytoprotection is impaired. TUNEL staining and caspase 3 levels in HS and I/R insulted hearts affirmed this hypothesis. To examine the role of the mitochondria in life/death decision in AC2d and 30d AC settings vs. control hearts, we studied the Bcl-2 apoptotic cascade and found increased levels of the anti-apoptotic Bcl-XL and decreased levels of the pro-apoptotic death promoter Bad in hearts from AC2d and AC animals. In these groups, cytochrome c (cyt c) was elevated in the mitochondria and remained unchanged in the cytosol. This adaptation was insufficient to negate apoptosis in AC2d rats. At this early acclimation phase (and in controls), increased caspase 8 activity confirmed activation of the extrinsic (Fas ligand) apoptosis pathway. In conclusion, the elevated Bcl-XL/Bad ratio and decreased cyt c leakage to the cytosol are insufficient to protect the heart and interactions with additional cytoprotective pathways involved in acclimation (elevated HSP70, ROS, and sarcolemmal adaptations to abolish extrinsic apoptosis pathways) are required to induce the apoptosis-resistant AC phenotype.
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Acknowledgement
This study was supported by the USA–Israel Binational Fund BSF Grant 2003-298 and (in part) by the Intramural Research Program of the NIH National Institute on Aging.
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Fig. 1
Bad and Bcl-XL protein expression in the mitochondrial fraction following recovery from HS in the C and AC groups. Bad expression did not change significantly between the groups. Bcl-XL was increased post-HS in the AC group and was significantly different from the C at three time points: HS+1, HS+1.5, and HS+24. Asterisk indicates significant difference from Con (P < 0.01–0.002), Tukey–Kremer (PPT 148 kb)
Fig. 2
Cytochrome c mRNA levels following HS. The only significant difference was immediately post-HS, when there was an up-regulation in the AC group and down-regulation in the C group. * - significant difference from Con (P < 0.024), Tukey-Kremer (PPT 127 kb)
Fig. 3
Caspase 9 mRNA levels (qRT-PCR) prior to and at selected time points during recovery from HS. Values are means±SE; mRNA levels were normalized to β-actin. Procaspase 9 was decreased in AC, as compared with C, hearts (2W ANOVA P < 0.003). In contrast, caspase 9 transcript was up-regulated post-HS in C but not in AC groups (2W ANOVA P < 0.001). Values are means±SE; asterisk indicates significant difference from C (P < 0.03–0.001) (PPT 126 kb)
Fig. 4
Caspase 9 and caspase 3 transcripts in left ventricle of the hearts of C, AC2d, and AC rats before and following I/R insult. Basal caspase 9 transcript in AC was significantly lower than that of C and AC2d. Basal caspase 3 transcripts (except for slight up-regulation on day 2 of the acclimation) did not differ significantly in all groups. I/R insult induced minor changes only in caspase 9 transcripts but not in caspase 3. Asterisk indicates significant difference from C (P < 0.003); circumflex accent indicates significant difference from basal (B) within the group (P < 0.002); number sign indicates significant difference from AC (P < 0.03; Tukey–Kramer) (PPT 200 kb)
Fig. 5
Caspase 8 transcripts in left ventricle of C, AC2d, and AC rats following I/R insult vs. basal normoxic C hearts. AC2d caspase 8 level was significantly higher than that of C and AC I/R heart, as well as vs. basal normoxic levels. Basal normoxic transcript levels did not differ significantly among the groups. Asterisk indicates significant difference from basal (P < 0.005); number sign indicates significant difference from C and AC (P < 0.02–0.04), 1W ANOVA (P < 0.005) followed by Tukey test (PPT 119 kb)
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Assayag, M., Gerstenblith, G., Stern, M.D. et al. Long- but not short-term heat acclimation produces an apoptosis-resistant cardiac phenotype: a lesson from heat stress and ischemic/reperfusion insults. Cell Stress and Chaperones 15, 651–664 (2010). https://doi.org/10.1007/s12192-010-0178-x
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DOI: https://doi.org/10.1007/s12192-010-0178-x