Tetramethylpyrazine Attenuates the Endotheliotoxicity and the Mitochondrial Dysfunction by Doxorubicin via 14-3-3γ/Bcl-2

Figure 1

Schematic representation of the experimental design in vivo and in vitro. In vivo, 75 mice were randomly divided into five different groups: Dox group, mice were routinely fed for 3 weeks; then intraperitoneal injected with six injections of 2.5 mg/kg Dox over 3 weeks for a cumulative dose of 15 mg/kg; TMP + Dox group, mice were administered 6 mg/kg TMP, once daily for 6 weeks via intragastric administration, an hour before Dox administration; TMP + Dox + pAD/14-3-3γ-shRNA group, mice were treated with a regimen similar to the TMP + Dox group for 4 weeks, then injected with pAD/14-3- 3γ-shRNA adenovirus; TMP + Dox + ABT-737 group, mice were treated with a regimen similar to the TMP + Dox group for 5 weeks, followed by a once daily intraperitoneal injection with ABT-737(20 μg/kg), an hour before TMP administration, for 1 week; Control group: mice were given an equal volume of PBS using a regimen similar to the TMP + Dox group. For the in vitro experimental groupings: HUVECs in the control group were cultured under normal conditions (37°C, 95% O₂ and 5% CO₂) over the entire experiment; HUVECs in the Dox group were treated with 1 μM Dox for 48 hours; HUVECs in the TMP + Dox group were treated similar to the Dox group, but cells were also co-incubated with 20 μM TMP for 48 hours, whereas HUVECs in the TMP + Dox + pAD/14-3-3γ-shRNA group were treated with pAD/14-3-3γ-shRNA for 2 hours before TMP treatment; HUVECs in the TMP + Dox + ABT-737 group were treated in a manner similar to the TMP + Dox group, but these cells were also co-incubated with 2 μM ABT-737 for an hour; HUVECs in the TMP + Dox + Atr group were treated in a manner similar to HUVECs in the TMP + Dox group, but were also co-incubated for 2 hours with 50 μM Atr; HUVECs in the Dox + CsA/MitoQ groups were treated in a manner similar to the Dox group, but the cells were also co-incubated with 1 μM CsA/0.5 μM MitoQ for 48 hours, respectively. HUVECs that underwent these treatments were combined to form Phase A-C (See the 2.3.2 in the text).