Objective: Perioperative cardiac complications during cardiovascular surgery remain the major cause of morbidity and mortality. Endothelial-derived exosomes, smallest nanovesicles, increase the survival of ischemic cardiac cells. Even if the ischemic preconditioning is effective in modulating the release of cardioprotective exosomes, drugs that mimic this effect are not known yet. Sevoflurane (SEVO) preconditioning attenuates ischemia and reperfusion (I/R) injury, but the underlying mechanisms are not fully understood. Hypothesis: The preconditioning by SEVO increases the levels of cardioprotective exosomes released by coronary endothelial cells. Methods: Murine coronary endothelial cells (MCEC) were cultured using exosome-depleted fetal bovine serum and then exposed to SEVO (0.35 mM= 1 MAC) for 6 h. Untreated MCEC were used as control. Afterward, cells were exposed to acute I/R injury mimicked by exposure to 1.5 mM hydrogen peroxide (H2O2) for 1 h. Cell viability and anion superoxide (O2-) production were assessed by MTT assay and dihydroethidium staining, respectively. Endothelial levels of peNOS/eNOS, an index of endothelial function, and pSTAT3/STAT3, a transcription factor linked to cardioprotection, were measured by Western blotting (WB). Endothelial exosomes (CD63- and HSP70-positive) were isolated from cell culture media of SEVO cells by serial ultracentrifugation and quantified by WB. In order to assess the exosome mediated cardioprotection, murine cardiomyocytes (HL-1) were treated for 6h with whole or exosomedepleted medium of SEVO-treated MCEC; although, untreated HL-1 were used as control. Then, acute I/R injury was induced by exposure to 1mM H2O2 for 1h. Results: SEVO preconditioning of MCEC significantly prevents the loss of viability induced by acute oxidative stress without affecting O2-, peNOS/eNOS and pSTAT3/STAT3 levels. SEVO significantly increases the release of CD63- and HSP70- positive exosomes compared to untreated cells. After 1h exposure to H2O2, HL-1 survival reduces to 31.76% (p b 0.05). The pre-treatment with whole conditioned medium of SEVO-MCEC increases HL-1 survival to 45.21% (p b 0.05). Conversely, the exosomes-depleted medium of SEVO-MCEC fails in evoking protection and the HL-1 survival decreases to 13.09 % (p b 0.05). Finally, exosomes released by SEVO-MCEC do not contain STAT3. Conclusions: This study shows, for the first time, that SEVO induces the release of CD63- and HSP70-positive exosomes, which protect cultured cardiomyocytes against acute I/R injury. Our results fit into an emerging concept whereby the uptake of HSP70-positive exosomes increases the cellular levels of HSP70, an effector of preconditioning.
|Titolo:||Sevoflurane preconditioning increases the release of cardioprotective exosomes from coronary endothelial cells|
LIONETTI, Vincenzo (Corresponding)
|Data di pubblicazione:||2018|
|Appare nelle tipologie:||1.5 Abstract|