Anti-apoptotic effect of morphine-induced delayed preconditioning on pulmonary artery endothelial cells with anoxia/reoxygenation injury

Background Opioid preconditioning （PC） reduces anoxia/reoxygenation （A/R） injury to various cells. However, it remains unclear whether opioid-induced delayed PC would show anti-apoptotic effects on pulmonary artery endothelial cells （PAECs） suffering from A/R injury. The present study was conducted to elucidate this issue and to investigate the potential mechanism of opioid-induced delayed PC. Methods Cultured porcine PAECs underwent 16-hour anoxia followed by 1-hour reoxygenation 24 hours after pretreatment with saline （NaCI; 0.9%） or morphine （1 μmol/L）. To determine the underlying mechanism, a non-selective KATe channel inhibitor glibenclamide （Glib; 10 μmol/L）, a nitric oxide （NO） synthase blocker NG-nitro-L-arginine methyl ester （L-NAME; 100 μmol/L）, and an opioid receptor antagonist naloxone （Nal; 10μmol/L） were given 30 minutes before the A/R load. The percentage of apoptotic cells was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling （TUNEL） staining, eNOS mRNA level was measured by real-time polymerase chain reaction （PCR）. NO content of PAECs supernatants was measured with the Griess reagent. Results Compared to the A/R PAECs, morphine-induced delayed PC significantly reduced PAECs apoptosis （（18.1±1.9）% vs （5.5±0.3）%; P 〈0.05）, increased NO release （（11.4±1.3） μmol/L vs （20.5±2.1） μmol/L, P 〈0.05）, and up-regulated eNOS gene expression nearly 9 times （P 〈0.05）. The anti-apoptosis effect of morphine was abolished by pretreatment with Glib, L-NAME and Nal, but the three agent-selves did not aggravate the A/R injury. Furthermore, L-NAME and Nal offset the enhanced release of NO caused by pretreatment with morphine. Conclusions Morphine-induced delayed PC prevents A/R injury of PAECs. This effect may be mediated by activation of KATe channel via opioid receptor and NO signaling pathways.