Ding Wen-gang, Zhou Hua-cheng, Cui Xiao-guang, Li Wen-zhi, Guo Yue-ping, Zhang Bing, Liu Wei
Department of Anesthesiology, Second Hospital of Harbin Medical University, Harbin, Heilongjiang 150086, China.
Chin Med J (Engl). 2008 Jul 20;121(14):1313-8.
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.
Cultured porcine PAECs underwent 16-hour anoxia followed by 1-hour reoxygenation 24 hours after pretreatment with saline (NaCl; 0.9%) or morphine (1 micromol/L). To determine the underlying mechanism, a non-selective K(ATP) channel inhibitor glibenclamide (Glib; 10 micromol/L), a nitric oxide (NO) synthase blocker NG-nitro-L-arginine methyl ester (L-NAME; 100 micromol/L), and an opioid receptor antagonist naloxone (Nal; 10 micromol/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.
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) micromol/L vs (20.5 +/- 2.1) micromol/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.
Morphine-induced delayed PC prevents A/R injury of PAECs. This effect may be mediated by activation of K(ATP) channel via opioid receptor and NO signaling pathways.
阿片类药物预处理（PC）可减轻多种细胞的缺氧/复氧（A/R）损伤。然而，阿片类药物诱导的延迟预处理是否会对遭受A/R损伤的肺动脉内皮细胞（PAECs）产生抗凋亡作用仍不清楚。本研究旨在阐明这一问题，并探讨阿片类药物诱导延迟PC的潜在机制。
用生理盐水（NaCl；0.9%）或吗啡（1 μmol/L）预处理培养的猪PAECs 24小时后，进行16小时缺氧，随后再进行1小时复氧。为确定潜在机制，在A/R负荷前30分钟给予非选择性钾离子通道阻滞剂格列本脲（Glib；10 μmol/L）、一氧化氮（NO）合酶阻滞剂NG-硝基-L-精氨酸甲酯（L-NAME；100 μmol/L）和阿片受体拮抗剂纳洛酮（Nal；10 μmol/L）。通过末端脱氧核苷酸转移酶介导的dUTP缺口末端标记（TUNEL）染色评估凋亡细胞百分比。通过实时聚合酶链反应（PCR）测量内皮型一氧化氮合酶（eNOS）mRNA水平。用格里斯试剂测量PAECs上清液中的NO含量。
与A/R处理的PAECs相比，吗啡诱导的延迟PC显著降低了PAECs凋亡（（18.1±1.9）%对（5.5±0.3）%；P<0.05），增加了NO释放（（11.4±1.3）μmol/L对（20.5±2.1）μmol/L，P<0.05），并使eNOS基因表达上调近9倍（P<0.05）。Glib、L-NAME和Nal预处理消除了吗啡的抗凋亡作用，但这三种药物本身并未加重A/R损伤。此外，L-NAME和Nal抵消了吗啡预处理引起的NO释放增加。
吗啡诱导的延迟PC可预防PAECs的A/R损伤。这种作用可能通过阿片受体和NO信号通路激活钾离子通道介导。
