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目的探讨不同间歇低氧暴露的内皮细胞和多形核白细胞(PMN)共培养后内皮细胞中核转录因子κB p65(NF-κB p65)的水平变化。方法 18只雄性Wistar大鼠随机分为常氧组和间歇低氧组(IH组)。IH组暴露于间歇低氧环境中,氧浓度波动于5.4%~20.7%,低氧频率为30次/h,8 h/d,6周后处死大鼠,腹主动脉取血,分离纯化PMN。PMN分别与常氧暴露内皮细胞、IH暴露内皮细胞(低氧频率12次/h,共4 h)共培养4 h。细胞按暴露情况分为常氧内皮+常氧PMN组,IH内皮+常氧PMN组,常氧内皮+IHPMN组,IH内皮+IHPMN组。Western blotting测定内皮细胞中NF-κB p65浓度,GAPDH为内参,以NF-κB p65/GAPDH标准化NF-κB p65的表达量。结果 IH内皮+常氧PMN组与常氧内皮+IHPMN组NF-κB p65表达量分别为2.49±0.39和2.14±0.33,两组间差异无统计学意义(P>0.05),但两组均较常氧内皮+常氧PMN组显著升高(0.68±0.20,P<0.05)。IH内皮+IHPMN组NF-κB p65表达水平为4.17±1.48,较其余三组显著升高(P<0.05)。结论间歇低氧可分别致内皮细胞和PMN发生炎症反应,而当两种细胞同时暴露于间歇低氧条件时炎症反应最为严重。上述两种细胞间的相互作用和引发的炎症反应可能是低氧内皮细胞损伤和睡眠呼吸暂停综合征并发心血管合并症的重要原因。
Objective To investigate the changes of nuclear factor κB p65 (NF-κB p65) in endothelial cells co-cultured with different intermittent hypoxic exposure endothelial cells and polymorphonuclear leukocytes (PMN). Methods Eighteen male Wistar rats were randomly divided into normoxia group and intermittent hypoxia group (IH group). IH group was exposed to intermittent hypoxic environment, the concentration of oxygen fluctuated between 5.4% and 20.7%, hypoxemia rate was 30 times / h, 8 h / d, 6 weeks after the rats were sacrificed, blood was taken from the abdominal aorta and PMN was isolated and purified . PMN were co-cultured with normoxia-exposed endothelial cells and IH-exposed endothelial cells (hypoxia frequency 12 times / h for 4 h) respectively for 4 h. The cells were divided into normoxia endothelial + normoxia PMN, IH endothelial + normoxia PMN, normoxia endothelial + IHPMN, IH endothelial + IHMNM according to the exposure. The concentration of NF-κB p65 in endothelial cells was determined by Western blotting. The expression of NF-κB p65 was normalized by NF-κB p65 / GAPDH. Results The expressions of NF-κB p65 in IHEC + normoxia PMN group and normoxia endothelial + IHPMN group were 2.49 ± 0.39 and 2.14 ± 0.33, respectively, with no significant difference between the two groups (P> 0.05) The normoxia endothelial + normoxia PMN group was significantly higher (0.68 ± 0.20, P <0.05). The expression of NF-κB p65 in IH + IHPMN group was 4.17 ± 1.48, which was significantly higher than the other three groups (P <0.05). Conclusions Intermittent hypoxia induced inflammatory responses in endothelial cells and PMNs, respectively, whereas inflammation was the most severe when both cells were exposed to intermittent hypoxic conditions. The interaction between the two kinds of cells and the inflammatory reaction may be the important reason of hypoxia endothelial cell injury and sleep apnea syndrome complicated with cardiovascular comorbidities.