论文部分内容阅读
目的 :探讨脂多糖 ( LPS)对培养的牛肺动脉内皮细胞 ( PAEC) 型 Na+ /H+交换器 ( NHE-1)表达的影响及其与内皮细胞死亡的关系。方法 :体外培养牛 PAEC,MTT比色检测细胞存活率 ,并提取总 RNA,应用反转录聚合酶链式反应 ( RT-PCR)及 DNA电泳技术 ,以 NHE-1与β-actin电泳条带光密度值之比 ( e/c)作数据分析 ,以其比值反映 NHE-1m RNA的表达程度。应用 Fluo3 /Amy荧光探针标记 ,激光共聚焦观察细胞内钙离子的浓度。结果 :在L PS( 0 .0 1～ 1.0 μg/ml)作用 48h后 ,MTT比色吸光度值分别为 0 .3 5± 0 .0 7,0 .3 1± 0 .0 7,0 .2 6± 0 .0 4,低于对照组 0 .44± 0 .0 6( P<0 .0 1)。e/c分别为 0 .43± 0 .11,0 .83± 0 .14 ,1.2 2± 0 .19,高于对照组 0 .2 8± 0 .0 8( P<0 .0 1,n=4)。激光共聚焦结果显示 L PS引起细胞内钙离子增多 ,呈剂量依赖型。结论 :LPS可以引起肺动脉内皮细胞NHE-1表达增加 ,抑制细胞增殖 ,导致细胞死亡 ,钙离子可能参与 NHE-1的激活及表达。
Objective: To investigate the effect of lipopolysaccharide (LPS) on the expression of Na + / H + exchanger (NHE-1) in cultured bovine pulmonary artery endothelial cells (PAEC) and its relationship with endothelial cell death. Methods: The bovine PAEC was cultured in vitro. The cell viability was detected by MTT colorimetric assay. The total RNA was extracted. Reverse transcription-polymerase chain reaction (RT-PCR) and DNA electrophoresis were used to detect the cell viability of NHE-1 and β-actin bands The ratio of optical density values (e / c) was used for data analysis to reflect the expression level of NHE-1 mRNA by its ratio. Fluo3 / Amy fluorescent probe labeling, laser confocal observation of intracellular calcium concentration. RESULTS: MTT colorimetric absorbance values were 0 .35 ± 0 .0 7,0 .31 ± 0 .0 7,0 .2 at LPS (0.01-1.0 μg / ml) for 48 h 6 ± 0. 04, lower than the control group 0. 44 ± 0. 0 6 (P <0. e / c were 0.43 ± 0.11,0. 83 ± 0.14, 1.2 2 ± 0.19, respectively, which were higher than that of the control group 0.828 ± 0.08 (P <0.01, n = 4). Laser confocal results showed that L PS caused an increase of intracellular calcium, in a dose-dependent manner. CONCLUSION: LPS can induce the increase of NHE-1 in pulmonary artery endothelial cells, inhibit cell proliferation and cause cell death, and calcium ion may be involved in the activation and expression of NHE-1.