肿瘤坏死因子α、白细胞介素1β对牛肺动脉内皮细胞凋亡的影响及意义

目的　研究肿瘤坏死因子α(TNF α)、白细胞介素 1β(IL 1β)对牛肺动脉内皮细胞 (BPEC)损伤的机制及在急性肺损伤 (ALI)发病过程中的作用。方法　建立BPEC的体外培养 ,采用流式细胞仪膜联蛋白V 异硫氰酸荧光素 (Annexin VFITC)、碘化吡啶 (PI)染色检测TNF α、IL 1β对BPEC凋亡的影响以及抗TNF α单克隆抗体、AC DEVD CHO (caspase 3的竞争性抑制剂 )的保护效应。结果　 (1)TNF α作用 2 4h ,随着其浓度的增加 (浓度为 50 0、10 0 0、2 0 0 0U/ml) ,BPEC凋亡率逐渐增加 [分别为(8 2 1± 0 70 ) %、(9 63± 0 71) %、(17 43± 1 99) % ] ,与对照组 [(3 0 9± 0 0 8) % ]比较差异有显著性(P均 <0 0 5) ;(2 )TNF α (2 0 0 0U/ml)培养时间延长 (分别为 6、12、2 4、3 6h) ,BPEC凋亡率逐渐增加 [分别为 (6 72± 0 3 8) %、(7 72± 1 66) %、(12 95± 0 3 2 ) %、(17 70± 1 79) % ,P均 <0 0 5] ;(3 )加入抗TNF α单抗、AC DEVD CHO的TNF α组的BPEC凋亡率 [(7 78± 0 2 1) %、(7 3 2± 0 11) % ]显著高于单纯TNF α(2 0 0 0U/ml)组 [(10 59± 0 49) % ,P均 <0 0 1] ,而加入IL 1β的TNF α组的凋亡率 [(10 73±0 60 ) % ]与单纯TNF α组比较差异无显著性 (P >0 0 5)。结论　ALI过程中是TNF

Effect of tumor necrosis factor-α and interleukin-1β on bovine pulmonary arterial endothelial cell apoptosis

OBJECTIVE: To investigate the effect of tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) on apoptosis of bovine pulmonary arterial endothelial cells (BPEC). METHODS: BPECs were incubated with TNF-alpha, IL-1beta or the combination of TNF-alpha and IL-1beta. BEPCs were also pretreated with TNF-alpha monoclonal antibody or Ac-DEVD-CHO (caspase-3 inhibitor), and followed by incubation with TNF-alpha. The apoptotic rate was measured by flow cytometry (Annexin V-FITC and PI staining). RESULTS: (1) TNF-alpha induced apoptosis in a dose-dependent manner after a 24 h incubation. With the increase of the TNF-alpha (500, 1,000, 2,000 U/ml respectively), the BPEC apoptotic rate [(8.21 +/- 0.70)%, (9.63 +/- 0.71)%, (17.43 +/- 1.99)%, respectively] was significantly higher than that of the control group [(3.09 +/- 0.08)%, P < 0.05]. (2) TNF-alpha induced apoptosis was time dependent. After incubated with TNF-alpha 2,000 U/ml, the BPEC apoptotic rate [(6.72 +/- 0.38)%, (7.72 +/- 1.66)%, (12.95 +/- 0.32)%, (17.70 +/- 1.79)%, P < 0.05] increased significantly with time [6, 12, 24, 36 h, respectively] of TNF-alpha incubation. (3) The BPEC apoptotic rate of anti-TNF-alpha monoclonal antibody group [(7.78 +/- 0.21)%] or the Ac-DEVD-CHO group [(7.32 +/- 0.11)%] was significantly higher than the TNF-alpha group alone [(10.59 +/- 0.49)%, P < 0.01], but the combination of TNF-alpha and IL-1beta group had no such significant effect (P > 0.05). CONCLUSIONS: It is TNF-alpha but not IL-1beta that induces pulmonary arterial endothelial cell apoptosis in acute lung injury (ALI). There is no synergistic effect between IL-1beta and TNF-alpha in the induction of BPEC apoptosis.