核因子kappaB的反义核酸对人内皮细胞间粘附分子-1表达的影响

Effect of antisense oligonucleotides of nuclear factor kappa B on expression of adhesion molecules intercellular adhesion molecule-1 in human vascular endothelial cells

目的 采用反义核酸技术观察核因子kappaB(NF κB)的合成及其对所调控的粘附分子表达的影响 ,阐明其作用机制并证明其有效性。方法 设立正义、反义核酸组和对照组 (单用肿瘤坏死因子 α刺激 )。观察FITC标记的寡核苷酸在内皮细胞内的摄取情况及采用流式细胞仪检测反义核酸对NF κB表达的影响 ;采用RT PCR、免疫组织化学法和流式细胞仪检测转染后细胞间粘附分子 (ICAM ) 1的表达情况。结果 反义核酸组NF κB的表达较对照组下降 35 .94%,较正义核酸组下降 47.14%(P <0 .0 5 ) ;反义核酸组人血管内皮细胞I CAM 1的mRNA表达及其在细胞表面的表达水平较对照组分别降低 12 .2 3%和 71.37%,较正义核酸组分别降低 18.16 %和 71.82 %(P <0 .0 5 )。结论 NF κB的反义核酸能有效降低NF κB的复制及合成 ,从而显著下调人血管内皮细胞ICAM 1的表达 ,其正义核酸无此效应。

Objective To investigate the effects of antisense oligonucleotides of nuclear factor kappa B(NF-κB) on TNF-α induced expression of NF-κB and intercellular adhesion molecule-1 (ICAM-1) in vitro. Methods The effects of antisense oligonucleotide of NF-κB and intracellular uptake of FITC-labeled oligonucleotides were detected by fluorescence active cell sort (FACS). Adhesion molecules expression was presented by RT-PCR at mRNA level, FACS on surfaces of human vascular endothelial cells (HUVECs) and on morphology by immunohistology. Results Expression of NF-κB in HUVECs transfected antisense ODNs of NF-κB reduced 35.94% comparing with expression of NF-κB in control cells stimulated with TNF-α only, and reduced 47.14% when compared with expression of NF-κB in sense ODNs of NF-κB transfected cells. ICAM-1 mRNA expression level of antisense group reduced 12.23% and 71.37%, on HUVECs surface comparing with that of the control group, and the sense group. Conclusion These observations demonstrate that antisense NF-κB can inhibit replication and synthesis of NF-κB and down-regulate ICAM-1 expression.