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目的探讨核因子kappa B(NF-κB)p65的小干扰RNA(siRNA)对延迟性排斥反应的抑制作用。方法血管内皮细胞(EOMA)分为7组:A组(空白对照),B组(阴性对照),C组(TNF-α处理),D组(siRNA处理),E组(scramble siRNA处理),F组(TNF-α+siRNA处理),G组(TNF-α+scramble siRNA处理)。用阳离子脂质体LipofectamineTM2000作为转染试剂将化学合成的小鼠NF-κB p65siRNA转染入相应的EOMA细胞,24h后在C组、F组和G组中加入TNF-α(终浓度为20ng/ml)。于TNF-α加入后的6h提取各组细胞总RNA,RT-PCR法测定EOMA细胞内E选择素(E-selectin)、细胞间黏附分子1(ICAM-1)、白细胞介素1α(IL-1α)和血管细胞黏附分子1(VCAM-1)水平。结果 F组EOMA细胞内E-selectin、ICAM-1、IL-1α和VCAM-1表达明显高于A组,的明显低于C组和G组(P<0.01)。结论特异性siRNA抑制NF-κB p65表达可以有效抑制延迟性排斥反应。
Objective To investigate the inhibitory effect of nuclear factor kappa B (NF-κB) p65 small interfering RNA (siRNA) on delayed rejection. Methods EOMA was divided into 7 groups: group A (blank control), group B (negative control), group C (TNF-α treatment), group D (siRNA treatment), group E (scramble siRNA treatment) Group F (TNF-α + siRNA treatment) and Group G (TNF-α + scramble siRNA treatment). Chemically synthesized mouse NF-κB p65siRNA was transfected into corresponding EOMA cells using lipofectamineTM 2000 as a transfection reagent. After 24 hours, TNF-α was added to groups C, F and G (the final concentration was 20 ng / ml). Total RNA was extracted 6 hours after the addition of TNF-α, and the expression of E-selectin, ICAM-1, IL- 1α) and vascular cell adhesion molecule 1 (VCAM-1) levels. Results The expressions of E-selectin, ICAM-1, IL-1α and VCAM-1 in group E were significantly higher than those in group A and significantly lower than those in group C and group G (P <0.01). Conclusion Specific siRNA can inhibit the expression of NF-κB p65 effectively in delayed rejection.