晚期糖化终产物受体、核因子-κB双基因反义RNA抑制糖化终产物刺激的炎症因子表达

Inhibitory effect of receptor of advanced glycation endproducts,nuclear factor-κB double gene antisense RNA on productions of inflammatory factor treated by advanced glycation endproducts

目的:观察晚期糖化终产物受体(receptor ofadvanced glycation endproducts,RAGE)、核因子-κB(NF-κB)双基因反义RNA对晚期糖化终产物(ad-vanced glycation endproducts,AGEs)刺激ECV304细胞分泌炎症因子的影响。方法:酶联免疫吸附测定(enzyme linked immunosorbent assay,ELISA)法观察AGEs对ECV304细胞分泌TNF-α和IL-6的影响,应用脂质体将RAGE、NF-κB单/双基因反义RNA转染ECV304,流式细胞仪、筛选低表达RAGE、NF-κB的细胞株,观察RAGE、NF-κB双基因反义RNA对AG-Es刺激ECV304细胞分泌TNF-α和IL-6的影响。结果:AGEs可引起ECV304细胞分泌TNF-α和IL-6增加,诱导作用具有时间和剂量依赖规律。稳定转染筛选出低表达RAGE、NF-κB的细胞株,在AGEs100 mg.L-1诱导下双基因转染细胞ECV-asRAGE-asP65克隆中RAGE、NF-κBp65表达抑制率分别为(62.2±8.7)%及(37.2±7.1)%。AGEs 100 mg.L-1刺激下ECV-asRAGE-asP65克隆分泌TNF-αI、L-6较空载体转染细胞、单基因转染细胞减少更明显(P<0.01)。结论:RAGE、NF-κB双基因反义RNA可抑制AGEs刺激的ECV304细胞的TNF-α和IL-6释放。

AIM：To observe the effects of receptor of advanced glycation endproducts （RAGE）, NF-κB double gene antisense RNA on the productions of TNF-α and IL-6 treated by advanced glycation endproducts （AGEs）. METHODS：The levels of TNF-α and IL-6 in the supernatants were measured by enzyme linked immunosorbent assay （ELISA） in ECV304 cells treated by AGEs. The RAGE, NF-κB single/double gene antisense RNA were transfected into ECV304 cell. The expressions of RAGE and NF-κB were detected by flow cytometry and RT-PCR. In different clone cells, the effects of antisense RNA on the productions of TNF-α, IL-6 were detennined by ELISA. RESULTS ： AGEs, instead of human serum albumin （HSA） , stimulated ECV304 cell to produce TNF-α and IL-6 with a time-and dose-dependent manner. The RAGE, NF-κB single/double gene antisense RNA were transfected into ECV304 cell. Induced by AGEs, the expressions of RAGE and NF-κB in double gene cotransfected cell were inhibited by （62.2 ± 8.7） % and （ 39.2 ± 7.1 ）%, respectively. Induced by AGEs, the amount of TNF-α, IL-6 in the medium were lower in single gene transfected cells ECV-asRAGE, ECV-asP65 than ECV-Vector（ P 〈 0.05）. The amount of TNF-α, IL-6 in the double gene transfected cells ECV-asRAGE-asP65 were lower than ECV-Vector and ECV-asRAGE, ECV-asP65 （ P 〈 0.05 ）. CONCLUSION： The production of TNF-α and IL-6 is increased in ECV304 cell treated by AGEs. RAGE, NF-κB double gene antisense RNA can inhibited the production of inflammatory factor treated by AGEs.