红霉素上调组蛋白去乙酰化酶2表达减轻氧化应激下THP-1细胞的糖皮质激素抵抗

Erythromycin restores oxidative stress-induced corticosteroid responsiveness of human THP-1 cells by up-regulating the expression of histone deacetylase 2

目的探讨红霉素(EM)对氧化应激下人THP-1单核细胞糖皮质激素抵抗的作用及机制。方法 EM预孵育THP-1细胞后予烟草烟雾提取物(CSE)刺激细胞,用质粒脂质体法将构建好的组蛋白去乙酰化酶2沉默RNA(HDAC2-siRNA)转染细胞,ELISA检测细胞培养上清白细胞介素8(IL-8)的含量,实时荧光定量PCR(qRT-PCR)和Western blot法分析HDAC2表达情况。结果 EM组的IL-8抑制率明显高于CSE组,而低于对照组(P<0.05),EM组地塞米松半数抑制浓度(IC50-Dex)低于CSE组,而高于对照组(P<0.05)。EM组HDAC2蛋白的表达高于CSE组,而低于对照组(P<0.05)。除此之外,应用HDAC2-siRNA转染细胞后HDAC2 mRNA及HDAC2蛋白的表达均明显低于对照组及空转组,而加用EM后其表达明显升高(P<0.05)。结论 CSE刺激下的THP-1细胞对地塞米松的敏感性明显降低,EM可通过上调HDAC2蛋白的表达减轻氧化应激下THP-1细胞的糖皮质激素抵抗。

Objective To investigate the effect of erythromycin （EM） on corticosteroid insensitivity of human THP-1 cells induced by cigarette smoke extract （CSE） and its mechanism. Methods THP-! cells were treated with EM followed by CSE stimulation. Histone deacetylase-2 （HDAC2） short interference RNA （HDAC2-siRNA） was transfected into the cells using LipofectamineTM 2000. Interleukin-8 （IL-8） level in supernatants was measured by ELISA and HDAC2 expression was determined by real-time quantitative PCR （qRT-PCR） and Western blotting. Results The inhibition ratio of IL-8 in the EM group was significantly higher than that in the CSE group, but lower than that in the control group （ P 〈 0.05）. The half-maximal inhibitory concentration of dexamethasone （ICs0-Dex） in the EM group was lower than that in the CSE group, but higher than that in the control group （ P〈O. 05）. The expression of HDAC2 protein in the EM group was higher than that in the CSE group, but lower than that in the control group （ P 〈 0.05 ）. Besides, HDAC2 mRNA and HDAC2 protein expressions were lower in the HDAC2-siRNA group than in the scrambled oligonucleotide （SC） group. EM could reverse HDAC2 mRNA and HDAC2 protein reduction induced by HDAC2-siRNA （ P 〈 0.05）. Conclusion Corticosteroid sensitivity of THP-1 cells could be reduced by CSE. EM could reverse the corticosteroid insensitivity by up-regulating the expression of HDAC2 protein.