摘要
目的:探讨载脂蛋白嵌合模拟肽Ac-hE-18A-NH2对氧化型低密度脂蛋白(oxidized LDL,ox-LDL)刺激下巨噬细胞肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)表达的影响及其作用机制.方法:Ox-LDL刺激RAW264.7巨噬细胞,给予不同浓度的模拟肽Ac-hE-18A-NH2(1～100μg/mL)干预,收集细胞,测定巨噬细胞TNF-α的分泌和mRN表达水平.Western印迹检测ATP结合盒转运蛋白A1 (ATP-binding cassette transporter A1,ABCA1)及P-IκB蛋白浓度.EMSA检测核因子-κB(nuclear factor-κB,NF-κB)活性.结果:Ox-LDL刺激使RAW264.7巨噬细胞TNF-α分泌和mRNA表达明显增强,细胞内胆固醇蓄积,促进IκB磷酸化,并激活NF-κB.Ac-hE-18A-NH2浓度依赖性地降低TNF-α分泌及mRNA表达,上调ABCA1 mRNA和蛋白的表达,减少细胞内胆固醇含量,抑制NF-κB活化,并抑制IκB磷酸化.相同的实验条件下及作用浓度,D-4F对于TNF-α分泌的抑制作用不如Ac-hE-18A-NH2.结论:Ac-hE-18A-NH2能抑制ox-LDL诱导的RAW264.7巨噬细胞TNF-α分泌和mRNA表达,IκB/NF-κB-TNF-α信号通路是其中作用途径之一.Ac-hE-18A-NH2的抗炎作用优于apoA-I模拟肽D-4F.
Objective: To evaluate the ef ect of Ac-hE-18A-NH2 on TNF-α secretion and mRNA expression in ox-LDL-stimulated A W264.7 macrophages and to elucidate the possible mechanisms. Methods: Macrophages were incubated in the medium containing various concentrationsof Ac-hE18A-NH2(1-50 μg/mL) with ox-LDL(50 μg/mL) stimulated. The TNF-α level and intracellular cholesterol content were measured by commercially available quantitation kits following the manufacturer's instructions. TNF-α and ATP-binding cassette transporter A1(ABCA1) mRNA expression were detected by real-time PCR. ABCA1 and IκB protein-expression in the macrophages were determined by Western blot. NF-κB activity was evaluated by electrophoretic mobility shit assay(EMSA). Results: Ox-LDL stimulation induced a signii cant increase in TNF-α secretion, mRNA expression, cholesterol accumulation and nuclear factor-κB(NF-κB) activity in RAW264.7 macrophages. Ac-hE-18A-NH2 reduced TNF-α secretion and mRNA expression, up-regulated the ABCA1 mRNA and protein expression, reduced the intracellular cholesterol content, and inhibited NF-κB activation in a dose-dependent manner. Under the same condition and the same concentration, Ac-hE-18A-NH2 was more efficient than D-4F(apoA-I mimetic peptide) in inhibiting the inl ammatory response induced by ox-LDL in the macrophages. Conclusion: Ac-hE-18A-NH2 may suppress TNF-α secretion and mRNA expression in ox-LDLstimulated A W264.7 macrophages via IκB-NF-κB signaling pathway. h e anti-inl ammatory ef ect of Ac-hE-18A-NH2 is bet er than that of apoA-I mimic peptide D-4F.
出处
《中南大学学报（医学版）》
CAS
CSCD
北大核心
2014年第3期232-238,共7页
Journal of Central South University ：Medical Science
基金
国家自然科学基金(30770857)~~
