CAJ | 학술논문

目的观察可溶性环氧化物水解酶抑制剂t-AUCB对小鼠巨噬细胞泡沫化及胆固醇流出率的影响,明确t-AUCB是否可抑制泡沫细胞的形成.方法培养小鼠巨噬细胞系RAW264.7,分组为空白对照组、不同浓度t-AUCB干预组、100 μmol/L t-AUCB+ GW9662组.t-AUCB干预组再分为4个亚组,分别用不同浓度t-AUCB(1,10,50,100 μmol/L)干预24h.100 μnol/L t-AUCB+ GW9662组为在t-AUCB前1h加入过氧化物酶增殖体激活型受体γ(PPARγ)拮抗剂GW9662 5μmol/L.采用油红O染色鉴定泡沫细胞的形成,液体闪烁计数器检测各组细胞3氚标记胆固醇(3 H-Cholesterol)的流出率.实时PCR和免疫印迹法分别测定巨噬细胞三磷酸腺苷结合盒转运子A1( ABCAl) mRNA和蛋白的表达.结果油红O染色24h后,空白对照组细胞大量红染,t-AUCB组细胞红染明显少于空白对照组(P＜0.05).100 μmol/L t-AUCB+GW9662组细胞大量红染,显著多于100 μmol/Lt-AUCB组(P＜0.05).空白对照组胆固醇流出率为(5.91±0.18)％.1、10、50和100 μmol/Lt-AUCB干预组胆固醇流出率分别为(7.03±0.33)％、(8.05±0.32)％、(9.04 ±0.14)％和(10.06±0.85)％,均明显高于空白对照组(P均＜0.05).100 μmol/L t-AUCB+ GW9662组胆固醇流出率为(6.33±0.15)％,明显低于100 μmol/L t-AUCB组,P＜0.05.各浓度t-AUCB干预组小鼠巨噬细胞ABCA1 mRNA和蛋白表达均明显高于空白对照组,P均＜0.05,100 μmol/L t-AUCB+ GW9662组ABCA1 mRNA和蛋白表达显著低于100 μmol/L t-AUCB组,P＜0.05.结论 t-AUCB可通过PPARγ-ABCA1途径改善巨噬细胞胆固醇流出,抑制巨噬细胞内胆固醇过度蓄积,从而抑制泡沫细胞的形成.
목적 관찰가용성배양화물수해매억제제t-AUCB대소서거서세포포말화급담고순류출솔적영향,명학t-AUCB시부가억제포말세포적형성.방법 배양소서거서세포계RAW264.7,분조위공백대조조、불동농도t-AUCB간예조、100 μmol/L t-AUCB+ GW9662조.t-AUCB간예조재분위4개아조,분별용불동농도t-AUCB(1,10,50,100 μmol/L)간예24h.100 μnol/L t-AUCB+ GW9662조위재t-AUCB전1h가입과양화물매증식체격활형수체γ(PPARγ)길항제GW9662 5μmol/L.채용유홍O염색감정포말세포적형성,액체섬삭계수기검측각조세포3천표기담고순(3 H-Cholesterol)적류출솔.실시PCR화면역인적법분별측정거서세포삼린산선감결합합전운자A1( ABCAl) mRNA화단백적표체.결과 유홍O염색24h후,공백대조조세포대량홍염,t-AUCB조세포홍염명현소우공백대조조(P＜0.05).100 μmol/L t-AUCB+GW9662조세포대량홍염,현저다우100 μmol/Lt-AUCB조(P＜0.05).공백대조조담고순류출솔위(5.91±0.18)％.1、10、50화100 μmol/Lt-AUCB간예조담고순류출솔분별위(7.03±0.33)％、(8.05±0.32)％、(9.04 ±0.14)％화(10.06±0.85)％,균명현고우공백대조조(P균＜0.05).100 μmol/L t-AUCB+ GW9662조담고순류출솔위(6.33±0.15)％,명현저우100 μmol/L t-AUCB조,P＜0.05.각농도t-AUCB간예조소서거서세포ABCA1 mRNA화단백표체균명현고우공백대조조,P균＜0.05,100 μmol/L t-AUCB+ GW9662조ABCA1 mRNA화단백표체현저저우100 μmol/L t-AUCB조,P＜0.05.결론 t-AUCB가통과PPARγ-ABCA1도경개선거서세포담고순류출,억제거서세포내담고순과도축적,종이억제포말세포적형성.
Objective To observe the effects of soluble epoxide hydrolase inhibitor t-AUCB on foam cell formation and cholesterol efflux in macrophage.Methods Mouse macrophages RAW264.7 were cultured and stimulated with ox-LDL (80 μmol/L) in the absence (group A) or presence of t-AUCB (1,10,50,100 μmol/L,group B) or t-AUCB (100 μmol/L) pretreated with PPARγ antagonist GW9662 (5 μmol/L,group C).The foam cell was identified by oil red O staining.The cholesterol efflux rates of 3 Hcholesterol in cells were measured by liquid scintillation counter.mRNA and protein expressions of ABCA1 were detected by real-time PCR or Western blot,respectively.Results Oil red O staining showed that t-AUCB (100 μmol/L) significantly inhibited foam cell formation which could be significantly reversed by GW9662 (all P ＜ 0.05).t-AUCB dese-dependently increased cholesterol efflux rates in mouse macrophage [(5.91+0.18)％ in group A,(7.03 ±0.33)％,(8.05 ±0.32)％,( 9.04 ±0.14)％,(10.06±0.85)％ in 1,10,50,100 μmol/L t-AUCB groups,all P＜0.05 vs.group A],which could be reversed by pretreatment with GW9662 [ (6.33 ±0.15)％ in 100 μmol/L t-AUCB + GW9662 group].t-AUCB also upregulated ABCA1 mRNA and protein expressions in a dose-dependent manner which could be significantly attenuated by pretreatment with GW9662.Condusion t-AUCB could inhibit foam cell formation by improving cholesterol efflux through activating PPARγ-ABCA1 pathway in macrophage.