可溶性环氧化物水解酶抑制剂可抑制小鼠巨噬细胞泡沫化

Soluble epoxide hydrolase inhibitor t-AUCB ameliorates ox-LDL induced conversion of macrophages into foam cells through activating the PPARγ/-ABCA1 pathway

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、100μmoL／Lt-AUCB＋GW9662组。t-AUCB干预组再分为4个亚组，分别用不同浓度t—AUCB（1，10，50，100μmol／L）干预24h。100μmoL／Lt-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GW96625μmol／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’氚标记胆固醇（^3H—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Al（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）。100μmoL／Lt-AUCB＋GW9662组细胞大量红染，显著多于100μmoL／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100μmol／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）。100μmol／Lt-AUCB＋GW9662组胆固醇流出率为（6．33±0．15）％，明显低于100μmol／Lt-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〈0．05，100μmol／Lt-AUCB＋GW9662组ABCA1 mRNA和蛋白表达显著低于100μmol／Lt-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。

Objective To observe the effects of soluble epoxide hydrolase inhibitor t-AUCB on foam cell formation and cholesterol efflux in maerophage. 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 （100μmoL/L） pretreated with PPAR＇y antagonist GW9662 （5 μmol/L, group C）. The foam cell was identified by oil red O staining. The cholesterol efflux rates of ^3H- cholesterol 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 dose-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. Conclusion t-AUCB could inhibit foam cell formation by improving cholesterol efflux through activating PPARγ-ABCA1 pathway in macrophage.