ABCA1对胎盘脂质代谢的影响

目的:探讨ATP结合盒转运蛋白A1(ABCA1)对胎盘脂质代谢的影响。方法:应用酶消化法分离胎盘原代滋养细胞,流式细胞术进行细胞纯度鉴定,免疫荧光检测细胞中ABCA1的表达情况。实验组应用肝X受体激动剂(T0901317)上调(以无菌磷酸缓冲盐溶液代替T0901317作为对照组)或siRNA下调(以mock-siRNA代替siRNA作为模拟转染组)滋养细胞中ABCA1的表达水平并应用RT-PCR及Western-blot进行验证,采用Amplex胆固醇检测试剂盒检测ABCA1表达改变后滋养细胞在实验开始后的2小时、4小时、6小时及8小时胆固醇流出率。结果:从足月胎盘中分离出原代滋养细胞,其纯度在90%以上,ABCA1在滋养细胞中存在表达。加入T0901317后,滋养细胞中ABCA1在蛋白水平的表达量(0.37±0.04)明显高于对照组(0.08±0.02),差异有统计学意义(P<0.05);而在转染siRNA后,滋养细胞中ABCA1在蛋白水平的表达量(0.10±0.01)低于对照组(0.16±0.02),差异有统计学意义(P<0.05)。在ABCA1表达上升后2小时、4小时、6小时、8小时,滋养细胞胆固醇流出率与对照组相比明显升高(4.70±0.03vs3.39±0.00、7.66±0.05vs5.28±0.02、14.73±0.03vs8.66±0.02、20.94±0.07vs13.47±0.03),差异均有统计学意义(P<0.05);而当ABCA1表达量下降后2小时、4小时、6小时、8小时,滋养细胞胆固醇流出率与对照组相比明显降低(2.30±0.03vs3.53±0.02、3.35±0.03vs5.17±0.01、5.37±0.05vs7.01±0.02、7.56±0.06vs11.49±0.03),差异均有统计学意义(P<0.05)。结论:当胎盘中ABCA1表达出现异常时,可通过影响滋养细胞胆固醇的转运而导致母胎界面脂质代谢出现紊乱。

The Role of ABCA1 on Placental Lipid Metabolism

Objective:To detective the potential role of ABCA1 on the process of placental lipid metabolism.Methods:The primary trophoblasts of placenta were isolated by enzyme digestion,while the cell purity was identified by flow cytometry,and the expression of ABCA1 was detected by immunofluorescence.Up-regulation of ABCA1 expression in trophoblasts was performed usingliver X-receptor agonist(sterile phosphate buffered salineas controlinstead of T0901317) or down-regulation of ABCA1 expression in trophoblast cells using siRNA(mock-siRNA as control instead of siRNA).Those regulations were verifiedby RT-PCR and Western-blog.The Amplex Cholesterol Detection Kit was used to detect the cholesterol efflux rate of trophoblast cells at 2 hours,4 hours,6 hours,and 8 hours after the start of the experiment.Results:We successfully isolated primary trophoblast cells from term placenta with a purity of more than 90%.The expression of ABCA1 in trophoblast cells was confirmed. After adding T0901317,the expression level of ABCA1 in the trophoblast cells(0.37±0.04) was significantly higher than that in the control group(0.08± 0.02)(P< 0.05).After transfection of siRNA,the expression level of ABCA1 in the trophoblast cells(0.10±0.01) was lower than that of the control group(0.16±0.02),and the difference was statistically significant( P<0.05).At 2 hours,4 hours,6 hours,and 8 hours after the increase of ABCA1 expression level,the function of trophoblastic transport of cholesterol was significantly higher than that of the control group(4.70±0.03 vs 3.39±0.00,7.66± 0.05 vs 5.28±0.02,14.73±0.03 vs 8.66±0.02,20.94±0.07 vs 13.47±0.03,respectively).The differences were statistically significant(P<0.05).When the expression of ABCA1 decreased after 2 hours,4 hours,6 hours and 8 hours,the function of cholesterol transport of was significantly lower than that of the control group(2.30±0.03 vs 3.53±0.02,3.35±0.03 vs 5.17±0.01,5.37±0.05 vs 7.01±0.02,7.56±0.06 vs 11.49±0.03,respectively).The differences were statistically significant( P<0.05).Conclusions:When the expression of ABCA1 in placenta was abnormal,it could affect the lipid metabolism of placenta by affecting the cholesterol transport function of trophoblast cells.