苜蓿皂苷对胆固醇逆向转运基因三磷酸腺苷结合盒转运体A1、清道夫受体BⅠmRNA表达的影响

Effects of Alfalfa Saponins m RNA Expressions of Reverse Cholesterol Transport Genes:ATP-Binding Cassette Transporter A1 and Scavenger Receptor Class B Type Ⅰ

本试验以大鼠肝脏和肝脏细胞(BRL细胞)及小鼠巨噬细胞(ANA-1细胞)为研究对象,从动物和细胞水平上研究苜蓿皂苷对胆固醇逆向转运基因三磷酸腺苷结合盒转运体A1(ABCA1)、清道夫受体BⅠ(SR-BⅠ)mRNA表达的影响。取雄性健康SD大鼠32只,随机分成4组,分别为正常对照组、苜蓿皂苷组、高脂模型组和高脂苜蓿皂苷组,每组8只。正常对照组和苜蓿皂苷组饲喂基础饲粮,其余2组均饲喂高脂饲粮,饲喂4周后,苜蓿皂苷组和高脂皂苷组从第5周开始每天灌胃240 mg/kg苜蓿皂苷,灌胃4周。采用胎牛血清作用于BRL细胞48 h,构建脂变模型,将BRL细胞分为正常对照组(正常细胞)、苜蓿皂苷组(正常细胞)、脂变模型组(脂变细胞)和脂变皂苷组(脂变细胞),苜蓿皂苷组、脂变皂苷组培养液中添加苜蓿皂苷(终浓度300μg/m L),培养24 h。采用氧化低密度脂蛋白(ox-LDL)作用于ANA-1细胞48 h,构建荷脂模型,将ANA-1细胞分为正常对照组(正常细胞)、苜蓿皂苷组(正常细胞)、荷脂模型组(荷脂细胞)和荷脂皂苷组(荷脂细胞),苜蓿皂苷组、荷脂皂苷组培养液中添加苜蓿皂苷(终浓度300μg/m L),培养24 h。采用荧光定量PCR法测定大鼠肝脏、BRL细胞和ANA-1细胞中ABCA1、SR-BⅠmRNA表达量。结果表明:1)苜蓿皂苷显著提高了正常大鼠肝脏ABCA1和SR-BⅠmRNA表达量(P<0.05),显著提高了高脂大鼠肝脏ABCA1 mRNA的表达量(P<0.05),对高脂大鼠肝脏SR-BⅠmRNA表达量影响不显著(P>0.05);2)苜蓿皂苷显著提高了正常BRL细胞ABCA1和SR-BⅠmRNA的表达量(P<0.05),而对脂变BRL细胞ABCA1和SR-BⅠmRNA表达量影响不大(P>0.05);3)苜蓿皂苷显著降低正常ANA-1细胞ABCA1 mRNA表达量(P<0.05)。由此可见,苜蓿皂苷可通过上调大鼠肝脏和正常肝脏细胞ABCA1和SR-BⅠmRNA的表达促进胆固醇的逆向转运,增强肝脏胆固醇排泄,从而发挥其对高脂血症的预防和治疗作用。

Rat liver,rat liver cells（BRL cells） and mouse macrophages（ANA-1 cells） were used to investigate the effects of alfalfa saponins（AS） on mRNA expressions of reverse cholesterol transport genes,which were ATP-binding cassette transporter A1（ABCA1） and scavenger receptor class B type Ⅰ（SR-B Ⅰ）,and discuss the effects of AS from animal and cellular levels.Thirty two healthy male SD rats were randomly divided into four groups： normal control group,AS group,hyperlipidemic model group and hyperlipidemic saponins group,and each group had 8 rats.Rats in normal control group and AS group were fed a basal diet,and those in the other two groups were fed a high-fat diet.After 4 weeks feeding,AS [240 mg/（kg ·d）]was intragastric administered to rats in AS group and hyperlipidemic saponins group from weeks 5 to 8.BRL cells were incubated with fetal bovine serum for 48 h to constitute hyperlipidemic model.BRL cells were divided into four groups,which were normal control group（normal cells）,AS group（normal cells）,hyperlipidemic model group（hyperlipidemic cells） and hyperlipidemic saponins group（hyperlipidemic cells）,and AS（final concentration 300 μg/m L） was added to culture medium in AS group and hyperlipidemic saponins group to culture for 24 h.ANA-1 cells were incubated with oxidized lowdensity lipoprotein for 48 h to constitute lipidloaded model.ANA-1 cells were divided into four groups,which were normal control group（normal cells）,AS group（normal cells）,lipid-loaded model group（lipid-loaded cells） and lipid-loaded saponins group（lipid-loaded cells）,and AS（final concentration 300 μg/m L） was added to culture medium in AS group and lipid-loaded saponins group to culture for 24 h.The mRNA expressions of ABCA1 and SR-BⅠ in rat liver,BRL cells and ANA-1 cells were determined by fluorescent quantitative PCR.The results showed as follows： 1） AS significantly increased the mRNA expressions of ABCA1 and SR-BⅠ in liver of normal rats and of ABCA1 in liver of hyperlipidemic rats（P〈0.05）,but had no significant effects on that of SR-BⅠ in liver of hyperlipidemic rats（P〉0.05）; 2） AS significantly increased the mRNA expressions of ABCA1 and SR-BⅠ in normal BRL cells（P〈0.05）,but had no significant effects on those in hyperlipidemic BRL cells（P〉0.05）; 3） AS significantly reduced the mRNA expression of ABCA1 in normal ANA-1 cells（P〈0.05）.In conclusion,AS might promote reverse cholesterol transport by up-regulating the mRNA expressions of ABCA1 and SR-B Ⅰ in rat liver and normal BRL cells,promote the excretion of liver cholesterol,and play prevention and curing effects on hyperlipidemia.