β-谷甾醇对脂多糖诱导的乳腺上皮细胞氧化应激及乳脂合成的影响

Effects ofβ-Sitosterol on Oxidative Stress and Milk Fat Synthesis of Mammary Epithelial Cells Induced by Lipopolysaccharide

本试验旨在探讨β-谷甾醇(BSS)对脂多糖(LPS)诱导的小鼠乳腺上皮细胞(HC11细胞)氧化应激及乳脂合成的影响。试验以体外培养的HC11细胞为模型,分为5组:对照组(CON组,生长培养基处理)、乳腺炎症模型组(LPS组,1μg/mL LPS处理)以及炎症模型药物组(BSS+LPS组,分别用5、10、20μmol/L的BSS预处理1 h后加入1μg/mL LPS),每组设置5个重复。检测HC11细胞氧化应激指标及抗氧化相关基因、乳脂合成相关基因mRNA和蛋白表达的变化。结果表明:1)与CON组相比,LPS组的HC11细胞丙二醛(MDA)、一氧化氮(NO)含量及诱导型一氧化氮合酶(iNOS)活性显著升高(P<0.05),超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GPx)、过氧化氢酶(CAT)活性和总抗氧化能力(T-AOC)显著降低(P<0.05)。与LPS组相比,5、10、20μmol/L BSS+LPS组的HC11细胞MDA、NO含量及iNOS活性显著降低(P<0.05),SOD、GPx、CAT活性和T-AOC显著升高(P<0.05)。2)与CON组相比,LPS组的HC11细胞核因子E2相关因子2(Nrf2)、血红素氧合酶-1(HO-1)和NAD(P)H醌氧化还原酶1(NQO1)mRNA和蛋白相对表达量显著降低(P<0.05)。与LPS组相比,5、10、20μmol/L BSS+LPS组的HC11细胞Nrf2、HO-1和NQO1 mRNA和蛋白相对表达量显著升高(P<0.05)。3)与CON组相比,LPS组的HC11细胞过氧化物酶体增殖物激活受体γ(PPARγ)和固醇调节元件结合蛋白1(SREBP1)mRNA和蛋白相对表达量显著降低(P<0.05),乙酰辅酶A羧化酶(ACC)、脂肪酸合成酶(FAS)和硬脂酰辅酶A去饱和酶(SCD)mRNA相对表达量显著降低(P<0.05)。与LPS组相比,5、10、20μmol/L BSS+LPS组的HC11细胞PPARγ和SREBP1 mRNA和蛋白相对表达量显著升高(P<0.05),ACC、FAS和SCD mRNA相对表达量显著升高(P<0.05)。由此可见,BSS能够抑制LPS诱导的HC11细胞氧化损伤,并能促进氧化应激状态下HC11细胞乳脂的合成。

The purpose of this experiment was to explore the effects ofβ-sitosterol(BSS)on oxidative stress and milk fat synthesis of mammary epithelial cells(HC11 cells)induced by lipopolysaccharide(LPS).The HC11 cells cultured in vitro were used as a model,and divided into 5 groups:control group(CON group,growth medium treated),mammary inflammatory model group(LPS group,1μg/mL LPS treated)and inflammatory model medicine groups(BSS+LPS groups,after 5,10 and 20μmol/L BSS treated 1 h,respectively,added 1μg/mL LPS),and set 5 replicates per group.The oxidative stress indexes and the mRNA expression and protein expression of antioxidant related genes and milk fat synthesis related genes in HC11 cells were detected.The results showed as follows:1)compared with the CON group,the malondialdehyde(MDA),nitric oxide(NO)contents and inducible nitric oxide synthase(iNOS)activity in HC11 cells of LPS group were significantly increased(P<0.05),and the superoxide dismutase(SOD),glutathione peroxidase(GPx),catalase(CAT)activities and total antioxidant capacity(T-AOC)were significantly decreased(P<0.05).Compared with the LPS group,the MDA,NO content and iNOS activity in HC11 cells of 5,10 and 20μmol/L BSS+LPS groups were significantly decreased(P<0.05),and the SOD,GPx,CAT activities and T-AOC were significantly increased(P<0.05).2)Compared with the CON group,the mRNA and protein relative expression levels of nuclear factor erythroid 2-related factor 2(Nrf2),heme oxygenase-1(HO-1)and NAD(P)H:quinone oxidoreductase 1(NQO1)in HC11 cells of LPS group were significantly decreased(P<0.05).Compared with the LPS group,the mRNA and protein relative expression levels of Nrf2,HO-1 and NQO1 in HC11 cells of 5,10 and 20μmol/L BSS+LPS groups were significantly increased(P<0.05).3)Compared with the CON group,the mRNA and protein relative expression levels of peroxisome proliferator activated receptorγ(PPARγ)and sterol regulatory element binding protein 1(SREBP1)in HC11 cells of LPS group were significantly decreased(P<0.05),and the mRNA relative expression levels of acetyl-CoA carboxylase(ACC),fatty acid synthase(FAS)and stearyl coenzyme A desaturated enzyme(SCD)were significantly decreased(P<0.05).Compared with the LPS group,the mRNA and protein relative expression levels of PPARγand SREBP1 in HC11 cells of 5,10 and 20μmol/L BSS+LPS groups were significantly increased(P<0.05),and the mRNA relative expression levels of ACC,FAS and SCD were significantly increased(P<0.05).In conclusion,BSS can inhibit LPS-induced HC11 cells oxidative damage,and promote milk fat synthesis in HC11 cells under oxidative stress.