体外实验中，17β-雌二醇、染料木黄酮或硫酸锌可增强β-隐黄质对成骨细胞MC3T3-E1细胞的合成代谢作用：锌对Runx2和α1(I)胶原mRNA表达具有独特影响。
Anabolic effect of beta-cryptoxanthin in osteoblastic MC3T3-E1 cells is enhanced with 17beta-estradiol, genistein, or zinc sulfate in vitro: the unique effect with zinc on Runx2 and alpha1(I) collagen mRNA expressions.
作者信息
Uchiyama Satoshi, Yamaguchi Masayoshi
机构信息
Laboratory of Endocrinology and Molecular Metabolism, Graduate School of Nutritional Sciences, University of Shizuoka, Shizuoka, Japan.
出版信息
Mol Cell Biochem. 2008 Jan;307(1-2):209-19. doi: 10.1007/s11010-007-9600-9. Epub 2007 Sep 25.
Whether the anabolic effect of beta-cryptoxanthin (CRP), a kind of carotenoid, on osteoblastic MC3T3-E1 cells are modulated in the presence of various hormones or nutrient factors were investigated. Cells were cultured for 72 h in a minimum essential medium containing 10% fetal bovine serum (FBS), and the cells with subconfluency were changed to a medium containing either vehicle or CRP (10(-8)-10(-6) M) in the presence or absence of various factors without FBS. Cells were cultured for 72 h. Protein content or alkaline phosphatase activity in osteoblastic cells were significantly increased after culture with CRP (10(-7) or 10(-6) M), 1,25-dihydroxyvitamin D(3) (VD(3); 10(-9) or 10(-8) M), 17beta-estradiol (E(2); 10(-9) M), genistein (10(-7) or 10(-6) M), or menaquinone-7 (MK-7; 10(-7) or 10(-6) M). The effect of CRP (10(-6) M) in increasing protein content in the cells was significantly enhanced in the presence of E(2) (10(-9) M) or genistein (10(-6) M). Gene expression in osteoblastic cells was determined using reverse transcription-polymerase chain reaction (RT-PCR). Culture with CRP (10(-7) or 10(-6) M) caused a significant increase in the expression of Runx2 and alkaline phosphatase mRNAs in the cells. Runx2 mRNA expression was significantly increased after culture with E(2) (10(-9) M) or MK-7 (10(-7) or 10(-6) M), but not VD(3) (10(-9) or 10(-8) M) or genistein (10(-7) or 10(-6) M). Alkaline phosphatase mRNA expression was significantly increased after culture with VD(3) (10(-9) or 10(-8) M), genistein (10(-7) or 10(-6) M), or MK-7 (10(-7) or 10(-6) M), but not E(2) (10(-10) or 10(-9) M). The effect of CRP (10(-7) or 10(-6) M) in increasing Runx2 or alkaline phosphatase mRNA expressions in the cells was not enhanced in the presence of VD(3), E(2), genistein, or MK-7. Culture with zinc sulfate (zinc; 10(-5) M) caused a significant increase in protein content or alkaline phosphatase activity in osteoblastic cells. The effect of CRP (10(-7) M) in increasing protein content or alkaline phosphatase activity in the cells was not significantly enhanced in the presence of zinc (10(-5) M). Culture with zinc (10(-5) M) caused a significant increase in alpha1(I) collagen mRNA expression, while it did not have a significant effect on Runx2 or osteocalcin mRNA expressions in the cells. The effect of CRP (10(-7) M) in increasing Runx2 or alpha1(I) collagen mRNA expressions was significantly enhanced in the presence of zinc (10(-6 )or 10(-5) M). Such an effect was not seen in the presence of cycloheximide (10(-7) M), an inhibitor of protein synthesis, or 5,6-dichloro-1-beta-D: -ribofuranosyl-benzimidazole (DRB; 10(-6) M), an inhibitor of transcriptional activity. This study demonstrates that the stimulatory effect of CRP on protein content in osteoblastic cells was additively enhanced with E(2) or genistein, and that the stimulatory effect of CRP on Runx2 or alpha1(I) collagen mRNA expressions was enhanced in the presence of zinc. Thus, the anabolic effect of CRP in osteoblastic MC3T3-E1 cells was modulated with a specific factor.
研究了在各种激素或营养因子存在的情况下，类胡萝卜素β-隐黄质（CRP）对成骨细胞MC3T3-E1细胞的合成代谢作用是否受到调节。细胞在含有10%胎牛血清（FBS）的最低必需培养基中培养72小时，将未达到汇合状态的细胞更换为不含FBS且含有载体或CRP（10⁻⁸ - 10⁻⁶ M）的培养基，并在有无各种因子的情况下培养72小时。用CRP（10⁻⁷或10⁻⁶ M）、1,25-二羟基维生素D₃（VD₃；10⁻⁹或10⁻⁸ M）、17β-雌二醇（E₂；10⁻⁹ M）、染料木黄酮（10⁻⁷或10⁻⁶ M）或甲基萘醌-7（MK-7；10⁻⁷或10⁻⁶ M）培养后，成骨细胞中的蛋白质含量或碱性磷酸酶活性显著增加。在E₂（10⁻⁹ M）或染料木黄酮（10⁻⁶ M）存在的情况下，CRP（10⁻⁶ M）增加细胞中蛋白质含量的作用显著增强。使用逆转录-聚合酶链反应（RT-PCR）测定成骨细胞中的基因表达。用CRP（10⁻⁷或10⁻⁶ M）培养导致细胞中Runx2和碱性磷酸酶mRNA的表达显著增加。用E₂（10⁻⁹ M）或MK-7（10⁻⁷或10⁻⁶ M）培养后，Runx2 mRNA表达显著增加，但用VD₃（10⁻⁹或10⁻⁸ M）或染料木黄酮（10⁻⁷或10⁻⁶ M）培养后未增加。用VD₃（10⁻⁹或10⁻⁸ M）、染料木黄酮（10⁻⁷或10⁻⁶ M）或MK-7（10⁻⁷或10⁻⁶ M）培养后，碱性磷酸酶mRNA表达显著增加，但用E₂（10⁻¹⁰或10⁻⁹ M）培养后未增加。在VD₃、E₂、染料木黄酮或MK-7存在的情况下，CRP（10⁻⁷或10⁻⁶ M）增加细胞中Runx2或碱性磷酸酶mRNA表达的作用未增强。用硫酸锌（锌；10⁻⁵ M）培养导致成骨细胞中的蛋白质含量或碱性磷酸酶活性显著增加。在锌（10⁻⁵ M）存在的情况下，CRP（10⁻⁷ M）增加细胞中蛋白质含量或碱性磷酸酶活性的作用未显著增强。用锌（10⁻⁵ M）培养导致α1（I）胶原mRNA表达显著增加，而对细胞中Runx2或骨钙素mRNA表达无显著影响。在锌（10⁻⁶或10⁻⁵ M）存在的情况下，CRP（10⁻⁷ M）增加Runx2或α1（I）胶原mRNA表达的作用显著增强。在蛋白质合成抑制剂环己酰亚胺（10⁻⁷ M）或转录活性抑制剂5,6-二氯-1-β-D-呋喃核糖基苯并咪唑（DRB；10⁻⁶ M）存在的情况下未观察到这种作用。本研究表明，CRP对成骨细胞中蛋白质含量的刺激作用与E₂或染料木黄酮呈累加增强，并且在锌存在的情况下，CRP对Runx2或α1（I）胶原mRNA表达的刺激作用增强。因此，CRP在成骨细胞MC3T3-E1细胞中的合成代谢作用受到特定因子的调节。
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