Yamaguchi M, Sugimoto E
Laboratory of Endocrinology and Molecular Metabolism, Graduate School of Nutritional Sciences, University of Shizuoka, Japan.
Mol Cell Biochem. 2000 Nov;214(1-2):97-102. doi: 10.1023/a:1007199120295.
The effect of genistein and daidzein on protein synthesis in osteoblastic MC3T3-E1 cells in vitro was investigated to determine a cellular mechanism by which the isoflavones stimulate bone formation. Cells were cultured for 48 h in alpha-minimal essential medium containing either vehicle, genistein (l0(-7) - 10(-5) M) or daidzein (10(-7) - 10(-5) M). The 5,500 g supernatant of cell homogenate was used for assay of protein synthesis with [3H]leucine incorporation in vitro. The culture with genistein or daidzein caused a significant elevation of protein synthesis in the cell homogenate. The effect of genistein ( 10(-5) M) or daidzein ( 10(-5) M) in elevating protein synthesis was significantly prevented, when cells were cultured for 48 h in a medium containing either actinomycin D (10(-7) M) or cycloheximide (10(-6) M) in the absence or presence of isoflavones. Moreover, when genistein (10(-7) 10(-5) M) or daidzein (10(-6) and 10(-5) M) was added to the reaction mixture containing the cell homogenate obtained from osteoblastic cells cultured without isoflavone, protein synthesis was significantly raised. This increase was markedly blocked by the addition of cycloheximide (10(-7) M). In addition, [3H]leucyl-tRNA synthetase activity in the cytosol of osteoblastic cells was significantly increased by the addition of genistein (10(-6) and 10(-5) M) or daidzein (10(-5) M) into the enzyme reaction mixture. The present study demonstrates that genistein or daidzein can stimulate protein synthesis in osteoblastic MC3T3-E1 cells. The isoflavones may have a stimulatory effect on osteoblastic bone formation due to increasing protein synthesis.
研究了染料木黄酮和大豆苷元对体外培养的成骨细胞MC3T3-E1蛋白质合成的影响，以确定异黄酮刺激骨形成的细胞机制。细胞在含有载体、染料木黄酮（10⁻⁷ - 10⁻⁵ M）或大豆苷元（10⁻⁷ - 10⁻⁵ M）的α-最低必需培养基中培养48小时。细胞匀浆的5500g上清液用于通过体外掺入[³H]亮氨酸来测定蛋白质合成。用染料木黄酮或大豆苷元培养可使细胞匀浆中的蛋白质合成显著升高。当细胞在含有放线菌素D（10⁻⁷ M）或环己酰亚胺（10⁻⁶ M）的培养基中培养48小时时，无论是否存在异黄酮，染料木黄酮（10⁻⁵ M）或大豆苷元（10⁻⁵ M）提高蛋白质合成的作用均被显著抑制。此外，当将染料木黄酮（10⁻⁷ - 10⁻⁵ M）或大豆苷元（10⁻⁶和10⁻⁵ M）添加到含有从未用异黄酮培养的成骨细胞获得的细胞匀浆的反应混合物中时，蛋白质合成显著提高。加入环己酰亚胺（10⁻⁷ M）可显著阻断这种增加。此外，将染料木黄酮（10⁻⁶和10⁻⁵ M）或大豆苷元（10⁻⁵ M）添加到酶反应混合物中可使成骨细胞胞质溶胶中的[³H]亮氨酰-tRNA合成酶活性显著增加。本研究表明，染料木黄酮或大豆苷元可刺激成骨细胞MC3T3-E1中的蛋白质合成。异黄酮可能由于增加蛋白质合成而对成骨细胞骨形成具有刺激作用。
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