Koren R, Hadari-Naor I, Zuck E, Rotem C, Liberman U A, Ravid A
Basil and Gerald Felsenstein Medical Research Center, Rabin Medical Center, Petah Tikva, Israel.
Cancer Res. 2001 Feb 15;61(4):1439-44.
The anticancer activity of the hormonal form of vitamin D, 1,25-dihydroxyvitamin D [1,25(OH)2D], is associated with inhibition of cell cycle progression, induction of differentiation, and apoptosis. In addition, 1,25(OH)2D3 augments the activity of anticancer agents that induce excessive reactive oxygen species generation in their target cells. This study aimed to find out whether 1,25(OH)2D3, acting as a single agent, is a prooxidant in cancer cells. The ratio between oxidized and reduced glulathione and the oxidation-dependent inactivation of glyceraldehyde-3phosphate dehydrogenase (GAPDH) are considered independent markers of cellular reactive oxygen species homeostasis and redox state. Treatment of MCF-7 breast cancer cells with 1,25(OH)2D3 (10-100 nM for 24-48 h) brought about a maximal increase of 41+/-13% (mean +/- SE) in the oxidized/reduced glutathione ratio without affecting total glutathione levels. The in situ activity of glutathione peroxidase and catalase were not affected by 1,25(OH)2D3, as assessed by the rate of H2O2 degradation by MCF-7 cell cultures. Neither did treatment with 1,25(OH)2D3 affect the levels of glutathione reductase or glutathione S-transferase as assayed in cell extracts. The hormone did not affect overall glutathione consumption and efflux as reflected in the rate of decline of total cellular glutathione after inhibition of its synthesis by buthionine sulfoximine. The extent of reversible oxidation-dependent inactivation of GAPDH in situ was determined by comparing the enzyme activity before and after reduction of cell extracts with DTT. The oxidized fraction was 0.13+/-0.02 of total GAPDH in control cultures and increased by 56+/-5.3% after treatment with 1,25(OH)2D3, which did not affect the total reduced enzyme activity. Treatment with 1,25(OH)2D3 resulted in a approximately 40% increase in glucose-6-phosphate dehydrogenase, the rate-limiting enzyme in the generation of NADPH. This enzyme is induced in response to various modes of oxidative challenge in mammalian cells. Taken together, these findings indicate that 1,25(OH)2D3 causes an increase in the overall cellular redox potential that could translate into modulation of redox-sensitive enzymes and transcription factors that regulate cell cycle progression, differentiation, and apoptosis.
维生素D的激素形式1,25 - 二羟基维生素D [1,25(OH)₂D]的抗癌活性与抑制细胞周期进程、诱导分化和凋亡有关。此外，1,25(OH)₂D₃可增强在其靶细胞中诱导过量活性氧生成的抗癌药物的活性。本研究旨在探究作为单一药物的1,25(OH)₂D₃在癌细胞中是否为促氧化剂。氧化型与还原型谷胱甘肽的比例以及甘油醛 - 3 - 磷酸脱氢酶（GAPDH）的氧化依赖性失活被视为细胞活性氧稳态和氧化还原状态的独立标志物。用1,25(OH)₂D₃（10 - 100 nM处理24 - 48小时）处理MCF - 7乳腺癌细胞，使氧化型/还原型谷胱甘肽比例最大增加41±13%（平均值±标准误），且不影响总谷胱甘肽水平。通过MCF - 7细胞培养物中H₂O₂降解速率评估，谷胱甘肽过氧化物酶和过氧化氢酶的原位活性不受1,25(OH)₂D₃影响。用1,25(OH)₂D₃处理也不影响在细胞提取物中检测的谷胱甘肽还原酶或谷胱甘肽S - 转移酶的水平。该激素不影响总体谷胱甘肽消耗和流出，这在丁硫氨酸亚砜胺抑制其合成后总细胞谷胱甘肽下降速率中得到体现。通过比较用二硫苏糖醇还原细胞提取物前后的酶活性，确定原位GAPDH可逆氧化依赖性失活的程度。在对照培养物中，氧化部分占总GAPDH的0.13±0.02，用1,25(OH)₂D₃处理后增加了56±5.3%，但不影响总还原酶活性。用1,25(OH)₂D₃处理导致葡萄糖 - 6 - 磷酸脱氢酶增加约40%，该酶是NADPH生成中的限速酶。该酶在哺乳动物细胞中响应各种氧化应激模式而被诱导。综上所述，这些发现表明1,25(OH)₂D₃导致细胞总体氧化还原电位升高，这可能转化为对调节细胞周期进程、分化和凋亡的氧化还原敏感酶和转录因子的调节。
