Department of Veterinary Medicine, Faculty of Agriculture, Kagoshima University, Kagoshima, Japan.
Nutrition. 2010 Jun;26(6):648-52. doi: 10.1016/j.nut.2009.08.005. Epub 2009 Nov 20.
Vitamin B6 (B6) suppresses the expression of cyclooxygenase-2 stimulated by lipopolysaccharide in mouse macrophage RAW264.7 cells. The greatest effect is recognized for pyridoxal (PL) compared with pyridoxamine (PM), pyridoxine (PN), and pyridoxal 5'-phosphate (PLP). However, it has not been elucidated why PL has the strongest effect. We compared the uptakes and cell surface interactions among PL, PM, PN, and PLP in RAW264.7 cells.
Cyclo-oxygenase-2 mRNA expression was evaluated by real-time polymerase chain reaction. Intracellular B6 concentrations were measured by high-performance liquid chromatography. Interactions of B6s with the cell surface were analyzed using a surface plasmon resonance biosensor. B6 uptake speeds were measured using [(3)H]-PN.
The intracellular PLP levels did not change significantly when cells were cultured in medium containing PL, PM, PN, or PLP. Only PL interacted with the cell surface. Although PM and PN were associated with the cell surface, their binding was only recognized during sample loading. After the change to phosphate buffered saline after sample loading, the binding resonances of PM and PN returned to baseline, whereas that of PL did not. Uptake of [(3)H]-PN was inhibited by non-labeled PN, PL, or PLP, but not PM, at 1 microM. The inhibition rate of PL was higher than those of PN and PLP.
The inhibition of cyclo-oxygenase-2 mRNA expression by PL may be related to the cell surface interaction of PL, rather than the intracellular PLP level. The uptake mechanism for PN and PL may differ from that for PM.
维生素 B6（B6）可抑制脂多糖刺激的小鼠巨噬细胞 RAW264.7 中环氧化酶-2 的表达。与吡哆胺（PM）、吡哆醇（PN）和吡哆醛 5′-磷酸（PLP）相比，吡哆醛（PL）的作用最大。然而，为什么 PL 具有最强的作用还没有阐明。我们比较了 PL、PM、PN 和 PLP 在 RAW264.7 细胞中的摄取和细胞表面相互作用。
通过实时聚合酶链反应评估环氧化酶-2 mRNA 的表达。通过高效液相色谱法测量细胞内 B6 浓度。使用表面等离子体共振生物传感器分析 B6 与细胞表面的相互作用。使用[（3）H]-PN 测量 B6 的摄取速度。
当细胞在含有 PL、PM、PN 或 PLP 的培养基中培养时，细胞内 PLP 水平没有明显变化。只有 PL 与细胞表面相互作用。虽然 PM 和 PN 与细胞表面结合，但仅在样品加载时才被识别。在样品加载后更换磷酸盐缓冲盐水时，PM 和 PN 的结合共振恢复到基线，而 PL 的结合共振没有恢复。在 1μM 时，非标记的 PN、PL 或 PLP 抑制[（3）H]-PN 的摄取，但 PM 不抑制。PL 的抑制率高于 PN 和 PLP。
PL 抑制环氧化酶-2 mRNA 的表达可能与 PL 的细胞表面相互作用有关，而与细胞内 PLP 水平无关。PN 和 PL 的摄取机制可能与 PM 不同。
