Pyruvate improves recovery after PARP-1-associated energy failure induced by oxidative stress in neonatal rat cerebrocortical slices
Pyruvate improves recovery after PARP-1-associated energy failure induced by oxidative stress in neonatal rat cerebrocortical slices
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DOI：
10.1038/sj.jcbfm.9600335
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发表时间：
2007-02-01
影响因子：
6.3
通讯作者：
Litt, Lawrence
中科院分区：
文献类型：
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作者：
Zeng, Jianying;Yang, Guo-Yuan;Litt, Lawrence
先前对多聚（ADP-核糖）聚合酶（PARP-1）过度激活的神经元和神经胶质细胞培养研究发现，NAD+耗尽、糖酵解停滞和细胞死亡可以通过外源三羧酸循环（TCA）代谢物，尤其是丙酮酸（pyr）来避免。丙酮酸神经保护作用归因于胞质 NAD+ 补充、TCA 代谢和抗氧化活性。我们研究了暴露于 H2O2 1 小时以激活 PARP-1 后大脑皮质切片呼吸的前两种机制。 H2O2 之后用含氧人工脑脊液灌注恢复 4 小时，其中： (1) 不含葡萄糖 (glc) 或丙酮酸盐； (2) 仅10mmol/L glc； (3) 仅10 mmol/L丙酮酸； (4) 10 mmol/L 葡萄糖和 10 mmol/L 丙酮酸盐。通过蛋白质印迹和免疫组织化学对多聚 ADPribosylation 进行定量。高氯酸提取物采用 14.1 T P-31 核磁共振波谱进行定量。接触 H2O2 后，ATP 和 NAD(+) 下降约 50%，PCr 下降 75%，ADP/ATP 比率约增加一倍。如果 PARP 抑制剂伴随 H2O2，则 ATP 和 NAD(+) 变化几乎被消除，但 PCr 变化则不然。丙酮酸和 GLC 联合使用的恢复效果优于单独使用 GLC，具有更高的 ATP（0.161 对比 0.075，P < 0.01）和 PCr 水平（0.144 对比 0.078，P < 0.01），并且 TUNEL 和 Fluoro-Jade B 染色中的活细胞计数更高。二维[H-1-C-13] HSQC 谱显示了C-13 glc 或pyr 回收期间的代谢。丙酮酸代谢主要通过丙酮酸脱氢酶，部分通过丙酮酸羧化。 PARP 损伤脑片的丙酮酸灌注有助于补充 NAD(+)，同时提供代谢燃料。虽然这可以促进恢复，但并不能排除丙酮酸的强抗氧化作用。
Previous neuron and glial cell culture studies of excessive poly (ADP-ribose) polymerase (PARP-1) activation found NAD+ depletion, glycolytic arrest, and cell death that could be avoided by exogenous tricarboxylic acid cycle (TCA) metabolites, especially pyruvate (pyr). Pyruvate neuroprotection has been attributed to cytosolic NAD+ replenishment, TCA metabolism, and antioxidant activity. We investigated the first two mechanisms in respiring cerebrocortical slices after a 1-h H2O2 exposure to activate PARP-1. H2O2 was followed by a 4- h recovery with oxy-artificial cerebrospinal fluid superfusion having either: (1) no glucose (glc) or pyruvate; (2) 10mmol/ L glc only; (3) 10 mmol/ L pyruvate only; (4) both 10 mmol/ L glc and 10 mmol/ L pyruvate. Poly-ADPribosylation was quantified from Western blots and immunohistochemistry. Perchloric acid extracts were quantified with 14.1 T P-31 nuclear magnetic resonance spectroscopy. Just after H2O2 exposure, ATP and NAD(+) decreased by approximate to 50%, PCr decreased by 75%, and the ADP/ ATP ratio approximately doubled. ATP and NAD(+) changes, but not PCr changes, were nearly eliminated if PARP inhibitors accompanied the H2O2. Recovery with both pyruvate and glc was better than with glc alone, having higher ATP (0.161 versus 0.075, P < 0.01) and PCr levels (0.144 versus 0.078, P < 0.01), and higher viable cell counts in TUNEL and Fluoro- Jade B staining. Two- dimensional [H-1-C-13] HSQC spectra showed metabolism during recovery of C-13 glc or pyr. Pyruvate metabolism was primarily via pyruvate dehydrogenase, with some via pyruvate carboxylation. Pyruvate superfusion of PARPinjured brain slices helps replenish NAD(+) while providing metabolic fuel. Although this augments recovery, a strong antioxidant role for pyruvate has not been ruled out.