Although phenylalanine (Phe) is known to be neurotoxic in phenylketonuria (PKU), its exact pathogenetic mechanisms of brain damage are still poorly known. Furthermore, much less is known about the role of the Phe derivatives phenylacetic (PAA), phenyllactic (PLA) and phenylpyruvic (PPA) acids that also accumulate in this this disorder on PKU neuropathology. Previous in vitro and in vivo studies have shown that Phe elicits oxidative stress in brain of rodents and that this deleterious process also occurs in peripheral tissues of phenylketonuric patients. In the present study, we investigated whether Phe and its derivatives PAA, PLA and PPA separately or in combination could induce reactive oxygen species (ROS) formation and provoke DNA damage in C6 glial cells. We also tested the role of L-carnitine (L-car), which has been recently considered an antioxidant agent and easily cross the blood brain barrier on the alterations of C6 redox status provoked by Phe and its metabolites. We first observed that cell viability was not changed by Phe and its metabolites. Furthermore, Phe, PAA, PLA and PPA, at concentrations found in plasma of PKU patients, provoked marked DNA damage in the glial cells separately and when combined. Of note, these effects were totally prevented (Phe, PAA and PPA) or attenuated (PLA) by L-car pre-treatment. In addition, a potent ROS formation also induced by Phe and PAA, whereas only moderate increases of ROS were caused by PPA and PLA. Pre-treatment with L-car also prevented Phe- and PAA-induced ROS generation, but not that provoked by PLA and PPA. Thus, our data show that Phe and its major metabolites accumulated in PKU provoke extensive DNA damage in glial cells probably by ROS formation and that L-car may potentially represent an adjuvant therapeutic agent in PKU treatment.

尽管已知苯丙氨酸 (Phe) 在苯丙酮尿症 (PKU) 中具有神经毒性，但其脑损伤的确切发病机制仍知之甚少。此外，人们对 Phe 衍生物苯乙酸 (PAA)、苯乳酸 (PLA) 和苯丙酮酸 (PPA) 的作用知之甚少，这些酸也积聚在这种疾病中对 PKU 神经病理学的影响。先前的体外和体内研究表明，Phe 在啮齿动物的大脑中引起氧化应激，并且这种有害过程也发生在苯丙酮尿症患者的外周组织中。在本研究中，我们研究了 Phe 及其衍生物 PAA、PLA 和 PPA 是否单独或组合可诱导活性氧 (ROS) 形成并引起 C6 神经胶质细胞中的 DNA 损伤。我们还测试了左旋肉碱（L-car）的作用，它最近被认为是一种抗氧化剂，在 Phe 及其代谢物引起的 C6 氧化还原状态的改变上很容易穿过血脑屏障。我们首先观察到 Phe 及其代谢物不会改变细胞活力。此外，在 PKU 患者血浆中发现的浓度下，Phe、PAA、PLA 和 PPA 在单独和组合时会在神经胶质细胞中引起显着的 DNA 损伤。值得注意的是，这些影响完全被 L-car 预处理阻止（Phe、PAA 和 PPA）或减弱（PLA）。此外，Phe 和 PAA 也诱导了有效的 ROS 形成，而 PPA 和 PLA 仅引起了 ROS 的适度增加。L-car 预处理也阻止了 Phe 和 PAA 诱导的 ROS 生成，但不是由 PLA 和 PPA 引起的。因此，