Moura Alana Pimentel, Grings Mateus, Marcowich Gustavo Flora, Bumbel Anna Paula, Parmeggiani Belisa, de Moura Alvorcem Leonardo, Wajner Moacir, Leipnitz Guilhian
Departamento de Bioquímica, Instituto de Ciências Básicas da Saúde, Universidade Federal de Rio Grande do Sul, Rua Ramiro Barcelos N° 2600 - Attached, Porto Alegre, RS, CEP 90035-003, Brazil.
Mol Cell Biochem. 2014 Oct;395(1-2):125-34. doi: 10.1007/s11010-014-2118-z. Epub 2014 Jun 18.
Patients with non-ketotic hyperglycinemia (NKH) present severe neurological symptoms and brain abnormalities involving cerebellum. Although the pathomechanisms underlying the cerebellum damage have not been studied, high tissue levels of glycine (GLY), the biochemical hallmark of this disorder have been suggested to contribute to the neuropathology of this disease. We investigated the in vitro effects of GLY on important parameters of oxidative stress and energy metabolism in cerebellum of 30-day-old rats. Our results show that GLY increased 2',7'-dichlorofluorescin oxidation, suggesting that reactive species production are augmented by GLY in the cerebellum. However, hydrogen peroxide generation was not altered by GLY. GLY also increased thiobarbituric acid-reactive substances (TBA-RS) levels and reduced the glutathione (GSH) content, indicating that this amino acid provokes lipid oxidative damage and compromises the non-enzymatic antioxidant defenses, respectively, in cerebellum. The antioxidants melatonin and trolox (the hydrosoluble analog of vitamin E) prevented the GLY-induced increase of TBA-RS and decrease of GSH in cerebellum, indicating the involvement of hydroxyl and peroxyl radicals in these effects. The NMDA receptor antagonist MK-801 also attenuated GLY-induced decrease of GSH, suggesting that this effect is mediated through NMDA receptor. In contrast, GLY did not alter the protein carbonyl formation and total and protein-bound sulfhydryl group content, as well as catalase and superoxide dismutase activities. Furthermore, GLY did not alter the activities of the respiratory chain complexes and creatine kinase. Our present data indicate that oxidative stress elicited by GLY in vitro may be a potential pathomechanism involved in the cerebellar dysfunction observed in NKH.
非酮症高甘氨酸血症(NKH)患者会出现严重的神经症状以及涉及小脑的脑部异常。尽管小脑损伤的发病机制尚未得到研究,但这种疾病的生化标志——高组织水平的甘氨酸(GLY),被认为与该疾病的神经病理学有关。我们研究了GLY对30日龄大鼠小脑氧化应激和能量代谢重要参数的体外影响。我们的结果表明,GLY增加了2',7'-二氯荧光素的氧化,这表明GLY在小脑中增加了活性物质的产生。然而,过氧化氢的生成并未因GLY而改变。GLY还增加了硫代巴比妥酸反应性物质(TBA-RS)的水平,并降低了谷胱甘肽(GSH)的含量,这表明这种氨基酸分别在小脑中引发了脂质氧化损伤并损害了非酶抗氧化防御。抗氧化剂褪黑素和生育三烯酚(维生素E的水溶性类似物)可预防GLY诱导的小脑中TBA-RS增加和GSH减少,这表明羟基和过氧自由基参与了这些作用。NMDA受体拮抗剂MK-801也减弱了GLY诱导的GSH减少,表明这种作用是通过NMDA受体介导的。相比之下,GLY并未改变蛋白质羰基的形成、总巯基和蛋白质结合巯基的含量,以及过氧化氢酶和超氧化物歧化酶的活性。此外,GLY并未改变呼吸链复合物和肌酸激酶的活性。我们目前的数据表明,GLY在体外引发的氧化应激可能是NKH中观察到的小脑功能障碍的潜在发病机制。