Cecatto Cristiane, Amaral Alexandre Umpierrez, Leipnitz Guilhian, Castilho Roger Frigério, Wajner Moacir
Departamento de Bioquímica, Instituto de Ciências Básicas da Saúde, Universidade Federal de Rio Grande do Sul, Rua Ramiro Barcelos, 2600-Anexo, Porto Alegre, RS, 90035-003, Brazil.
Neurotox Res. 2014 Aug;26(2):168-78. doi: 10.1007/s12640-014-9460-5. Epub 2014 Feb 21.
Predominant accumulation of ethylmalonic acid (EMA) in tissues and biological fluids is a characteristic of patients affected by short chain acyl-CoA dehydrogenase deficiency and ethylmalonic encephalopathy. Neurological abnormalities are frequently found in these disorders, but the mechanisms underlying the brain injury are still obscure. Since hyperlacticacidemia is also found in many affected patients indicating a mitochondrial dysfunction; in the present work, we evaluated the in vitro and ex vivo effects of EMA plus Ca(2+) on mitochondrial integrity and redox balance in succinate-supported brain organelles. We verified that the evaluated parameters were disturbed only when EMA was associated with exogenous micromolar Ca(2+) concentrations. Thus, we found that this short chain organic acid plus Ca(2+) dissipated the membrane potential and provoked mitochondrial swelling, as well as impaired the mitochondrial Ca(2+) retention capacity, resulting in a rapid Ca(2+) release and decreased NAD(P)H matrix content. In contrast, EMA was not able to stimulate mitochondrial hydrogen peroxide generation. We also observed that all these effects were prevented by the mitochondrial Ca(2+) uptake inhibitor ruthenium red and the mitochondrial permeability transition (MPT) inhibitors cyclosporin A (CsA) and ADP. Furthermore, mitochondria isolated from rat brains after in vivo intrastriatal administration of EMA was more susceptible to Ca(2+)-induced swelling, which was fully prevented by CsA and ADP. Finally, EMA significantly decreased striatal slice viability, which was attenuated by CsA. The data strongly indicate that EMA reduced the mitochondrial threshold for Ca(2+)-induced MPT reinforcing the role of this cation in EMA-induced disruption of mitochondrial bioenergetics. It is, therefore, presumed that EMA acting synergistically with Ca(2+) compromises mitochondrial energy homeostasis in the central nervous system that may explain at least in part the neurologic alterations presented by patients with abnormal levels of this organic acid.
组织和生物体液中乙基丙二酸（EMA）的主要蓄积是短链酰基辅酶A脱氢酶缺乏症和乙基丙二酸脑病患者的特征。这些疾病中经常发现神经学异常，但脑损伤的潜在机制仍不清楚。由于在许多受影响的患者中也发现高乳酸血症，表明存在线粒体功能障碍；在本研究中，我们评估了EMA加Ca(2+)对琥珀酸支持的脑细胞器中线粒体完整性和氧化还原平衡的体外和离体作用。我们证实，仅当EMA与外源微摩尔浓度的Ca(2+)结合时，评估的参数才会受到干扰。因此，我们发现这种短链有机酸加Ca(2+)会使膜电位消散，引起线粒体肿胀，并损害线粒体Ca(2+)保留能力，导致Ca(2+)快速释放和NAD(P)H基质含量降低。相比之下，EMA不能刺激线粒体过氧化氢的产生。我们还观察到，线粒体Ca(2+)摄取抑制剂钌红以及线粒体通透性转换（MPT）抑制剂环孢素A（CsA）和ADP可防止所有这些作用。此外，体内纹状体内注射EMA后从大鼠脑分离的线粒体对Ca(2+)诱导的肿胀更敏感，而CsA和ADP可完全防止这种肿胀。最后，EMA显著降低纹状体切片活力，而CsA可减轻这种降低。数据强烈表明，EMA降低了Ca(2+)诱导MPT的线粒体阈值，加强了该阳离子在EMA诱导的线粒体生物能量学破坏中的作用。因此，推测EMA与Ca(2+)协同作用会损害中枢神经系统中的线粒体能量稳态，这可能至少部分解释了这种有机酸水平异常的患者出现的神经学改变。
