Baguma Marius, Kessels Sofie, Bito Virginie, Brône Bert, Triller Antoine, Maynard Stéphanie, Legendre Pascal, Rigo Jean-Michel, Le Corronc Hervé, Chabwine Joelle Nsimire
UHasselt, Neuroscience Research Group (NEURO), BIOMED, Agoralaan, 3590 Diepenbeek, Belgium; Université Catholique de Bukavu (UCB), Center for Tropical Diseases and Global Health (CTDGH), Bukavu, Democratic Republic Congo; Hôpital Provincial Général de Référence de Bukavu (HPGRB), Neurology Ward, Bukavu, Democratic Republic Congo.
UHasselt, Neuroscience Research Group (NEURO), BIOMED, Agoralaan, 3590 Diepenbeek, Belgium.
Neurotoxicology. 2024 Dec;105:323-333. doi: 10.1016/j.neuro.2024.11.004. Epub 2024 Nov 26.
Chronic cassava-derived cyanide poisoning is associated with the appearance of konzo, a tropical spastic paraparesis due to selective upper motor neuron damage. Whether the disease is caused by a direct action of cyanide or its metabolites is still an open question. This preliminary study assessed the neurotoxic effects of thiocyanate (SCN) and cyanate (OCN), two cyanide metabolites hypothesized to be plausible toxic agents in konzo.
Cultured mouse neuroblastoma (Neuro-2A) and human neuroblastoma (SH-SY5Y) cell lines were incubated (24, 48, and 72 hours) in sodium OCN or sodium SCN in a disease-relevant concentration range. Cell viability, caspase (3, 8, and 9) activities, and reactive oxygen species (ROS) generation were evaluated using appropriate assay kits. Additionally, electrophysiological responses induced by OCN and SCN in primary spinal cord neurons (from Sprague Dawley rats) were assessed by whole-cell patch-clamp techniques.
Both OCN and SCN were toxic in a dose-dependent way, even if SCN toxicity appeared at very high concentrations (30 mM, corresponding to more than 100-fold higher than normal plasmatic levels), contrary to OCN (0.3-3 mM). OCN was markedly more toxic in a poor culture medium (MEM; IC50 = 3.2 mM) compared to a glucose- and amino acid-rich medium (DMEM; IC50=7.6 mM). OCN treatment increased the ROS generation by 8.9 folds, as well as the Caspase-3, Caspase-8, and Caspase-9 activities by 3.2, 2.5, and 2.6 folds, respectively. Finally, OCN (and SCN to a lesser extent) induced depolarizing currents in primary spinal cord neurons, through an activation of ionotropic glutamate receptors.
Our results suggest OCN as the most plausible neurotoxic agent involved in konzo, while SCN toxicity could be questioned at such high concentrations. Also, they support apoptosis, oxidative stress, and excitotoxicity as probable mechanisms of OCN neurotoxicity.
慢性木薯衍生氰化物中毒与konzo的出现有关，konzo是一种由于选择性上运动神经元损伤导致的热带痉挛性截瘫。该疾病是由氰化物或其代谢产物的直接作用引起的，这仍然是一个悬而未决的问题。这项初步研究评估了硫氰酸盐（SCN）和氰酸盐（OCN）的神经毒性作用，这两种氰化物代谢产物被认为可能是konzo中的有毒物质。
将培养的小鼠神经母细胞瘤（Neuro-2A）和人神经母细胞瘤（SH-SY5Y）细胞系在与疾病相关的浓度范围内，用氰酸钠或硫氰酸钠孵育（24、48和72小时）。使用适当的检测试剂盒评估细胞活力、半胱天冬酶（3、8和9）活性以及活性氧（ROS）生成。此外，通过全细胞膜片钳技术评估了氰酸钠和硫氰酸钠对原代脊髓神经元（来自Sprague Dawley大鼠）诱导的电生理反应。
OCN和SCN均呈剂量依赖性毒性，尽管SCN毒性在非常高的浓度（30 mM，比正常血浆水平高100多倍）时才出现，而OCN的毒性浓度为0 . 3 - 3 mM。与富含葡萄糖和氨基酸的培养基（DMEM；IC50 = 7.6 mM）相比，在贫培养基（MEM；IC50 = 3.2 mM）中OCN的毒性明显更大。OCN处理使ROS生成增加了8.9倍，半胱天冬酶-3、半胱天冬酶-8和半胱天冬酶-9活性分别增加了3.2倍、2.5倍和2.6倍。最后，OCN（以及程度较轻的SCN）通过激活离子型谷氨酸受体在原代脊髓神经元中诱导去极化电流。
我们的结果表明OCN是konzo中最可能的神经毒性物质，而SCN在如此高的浓度下的毒性可能存疑。此外，这些结果支持细胞凋亡、氧化应激和兴奋性毒性是OCN神经毒性的可能机制。
