Mehrzad Jalil, Malvandi Amir Mohammad, Alipour Mohsen, Hosseinkhani Saman
Department of Microbiology and Immunology, Faculty of Veterinary Medicine, University of Tehran, Tehran, Iran.
Institute of Experimental Neurology (INSPE), San Raffaele Hospital, Via Olgettina 58, Milan, Italy.
Toxicol Lett. 2017 Sep 5;279:96-106. doi: 10.1016/j.toxlet.2017.07.902. Epub 2017 Aug 5.
Aflatoxin B (AFB) is a well-known member of aflatoxins (AFs) that is considered among highly stable toxic contaminants of food, worldwide. The impact of AFB on neural cells and systems has poorly been understood. To assess the cellular effects of AFB on brain, we used murine pure primary astrocytes, sub ventricular zone-derived neural precursor cells (NPCs) and microglia cell line (BV2). Cells were exposed separately to environmentally relevant level (20ng/ml) of AFB for 1, 2, 3, 6, 12, 24 and 48h in culture. At each time points, total free radicals production measured by luminol-enhanced cellular chemiluminescence (CL) assay; cytokines production of IL-1β, IL-6, TNF-α and IL-10 were analyzed using Bioplex ELISA and a set of genes involved in the immediate response to danger such as TLR2, TLR4 and iNOS etc. were evaluated by multiplex qPCR. Upon AFB exposure production, of the total free radicals significantly increased only in microglial cells after 24h and slightly elevated in the other examined cells. AFB also induced secretion of pro-inflammatory cytokines (i.e. TNF-α and IL-6) on both microglial cells (more TNF-α) and astrocytes (more IL-6). mRNA expression of TLR2, TLR4, MyD88 and NF-κB were up-regulated with different timing and levels among cells. Immunotoxicologically, microglial cells, and astrocytes, but not NPCs, are capable of sensing a low level of AFB. Thus, the pro-inflammatory effects of an environmentally relevant dose of AFB on CNS-derived cells in vitro could potentially explain the immune dysregulation in neurodegenerative disorders.
黄曲霉毒素B（AFB）是黄曲霉毒素（AFs）中一种广为人知的毒素，被认为是全球食品中高度稳定的有毒污染物之一。AFB对神经细胞和系统的影响目前了解甚少。为了评估AFB对大脑的细胞效应，我们使用了小鼠纯原代星形胶质细胞、脑室下区来源的神经前体细胞（NPCs）和小胶质细胞系（BV2）。将细胞分别暴露于环境相关水平（20ng/ml）的AFB中，在培养中培养1、2、3、6、12、24和48小时。在每个时间点，通过鲁米诺增强细胞化学发光（CL）测定法测量总自由基产生；使用Bioplex ELISA分析IL-1β、IL-6、TNF-α和IL-10的细胞因子产生，并通过多重qPCR评估一组参与对危险即时反应的基因，如TLR2、TLR4和iNOS等。暴露于AFB后，仅在24小时后小胶质细胞中的总自由基产生显著增加，而在其他检测细胞中略有升高。AFB还诱导小胶质细胞（更多TNF-α）和星形胶质细胞（更多IL-6）分泌促炎细胞因子（即TNF-α和IL-6）。TLR2、TLR4、MyD88和NF-κB的mRNA表达在细胞中以不同的时间和水平上调。免疫毒理学研究表明，小胶质细胞和星形胶质细胞能够感知低水平的AFB，而NPCs则不能。因此，环境相关剂量的AFB对体外中枢神经系统来源细胞的促炎作用可能潜在地解释神经退行性疾病中的免疫失调。
