Intranasal instillation of aflatoxin B(1) in rats: bioactivation in the nasal mucosa and neuronal transport to the olfactory bulb.

Aflatoxin B(1) (AFB(1)) may be present in moldy dust. Inhalation of contaminated dust particles may result in high local exposure of the nasal mucosa. The present study was designed to assess bioactivation and toxicity of AFB(1) in the nasal mucosa after intranasal administration of the mycotoxin in rats and also to examine if translocation of the mycotoxin occurs from the nasal mucosa to the brain along olfactory neurons. Female Sprague-Dawley rats were given (3)H-AFB(1) (0.2, 1 or 20 microg) intranasally and were sacrificed at various intervals (1 h to 20 d). Tissues were examined autoradiographically or histopathologically. Quantitative data were obtained by beta-spectrometry in rats given (3)H-AFB(1) intranasally or orally (for comparison). The data indicated that intranasal administration of AFB(1) resulted in formation of tissue-bound metabolites in sustentacular cells, in some cells of Bowman's glands, and in a population of neuronal cells in the olfactory mucosa, whereas in the respiratory nasal mucosa, there was selective bioactivation of AFB(1) in mucous cells. Intranasal instillation of 20 microg AFB(1) resulted in disorganized undulating olfactory epithelium, with injured neuronal and sustentacular cells. In the respiratory epithelium, there was selective destruction of mucous cells. beta-Spectrometry and autoradiography with tape-sections of the head of rats given (3)H-AFB(1) intranasally indicated transport of AFB(1) and/or AFB(1) metabolites along the axons of the primary olfactory neurons to their terminations in the glomeruli of the olfactory bulb. The data indicate that the materials transported in the olfactory nerves represent AFB(1) and/or some of its nonreactive metabolites. It is concluded that application of AFB(1) on the nasal mucosa in rats results in high local bioactivation of the mycotoxin in this tissue and translocation of AFB(1) and/or its metabolites to the olfactory bulb.     

中药材中真菌毒素的检测与脱毒研究进展

中药材在种植、采收、贮存及加工各个环节中容易受到真菌的污染,产生一系列有毒次生代谢产物（真菌毒素）,直接影响药材质量的安全性与有效性,还会严重威胁人体生命健康。针对中药材中潜在的多种真菌毒素,建立高效的净化富集与灵敏、专属的检测方法,对提升中药质量标准、保障中药用药安全具有重要意义。通过对国内外重要真菌毒素的限量标准、中药材中真菌毒素检测使用的样品前处理方法、关键检测技术及降解脱毒方法进行综述,并展望中药材真菌毒素研究的发展趋势,为保障中药材的质量安全提供参考。     

Congenital aflatoxicosis,mal-detoxification genomics & ontogeny trigger immune-mediated Kotb disease biliary atresia variant: SANRA compliant review

Biliary atresia (BA) is the most common indication for pediatric liver transplantation. We describe The BA variant: Kotb disease. Liver tissue in the Kotb disease BA is massively damaged by congenital aflatoxicosis resulting in inflammation, adhesions, fibrosis, bile duct proliferation, scarring, cholestasis, focal syncytial giant cell transformation, and typical immune response involving infiltration by CD4+, CD8+, CD68+, CD14+, neutrophil infiltration, neutrophil elastase spill, heavy loads of aflatoxin B1, accelerated cirrhosis, disruption of p53 and GSTPi, and have null glutathione S transferase M1 (GSTM1). All their mothers are heterozygous for GSTM1. This inability to detoxify aflatoxicosis results in progressive inflammatory adhesions and obliterative cholangiopathy early in life. The typical disruption of both p53 and GSTPi causes loss of fidelity of hepatic regeneration. Hence, regeneration in Kotb disease BA typically promotes accelerated cirrhosis. The immune response in Kotb disease BA is for damage control and initiation of regeneration, yet, this friendly fire incurs massive structural collateral damage. The Kotb disease BA is about actual ongoing hepatic entrapment of aflatoxins with lack of ability of safe disposal due to child detoxification-genomics disarray.The Kotb disease BA is a product of the interaction of persistent congenital aflatoxicosis, genetic lack of GSTM1 detoxification, ontogenically impaired activity of other hepatic detoxification, massive neutrophil-elastase, immune-induced damage, and disturbed regeneration. Ante-natal and neonatal screening for aflatoxicosis, avoiding cord milking, and stringent control of aflatoxicosis content of human, poultry and live-stock feeds might prove effective for prevention, prompt diagnosis and management based on our recent understanding of its patho-genomics.     

Assessing the Impacts of Preanalytical Field Sampling Challenges on the Reliability of Serum Aflatoxin B1-Lysine Measurements by Use of LC-MS/MS

Aflatoxin exposure is endemic in developing countries with warm, humid climates that promote toxigenic mold growth on crops and foodstuffs. Estimating human aflatoxin exposure is key to identifying and abating contamination sources. Serum aflatoxin B1 bound to albumin lysine (AFB1-lys) is a preferred exposure biomarker, but field sample collection, processing, transportation, and storage logistics are challenging. We validated an improved LC-MS/MS method for serum AFB1-lys and applied it to three field sampling challenges: transportation/storage (elevated temperature); collection/processing (hemolysis); and sample type substitution (heparinized plasma). Our new LC-MS/MS method had a LOD of 0.03 ng/mL, accuracy (mean spike recovery) of 112%, total imprecision (replicate pool measurements) ≤5% at ≥0.2 ng/mL, and results that were 95.1% similar (mean percentage similarity) to an established method. AFB1-lys in human serum spiked with serum from aflatoxin-dosed rats was stable for 14 days at both ambient (22.5 °C) and elevated (38 °C) temperatures. Simulated hemolysis (adding 0.25–3 mg hemoglobin) did not affect AFB1-lys accuracy at ≥0.5 ng/mL but caused 10–25% signal suppression. Heparinized plasma AFB1-lys was 99.0% similar to serum but interfered with albumin measurements (bromocresol green) causing spurious low bias. Further investigation is warranted, but our findings suggest that AFB1-lys is pre-analytically robust.     

银杏叶对黄曲霉毒素B1诱发大鼠肝癌IGF-ⅡmRNA及蛋白表达影响的观察

目的：动态观察银杏叶提取物（EGb761）在抑制黄曲霉毒素B1（AFB1）诱发大鼠肝癌（HCO）过程中对肝组织相关基因IGF-ⅡmRNA及蛋白表达水平的影响,从分子生物学水平揭示银杏叶抗癌的机制及其效果。方法：将70只4周龄Wistar雄性大鼠随机分为AFB1组（25只）、AFB1＋EGb761组（25只）及对照组（20只）。在诱发大鼠HCC过程中,分别于第13周、33周及53周对大鼠进行肝组织活检;实验至第73周处死全部动物取肝组织。利用实时荧光定量PCR和蛋白质印迹法动态检测肝组织中IGF-ⅡmRNA及相应蛋白的表达情况。结果：AFB1组原发性HCC发生率为58．8％（10／17）;AFB1＋EGb761组发生率为29．4％（5／17）;对照组为0（o／16）。AFBl＋EGb761组HCC发生率显著低于AFB1组（P〈0．05）。AFBl＋EGb761组肝组织IGF_2mRNA及相应蛋白表达扮平在第53周及73周较AFB1组均显著下降,差异有统计学意义,P％0．05。结论：银杏叶提取物（EGb761）具有抑制AFBl致大鼠HCC的作用。其机制可能与调控肝细胞增殖基因IGF-ⅡmRNA及蛋白表达水平有关。     

Urinary Aflatoxin M1 Concentration and Its Determinants in School-Age Children in Southern Ethiopia

Aflatoxins are mycotoxins that can contaminate grains, legumes, and oil seeds. These toxic compounds are an especially serious problem in tropical and sub-tropical climates. The objective of this study was to raise awareness of aflatoxin exposure among primary school children in Shebedino woreda, southern Ethiopia, by measuring urinary aflatoxin M1 (AFM1). The study employed a cross-sectional design and systematic random sampling of children from eight schools in the district. The mean ± SD age of the children was 9.0 ± 1.8 years. Most (84.6%) households were food insecure with 17.9% severely food insecure. Urinary AFM1 was detected in more than 93% of the children. The median [IQR] concentration of AFM1/Creat was 480 [203, 1085] pg/mg. Based on a multiple regression analysis: DDS, consumption of haricot bean or milk, source of drinking water, maternal education, and household food insecurity access scale scores were significantly associated with urinary AFM1/Creat. In conclusion, a high prevalence of urinary AFM1 was observed in this study. However, the relation between AFM1 and dietary intake was analyzed based on self-reported dietary data; hence, all of the staple foods as well as animal feeds in the study area should be assessed for aflatoxin contamination.     

Design of a Diagnostic Immunoassay for Aflatoxin M1 Based on a Plant-Produced Antibody

A new green competitive ELISA for aflatoxin M1 quantification in raw milk was developed. This diagnostic tool is based on an anti AFM1 mAb produced by plant molecular farming in alternative to classical systems. Our assay, showing an IC₅₀ below 25 ng/L, fits with the requirements of EU legislation limits for AFM1 (50 ng/L). Optimal accuracy was achieved in correspondence of the decision levels (25 and 50 ng/L), and the assay enabled AFM1 quantification in the range 5–110 ng/L, with limit of detection 3 ng/L. Moreover, to evaluate a real applicability in diagnostics, raw milk-spiked samples were analysed, achieving satisfactory recovery rates of AFM1. In conclusion, an efficient and ready-to-use diagnostic assay for the quantification of aflatoxin M1 in milk, based on a plant-produced recombinant mAb, has been successfully developed.     

A longitudinal assessment of aflatoxin M1 excretion in breast milk of selected Egyptian mothers.

Aflatoxins are potent toxins and carcinogens which can be excreted in the milk of exposed lactating mothers mainly in the form of aflatoxin M(1) (AFM(1)). We previously evaluated the level and frequency of AFM(1) in breast milk in a group of Egyptian mothers attending the New El-Qalyub Hospital, Qalyubiyah governorate, Egypt. In this study, fifty of those women who were AFM(1) positive were revisited monthly for 12 months to assess the temporal variation in breast milk AFM(1). AFM(1) was detected in 248 of 443 (56%) samples. In a multilevel model of the data there was a highly significant (p<0.001) effect of month of sampling on the frequency of AFM(1) detection with summer months having the highest frequency (>80%) and winter months the lowest frequency (<20%) of detection. AFM(1) was observed most frequently in June [OR 63, 95% CI (7.6, 522)]. The level of AFM(1) detection also followed this seasonal pattern with highest mean level in July (64 pg/ml milk, range 6.3-497 pg/ml milk) and the lowest mean level in January (8 pg/ml milk, range 4.2-108 pg/ml milk). The duration of lactation [p=0.0035, OR=1.08, 95% CI (1.02, 1.13)], and peanut consumption [p=0.06, OR=1.69, 95% CI (0.9, 2.9)] also contributed to the model. The identification and understanding of factors determining the presence of toxicants in human milk is important and may provide a knowledge driven basis for controlling the transfer of chemicals to infants.