Effects of aflatoxin and fumonisin on gene expression of growth factors and inflammation-related genes in a human hepatocyte cell line.

Aflatoxin B1 (AFB1) and fumonisin B1 (FB1) are mycotoxins widely distributed in maize and maized-based products, often occurring together. The implications of co-exposure to aflatoxin and fumonsin for human health are numerous, but a particular concern is the potential of FB1 to modulate AFB1 hepatotoxicity. This study evaluated the toxicity of these mycotoxins, alone or combined, in a human non-tumorigenic liver cell line, HHL-16 cells, and assessed the effects of AFB1 and FB1 on expression of genes involved in immune and growth factor pathways. The results demonstrated that in HHL-16 cells, both AFB1 and FB1 had dose-dependent and time-dependent toxicity, and the combination of them showed a synergistic toxicity in the cells. Moreover, AFB1 caused upregulation of IL6, CCL20, and BMP2, and downregulation of NDP. In combination of AFB1 with FB1, gene expression levels of IL6 and BMP2 were significantly higher compared to individual FB1 treatment, and had a tendency to be higher than individual AFB1 treatment. This study shows that FB1 may increase the hepatoxicity of AFB1 through increasing the inflammatory response and disrupting cell growth pathways.

Highly resistant and sensitive colorimetric immunochromatographic assay for sibutramine (SBT) illegally adulterated into diet food based on PDA/AuNP labelling.

A gold nanoparticle (AuNP) based immunochromatographic assay strip is a valuable tool for monitoring chemicals in foods. However, the sensitive ICA strip for SBT is rarely reported due to the fact that monoclonal antibodies (mAbs) against SBT with high affinity are commercially unavailable. Herein, a monoclonal antibody against SBT was prepared through a designed hapten with a carboxyl end-capped space arm. The obtained mAb showed high affinity for SBT and N-desmethylsibutramine, a metabolite of SBT. Furthermore, a series of core-shell NPs, polydopamine (PDA) coated AuNPs (PDA/AuNPs) with controlled shell thickness and packing density were synthesized. The obtained PDA/AuNP-mAb conjugate demonstrated high tolerance to salt and good stability in a wide pH range, which is beneficial for improving the matrix interference common in ICA. As a result, PDA/AuNP-based ICA could quantify SBT in the range of 3.39-69.60 ng mL-1 with a limit of detection (LOD) of 0.98 ng mL-1. This novel ICA improved the sensitivity of the traditional AuNP-based ICA by nearly 12 times. Method validation was conducted with spiked samples of slimming food and human serum and compared with HPLC-MS/MS. Consistent results indicated that high sensitivity, accuracy, and reliability of the PDA/AuNP-based ICA approach were achieved. To the best of our knowledge, this study reported the most sensitive immunoassay for SBT thus far.

An aggregation-induced emission immunoassay for broad detection of polychlorinated biphenyls in chicken and crab.

Polychlorinated biphenyls (PCBs) are persistent organic pollutants (POPs) with multiple variants, which may be harmful to human health by absorption and bioaccumulation. To ensure food safety, it is necessary to develop multi-residue immunoassays for broad recognition of PCBs. In this study, by mimicking the generic core structure of PCBs, three haptens have been designed and synthesized for monoclonal antibody (mAb) generation. A carboxylic acid derivative of PCB80 was a hapten that induced a mAb with broad recognition of PCBs. The results of ELISA further identified that the mAb could recognize 11 different kinds of PCBs; half-maximal inhibition concentrations (IC50) ranged from 33.12 to 476.42 ng/mL. Subsequently, using aggregation-induced emission luminogen (AIEgen) nanobeads as the tracer for the output signal, the IC50 value of the various PCBs was improved to 6.38-252.1 ng/mL. The limit of detection (LOD) varied from 0.32 to 42.15 ng/mL. Recoveries of 76.90-95.74% and intra-assay coefficients of variation of 8.5-14.4% were obtained with spiked chicken and crab meat samples. Matrix interference was eliminated by dilution, and no false-positive and false-negative results were observed. The developed assay provides a simple, broad-spectrum, and sensitive tool for detecting PCBs, with high-throughput possibilities for large-scale screening of PCBs in food.

Organ Specific Differences in Alteration of Aquaporin Expression in Rats Treated with Sennoside A, Senna Anthraquinones and Rhubarb Anthraquinones.

Senna and rhubarb are often used as routine laxatives, but there are differences in mechanism of action and potential side effects. Here, we studied metabolites of senna anthraquinones (SAQ), rhubarb anthraquinones (RAQ) and their chemical marker, sennoside A (SA), in a rat diarrhea model. In in vitro biotransformation experiments, SAQ, RAQ and SA were incubated with rat fecal flora solution and the metabolites produced were analyzed using HPLC. In in vivo studies, the same compounds were investigated for purgation induction, with measurement of histopathology and Aqps gene expression in six organs. The results indicated that SAQ and RAQ had similar principal constituents but could be degraded into different metabolites. A similar profile of Aqps down-regulation for all compounds was seen in the colon, suggesting a similar mechanism of action for purgation. However, in the kidneys and livers of the diarrhea-rats, down-regulation of Aqps was found in the RAQ-rats whereas up-regulation of Aqps was seen in the SAQ-rats. Furthermore, the RAQ-rats showed lower Aqp2 protein expression in the kidneys, whilst the SA-rats and SAQ-rats had higher Aqp2 protein expression in the kidneys. This may have implications for side effects of SAQ or RAQ in patients with chronic kidney or liver diseases.

Aflatoxin Exposure during Early Life Is Associated with Differential DNA Methylation in Two-Year-Old Gambian Children.

Background: DNA methylation is an epigenetic control mechanism that may be altered by environmental exposures. We have previously reported that in utero exposure to the mycotoxin and liver carcinogen aflatoxin B1 from the maternal diet, as measured using biomarkers in the mothers' blood, was associated with differential DNA methylation in white blood cells of 6-month-old infants from The Gambia. Methods: Here we examined aflatoxin B1-associated differential DNA methylation in white blood cells of 24-month-old children from the same population (n = 244), in relation to the child's dietary exposure assessed using aflatoxin albumin biomarkers in blood samples collected at 6, 12 and 18 months of age. HM450 BeadChip arrays were used to assess DNA methylation, with data compared to aflatoxin albumin adduct levels using two approaches; a continuous model comparing aflatoxin adducts measured in samples collected at 18 months to DNA methylation at 24 months, and a categorical time-dose model that took into account aflatoxin adduct levels at 6, 12 and 18 months, for comparison to DNA methylation at 24 months. Results: Geometric mean (95% confidence intervals) for aflatoxin albumin levels were 3.78 (3.29, 4.34) at 6 months, 25.1 (21.67, 29.13) at 12 months and 49.48 (43.34, 56.49) at 18 months of age. A number of differentially methylated CpG positions and regions were associated with aflatoxin exposure, some of which affected gene expression. Pathway analysis highlighted effects on genes involved with with inflammatory, signalling and growth pathways. Conclusions: This study provides further evidence that exposure to aflatoxin in early childhood may impact on DNA methylation.

Recent trends in detecting, controlling, and detoxifying of patulin mycotoxin using biotechnology methods.

Patulin (PAT) is a mycotoxin that can contaminate many foods and especially fruits and fruit-based products. Therefore, accurate and effective testing is necessary to enable producers to comply with regulations and promote food safety. Traditional approaches involving the use of chemical compounds or physical treatments in food have provided practical methods that have been used to date. However, growing concerns about environmental and health problems associated with these approaches call for new alternatives. In contrast, recent advances in biotechnology have revolutionized the understanding of living organisms and brought more effective biological tools. This review, therefore, focuses on the study of biotechnology approaches for the detection, control, and mitigation of PAT in food. Future aspects of biotechnology development to overcome the food safety problem posed by PAT were also examined. We find that biotechnology advances offer novel, more effective, and environmental friendly approaches for the control and elimination of PAT in food compared to traditional methods. Biosensors represent the future of PAT detection and use biological tools such as aptamer, enzyme, and antibody. PAT prevention strategies include microbial biocontrol, the use of antifungal biomolecules, and the use of microorganisms in combination with antifungal molecules. PAT detoxification aims at the breakdown and removal of PAT in food by using enzymes, microorganisms, and various adsorbent biopolymers. Finally, biotechnology advances will be dependent on the understanding of fundamental biology of living organisms regarding PAT synthesis and resistance mechanisms.

Comparison of urinary aflatoxin M1 and aflatoxin albumin adducts as biomarkers for assessing aflatoxin exposure in Tanzanian children.

PURPOSE: To determine levels of urinary aflatoxin M1 (AFM1) in children and correlate the concentrations with previously reported aflatoxin albumin adduct (AF-alb) levels in these children. MATERIALS AND METHODS: Matched urine and blood samples were collected from 84 Tanzanian children aged 6-14 months old. From 31 children in one village (Kigwa), samples were collected at three time points six months apart. Samples were collected from 31 and 22 children from two different regions at the second time point only. Urinary AFM1 was measured using a commercial enzyme-linked immunosorbent assay (ELISA) kit with a modified protocol to improve sensitivity. AF-alb was measured using an established ELISA method. RESULTS: The relative ranking of the three villages for exposure to aflatoxin based on either AFM1 or AF-alb biomarker measurements was the same. In Kigwa village, both AFM1 and AF-alb levels were higher at six months post-harvest compared to baseline. However, at the next visit, the AFM1 levels dropped from a GM (interquartile range) of 71.0 (44.7, 112.6) at visit two to 49.3 (31.5, 77.3) pg/ml urine, whereas AF-alb levels increased from 47.3 (29.7, 75.2) to 52.7 (35.4, 78.3) pg/mg albumin between these two visits, reflecting the fact that AFM1 measures short-term exposure, whereas AF-alb measures longer term exposure. There was a correlation between AFB1 intake and AFM1 excretion (r= 0.442, p ≤ 0.001). CONCLUSIONS: Urinary AFM1 is a good biomarker for AFB1 exposure in Tanzanian children, reflecting geographical and temporal variations in exposure to this foodborne toxin.

Impaired growth in rural Gambian infants exposed to aflatoxin: a prospective cohort study.

BACKGROUND: Exposure to aflatoxin, a mycotoxin produced by fungi that commonly contaminates cereal crops across sub-Saharan Africa, has been associated with impaired child growth. We investigated the impact of aflatoxin exposure on the growth of Gambian infants from birth to two years of age, and the impact on insulin-like growth factor (IGF)-axis proteins. METHODS: A subsample (N = 374) of infants from the Early Nutrition and Immune Development (ENID) trial (ISRCTN49285450) were included in this study. Aflatoxin-albumin adducts (AF-alb) were measured in blood collected from infants at 6, 12 and 18 months of age. IGF-1 and IGFBP-3 were measured in blood collected at 12 and 18 months. Anthropometric measurements taken at 6, 12, 18 and 24 months of age were converted to z-scores against the WHO reference. The relationship between aflatoxin exposure and growth was analysed using multi-level modelling. RESULTS: Inverse relationships were observed between lnAF-alb and length-for-age (LAZ), weight-for-age (WAZ), and weight-for-length (WLZ) z-scores from 6 to 18 months of age (ß = - 0·04, P = 0·015; ß = - 0·05, P = 0.003; ß = - 0·06, P = 0·007; respectively). There was an inverse relationship between lnAF-alb at 6 months and change in WLZ between 6 and 12 months (ß = - 0·01; P = 0·013). LnAF-alb at 12 months was associated with changes in LAZ and infant length between 12 and 18 months of age (ß = - 0·01, P = 0·003; ß = - 0·003, P = 0·02; respectively). LnAF-alb at 6 months was associated with IGFBP-3 at 12 months (r = - 0·12; P = 0·043). CONCLUSIONS: This study found a small but significant effect of aflatoxin exposure on the growth of Gambian infants. This relationship is not apparently explained by aflatoxin induced changes in the IGF-axis.

Interventions targeting child undernutrition in developing countries may be undermined by dietary exposure to aflatoxin.

Child undernutrition, a form of malnutrition, is a major public health burden in developing countries. Supplementation interventions targeting the major micronutrient deficiencies have only reduced the burden of child undernutrition to a certain extent, indicating that there are other underlying determinants that need to be addressed. Aflatoxin exposure, which is also highly prevalent in developing countries, may be considered an aggravating factor for child undernutrition. Increasing evidence suggests that aflatoxin exposure can occur in any stage of life, including in utero through a trans-placental pathway and in early childhood (through contaminated weaning food and family food). Early life exposure to aflatoxin is associated with adverse effects on low birth weight, stunting, immune suppression, and the liver function damage. The mechanisms underlying impaired growth and aflatoxin exposure are still unclear but intestinal function damage, reduced immune function, and alteration in the insulin-like growth factor axis caused by the liver damage are the suggested hypotheses. Given the fact that both aflatoxin and child undernutrition are common in sub-Saharan Africa, effective interventions aimed at reducing undernutrition cannot be satisfactorily achieved until the interactive relationship between aflatoxin and child undernutrition is clearly understood, and an aflatoxin mitigation strategy takes effect in those vulnerable mothers and children.

Pandemic H1N1 Vaccination and Incidence of Acute Disseminated Encephalomyelitis in Manitoba.

BACKGROUND: An increased incidence of hospital admissions coded as acute disseminated encephalomyelitis (ADEM) was noted in Winnipeg, Manitoba, Canada, during the second wave of the influenza pandemic from October 2009 to March 2010. However, it was not clear whether this was due to heightened awareness of potential neurological complications of influenza or influenza vaccination or an actual increase in the number of cases. METHODS: We extracted data from the charts of 139 patients hospitalized with an International Classification of Diseases-10 discharge code indicating ADEM (G04.0) or unspecified noninfectious encephalitis or myelitis (G04.8, G04.9) between January 2006 and December 2012. Clinical and laboratory data were reviewed by a neurologist, and diagnoses were determined using the Brighton criteria. RESULTS: Over the entire study period, there were 22 cases of ADEM. During the peak pandemic period (April-December 2009), seven patients were hospitalized with ADEM, corresponding to a rate of 7.8/million/year; 4.7 (95% confidence interval: 1.9-11.4) times higher than the rate before or after the pandemic period. Only one patient with ADEM had received the monovalent A(H1N1)pdm09 vaccine within 12 weeks of hospitalization. CONCLUSIONS: We have found an increased incidence of ADEM during the pandemic period that may be related, at least in part, to the increased incidence of influenza during that period. However, there was no temporal relationship with the administration of A(H1N1)pdm09 or seasonal influenza vaccines. Our study provides reassurance that use of these vaccines was not associated with increased risk of ADEM.

Restrictive Cardiomyopathy Resulting from a Troponin I Type 3 Mutation in a Chinese Family.

Objective To identify the pathogenic variant responsible for restrictive cardiomyopathy (RCM) in a Chinese family.Methods Next generation sequencing was used for detecting the mutation and Results verified by sequencing. We used restriction enzyme digestion to test the mutation in the family members and 200 unrelated normal subjects without any cardiac inherited diseases when the mutation was identified.Results Five individuals died from cardiac diseases, two of whom suffered from sudden cardiac death. Two individuals have suffered from chronic cardiac disorders. Mutation analysis revealed a novel missense mutation in exon 7 of troponin I type 3 (TNNI3), resulting in substitution of serine (S) with proline (P) at amino acid position 150, which cosegregated with the disease in the family, which is predicted to be probably damaging using PolyPhen-2. The mutation was not detected in the 200 unrelated subjects we tested.Conclusion Using next generation sequencing, which has very recently been shown to be successful in identifying novel causative mutations of rare Mendelian disorders, we found a novel mutation of TNNI3 in a Chinese family with RCM.

Systematic investigation of the physicochemical factors that contribute to the toxicity of ZnO nanoparticles.

ZnO nanoparticles (NPs) are prone to dissolution, and uncertainty remains whether biological/cellular responses to ZnO NPs are solely due to the release of Zn(2+) or whether the NPs themselves have additional toxic effects. We address this by establishing ZnO NP solubility in dispersion media (Dulbecco's modified Eagle's medium, DMEM) held under conditions identical to those employed for cell culture (37 °C, 5% CO2, and pH 7.68) and by systematic comparison of cell-NP interaction for three different ZnO NP preparations. For NPs at concentrations up to 5.5 µg ZnO/mL, dissolution is complete (with the majority of the soluble zinc complexed to dissolved ligands in the medium), taking ca. 1 h for uncoated and ca. 6 h for polymer coated ones. Above 5.5 µg/mL, the results are consistent with the formation of zinc carbonate, keeping the solubilized zinc fixed to 67 µM of which only 0.45 µM is as free Zn(2+), i.e., not complexed to dissolved ligands. At these relatively high concentrations, NPs with an aliphatic polyether-coating show slower dissolution (i.e., slower free Zn(2+) release) and reprecipitation kinetics compared to those of uncoated NPs, requiring more than 48 h to reach thermodynamic equilibrium. Cytotoxicity (MTT) and DNA damage (Comet) assay dose-response curves for three epithelial cell lines suggest that dissolution and reprecipitation dominate for uncoated ZnO NPs. Transmission electron microscopy combined with the monitoring of intracellular Zn(2+) concentrations and ZnO-NP interactions with model lipid membranes indicate that an aliphatic polyether coat on ZnO NPs increases cellular uptake, enhancing toxicity by enabling intracellular dissolution and release of Zn(2+). Similarly, we demonstrate that needle-like NP morphologies enhance toxicity by apparently frustrating cellular uptake. To limit toxicity, ZnO NPs with nonacicular morphologies and coatings that only weakly interact with cellular membranes are recommended.

Seasonal and gestation stage associated differences in aflatoxin exposure in pregnant Gambian women.

OBJECTIVE: Aflatoxin is known to cross the placental barrier and exposures in utero could influence genomic programming, foetal growth and development, resulting in long-term health effects. We aimed to determine aflatoxin exposure in Gambian women at two stages of pregnancy and during the rainy and dry seasons. METHODS: We examined aflatoxin exposure in pregnant Gambian women at early (<16 weeks) and later (16 weeks onward) stages of pregnancy and at different times of the year, during the rainy (June to October 2009) or dry (November to May 2010) season, using aflatoxin-albumin adducts (AF-alb). RESULTS: Mean AF-alb was higher during the dry season than in the rainy season, in both early and later pregnancy although the difference was strongest in later pregnancy. There was a modest increase in AF-alb in later than early pregnancy (geometric mean 41.8 vs. 34.5 pg/mg, P < 0.05), but this was restricted to the dry season when exposures were generally higher. CONCLUSIONS: The study confirmed that Gambian pregnant women were exposed to aflatoxin throughout the pregnancy, with higher levels in the dry season. There was some evidence in the dry season that women in later pregnancy had higher AF-alb levels than those in earlier pregnancy. Further research on the effects of exposure to this potent mutagen and carcinogen throughout pregnancy, including the epigenetic modification of foetal gene expression and impact on pre- and post-natal growth and development, are merited.

A pilot study to evaluate aflatoxin exposure in a rural Ugandan population.

OBJECTIVES: The fungal metabolite aflatoxin is a common contaminant of foodstuffs, especially when stored in damp conditions. In humans, high levels can result in acute hepatic necrosis and death, while chronic exposure is carcinogenic. We conducted a pilot study nested within an existing population cohort (the General Population Cohort), to assess exposure to aflatoxin, among people living in rural south-western Uganda. METHODS: Sera from 100 adults and 96 children under 3 years of age (85 male, 111 female) were tested for aflatoxin-albumin adduct (AF-alb), using an ELISA assay. Socio-demographic and dietary data were obtained for all participants; HIV serostatus was available for 90 adults and liver function tests (LFTs) for 99. RESULTS: Every adult and all but four children had detectable AF-alb adduct, including five babies reported to be exclusively breastfed. Levels ranged from 0 to 237.7 pg/mg albumin and did not differ significantly between men and women, by age or by HIV serostatus; 25% had levels above 15.1 pg/mg albumin. There was evidence of heterogeneity between villages (P = 0.003); those closest to trading centres had higher levels. Adults who consumed more Matooke (bananas) had lower levels of AF-alb adduct (P = 0.02) than adults who did not, possibly because their diet contained fewer aflatoxin-contaminated foods such as posho (made from maize). Children who consumed soya, which is not grown locally, had levels of AF-alb adduct that were almost twice as high as those who did not eat soya (P = 0.04). CONCLUSIONS: Exposure to aflatoxin is ubiquitous among the rural Ugandans studied, with a significant number of people having relatively high levels. Sources of exposure need to be better understood to instigate practical and sustainable interventions.

Variation of DNA damage levels in peripheral blood mononuclear cells isolated in different laboratories.

This study investigated the levels of DNA strand breaks and formamidopyrimidine DNA glycosylase (FPG) sensitive sites, as assessed by the comet assay, in peripheral blood mononuclear cells (PBMC) from healthy women from five different countries in Europe. The laboratory in each country (referred to as 'centre') collected and cryopreserved PBMC samples from three donors, using a standardised cell isolation protocol. The samples were analysed in 13 different laboratories for DNA damage, which is measured by the comet assay. The study aim was to assess variation in DNA damage in PBMC samples that were collected in the same way and processed using the same blood isolation procedure. The inter-laboratory variation was the prominent contributor to the overall variation. The inter-laboratory coefficient of variation decreased for both DNA strand breaks (from 68 to 26%) and FPG sensitive sites (from 57 to 12%) by standardisation of the primary comet assay endpoint with calibration curve samples. The level of DNA strand breaks in the samples from two of the centres (0.56-0.61 lesions/10(6) bp) was significantly higher compared with the other three centres (0.41-0.45 lesions/10(6) bp). In contrast, there was no difference between the levels of FPG sensitive sites in PBMC samples from healthy donors in the different centres (0.41-0.52 lesion/10(6) bp).

Quantitative correlation of aflatoxin biomarker with dietary intake of aflatoxin in Tanzanian children.

The association between aflatoxin intake from maize-based weaning food and aflatoxin albumin adducts (AF-alb) was investigated in 148 Tanzanian children aged between 12 and 22 months, at 2 visits 6 months apart. At the first visit (storage season) there was a significant correlation at the individual level between AF-alb (geometric mean 43.2 pg/mg albumin) and aflatoxin intake (geometric mean 81.7 ng/kg b.w./d) through maize-based weaning food (r = 0.51, p < 0.01). Overall, this correlation was r = 0.43 (p < 0.01). The AF-alb level in weaning-age children in Tanzania closely reflects aflatoxin intake from maize in weaning food. Exposure levels suggest children may be at risk from aflatoxin associated health effects.

Aquaporins alteration revealed kidney damages in cerebral ischemia/reperfusion rats.

Background: Restoration of blood supply is a desired goal for the treatment of acute ischemic stroke. However, the restoration often leads to cerebral ischemia-reperfusion injury (CIR/I), which greatly increases the risk of non-neural organ damage. In particular, the acute kidney injury might be one of the most common complications. Aims: The study aimed to understand the damage occurred and the potential molecular mechanisms. Methods: The study was explored on the CIR/I rats generated by performing middle cerebral artery occlusion/reperfusion (MCAO/Reperfusion). The rats were evaluated with injury on the brains, followed by the non-neural organs including kidneys, livers, colons and stomachs. They were examined further with histopathological changes, and gene expression alterations by using RT-qPCR of ten aquaporins (Aqps) subtypes including Aqp1～Aqp9 and Aqp11. Furthermore, the Aqps expression profiles were constructed for each organ and analyzed by performing Principle Component Analysis. In addition, immunohistochemistry was explored to look at the protein expression of Aqp1, Aqp2, Aqp3 and Aqp4 in the rat kidneys. Results: There was a prominent down-regulation profile in the MCAO/Reperfusion rat kidneys. The protein expression of Aqp1, Aqp2, Aqp3 and Aqp4 was decreased in the kidneys of the MCAO/Reperfusion rats. We suggested that the kidney was in the highest risk to be damaged following the CIR/I. Down-regulation of Aqp2, Aqp3 and Aqp4 was involved in the acute kidney injury induced by the CIR/I.

An ECVAG inter-laboratory validation study of the comet assay: inter-laboratory and intra-laboratory variations of DNA strand breaks and FPG-sensitive sites in human mononuclear cells.

The alkaline comet assay is an established, sensitive method extensively used in biomonitoring studies. This method can be modified to measure a range of different types of DNA damage. However, considerable differences in the protocols used by different research groups affect the inter-laboratory comparisons of results. The aim of this study was to assess the inter-laboratory, intra-laboratory, sample and residual (unexplained) variations in DNA strand breaks and formamidopyrimidine DNA glycosylase (FPG)-sensitive sites measured by the comet assay by using a balanced Latin square design. Fourteen participating laboratories used their own comet assay protocols to measure the level of DNA strand breaks and FPG-sensitive sites in coded samples containing peripheral blood mononuclear cells (PBMC) and the level of DNA strand breaks in coded calibration curve samples (cells exposed to different doses of ionising radiation) on three different days of analysis. Eleven laboratories found dose-response relationships in the coded calibration curve samples on two or three days of analysis, whereas three laboratories had technical problems in their assay. In the coded calibration curve samples, the dose of ionising radiation, inter-laboratory variation, intra-laboratory variation and residual variation contributed to 60.9, 19.4, 0.1 and 19.5%, respectively, of the total variation. In the coded PBMC samples, the inter-laboratory variation explained the largest fraction of the overall variation of DNA strand breaks (79.2%) and the residual variation (19.9%) was much larger than the intra-laboratory (0.3%) and inter-subject (0.5%) variation. The same partitioning of the overall variation of FPG-sensitive sites in the PBMC samples indicated that the inter-laboratory variation was the strongest contributor (56.7%), whereas the residual (42.9%), intra-laboratory (0.2%) and inter-subject (0.3%) variations again contributed less to the overall variation. The results suggest that the variation in DNA damage, measured by comet assay, in PBMC from healthy subjects is assay variation rather than variation between subjects.

Assessment of the Adverse Health Effects of Aflatoxin Exposure from Unpackaged Peanut Oil in Guangdong, China.

Aflatoxins are liver carcinogens and are common contaminants in unpackaged peanut (UPP) oil. However, the health risks associated with consuming aflatoxins in UPP oil remain unclear. In this study, aflatoxin contamination in 143 UPP oil samples from Guangdong Province were assessed via liquid chromatography-tandem mass spectrometry (LC-MS). We also recruited 168 human subjects, who consumed this oil, to measure their liver functions and lipid metabolism status. Aflatoxin B1 (AFB1) was detected in 79.72% of the UPP oil samples, with levels ranging from 0.02 to 174.13 µg/kg. The average daily human intake of AFB1 from UPP oil was 3.14 ng/kg·bw/day; therefore, the incidence of liver cancer, caused by intake of 1 ng/kg·bw/day AFB1, was estimated to be 5.32 cases out of every 100,000 persons per year. Meanwhile, Hepatitis B virus (HBV) infection and AFB1 exposure exerted a synergistic effect to cause liver dysfunction. In addition, the triglycerides (TG) abnormal rate was statistically significant when using AFB1 to estimate daily intake (EDI) quartile spacing grouping (p = 0.011). In conclusion, high aflatoxin exposure may exacerbate the harmful effects of HBV infection on liver function. Contamination of UPP oil with aflatoxins in Guangdong urgently requires more attention, and public health management of the consumer population is urgently required.

Inter-laboratory variation in DNA damage using a standard comet assay protocol.

There are substantial inter-laboratory variations in the levels of DNA damage measured by the comet assay. The aim of this study was to investigate whether adherence to a standard comet assay protocol would reduce inter-laboratory variation in reported values of DNA damage. Fourteen laboratories determined the baseline level of DNA strand breaks (SBs)/alkaline labile sites and formamidopyrimidine DNA glycosylase (FPG)-sensitive sites in coded samples of mononuclear blood cells (MNBCs) from healthy volunteers. There were technical problems in seven laboratories in adopting the standard protocol, which were not related to the level of experience. Therefore, the inter-laboratory variation in DNA damage was only analysed using the results from laboratories that had obtained complete data with the standard comet assay protocol. This analysis showed that the differences between reported levels of DNA SBs/alkaline labile sites in MNBCs were not reduced by applying the standard assay protocol as compared with the laboratory's own protocol. There was large inter-laboratory variation in FPG-sensitive sites by the laboratory-specific protocol and the variation was reduced when the samples were analysed by the standard protocol. The SBs and FPG-sensitive sites were measured in the same experiment, indicating that the large spread in the latter lesions was the main reason for the reduced inter-laboratory variation. However, it remains worrying that half of the participating laboratories obtained poor results using the standard procedure. This study indicates that future comet assay validation trials should take steps to evaluate the implementation of standard procedures in participating laboratories.