Location: Toxicology & Mycotoxin Research2012 Annual Report
1a. Objectives (from AD-416):
1. Determine the biochemical and molecular basis for the species specificity of fumonisins using animal models. 2. Determine the dietary “no observed effect” and “lowest observed effect levels” for neural tube defect induction and determine the dose-response thresholds for elevation in sphingolipid biomarkers in blood spots and fumonisins in urine using animal models. 3. Determine the relationship between fumonisin consumption, urinary fumonisin (exposure biomarker) and changes in sphingolipids in blood spots (effect biomarker) in human populations consuming corn. 4. Determine the specific mechanism(s) by which fumonisins are readily taken up by corn plant roots and yet have limited translocation into above ground vegetative tissues. 5. Determine the effectiveness of alkaline cooking for reducing the toxic potential of fumonisin-contaminated whole kernel corn.
1b. Approach (from AD-416):
1. Conduct dose-response studies to determine the minimum oral dose of FB1 that disrupts sphingolipid metabolism and induces toxicity (increased apoptosis) in rat kidney and mouse liver. 2. Conduct dose-response studies in susceptible mouse strains to determine the thresholds for changes in biomarkers of exposure and effect and induction of neural tube defects. 3. Conduct epidemiological studies to identify humans consuming large amounts of corn-based foods in communities where FB is infrequently detected and frequently detected and sample and analyze urine (FBs) and blood spots (sphingolipids). 4. Conduct dose response studies to determine FB1 affects on plant transpiration and levels of sphingoid bases and their 1-phosphates in roots and aerial tissues in FB1-sensitive and -insensitive genotypes of corn. 5. Utilize FB-contaminated whole kernel corn to determine the processing conditions that maximize FB1 reduction using chemical analysis and in vivo animal bioassays.
3. Progress Report:
Objective 1: Rat liver and kidney were analyzed and confirmed that deoxysphinganine (DSa) is not detected in rat tissue. Analysis of mouse liver and kidney confirmed that DSa is produced in large amounts in animals treated with fumonisin B1 (FB1). These results provide evidence for the species and target organ specific responses in rats and mice with regards to the consequences of ceramide synthase inhibition. Objective 2: A study was conducted in collaboration with Creighton University to determine the oral dose of FB1 for induction of maternal toxicity, elevation in urinary FB1, and elevated sphingoid bases and sphingoid base 1-phosphates in maternal kidney, liver, and blood. This study showed that elevation in sphingoid base 1-phosphates in blood spots was correlated with subtle evidence of liver toxicity. Doses of FB1 of less than 10 mg/kg body weight per day for 4 days, induced little or no evidence suggestive of disrupted sphingolipid metabolism or liver toxicity. Follow-up studies were initiated to confirm or not results showing that neural tube defect induction by FB1 was significantly reduced in mice fed folate deficient diets. Follow-up confirmation is needed due to the unexpected and, based on earlier understandings of interactions between folate and fumonisins, counterintuitive findings of an earlier study. Objective 3: Biomarkers for FB exposure are being used in Institutional Review Board (IRB) approved studies in humans in collaboration with Guatemalan scientists. Approximately 1,200 urine and blood spot samples and 90 maize samples have been collected and analyzed from three locations in rural Guatemala. The results show that FB1 in urine is closely correlated with the level of FB in the maize collected from each locality and the urinary FB1 is significantly correlated with evidence indicating elevated levels of sphingoid base 1-phosphates in blood spots. At the request of the Guatemalan Ministry of Health (MOH) a modified human subjects' protocol was submitted and approved. A request was made to the NIH National Institute of Child Health & Human Development to modify the study design so as to identify two new sampling locations to be used to validate the findings thus far. This request was approved. Objective 4: Studies were conducted to determine if water uptake and transpiration in corn plants are modulated during the plant-Fusarium interaction. Transpiration may be negatively impacted by accumulation of sphingoid base 1-phosphates. We developed a growth chamber assay to address transpiration responses in plants treated with either FB alone or with F. verticillioides. Treatment with abscisic acid (ABA) plus FB1 resulted in reduced accumulation of FB1 in seedling roots compared to seedlings treated only with FB1. FB1 alone and infection with F. verticillioides both had a moderate effect on transpiration. Objective 5: An in vivo bioassay was conducted and showed that nixtamalization (alkaline cooking followed by three fresh water rinses) of whole kernel corn, as practiced in households and industry, significantly reduces the amount of measurable FB1 and, consequently, the toxicity of FB in the corn.
1. Evidence for fumonisin-induced disruption of sphingolipid metabolism. Studies conducted in Guatemala, in collaboration with the Centro de Investigaciones en Nutricion y Salud in Guatemala (CIENSA), Creighton University, and Duke University show that fumonisin exposure, based on the levels of urinary fumonisin B1, is significantly correlated with the level of sphingoid base 1-phosphates in blood spots and the increase in the sphinganine 1-phosphate to sphingosine 1-phosphate ratio. This finding is consistent with the hypothesis that high levels of fumonisin exposure in humans can lead to disruption of sphingolipid metabolism through inhibition of ceramide synthase. This is significant because every animal disease known to be caused by fumonisin has been shown to be closely correlated with, and preceded by, evidence of disruption of sphingolipid metabolism. The findings also provide a research tool for assessing the threshold for disruption of sphingolipid metabolism in humans and for designing epidemiological studies to evaluate the potential of fumonisin exposure as a contributing factor to human disease.
2. Alkaline processing of maize unequivocally reduces fumonisin toxicity in maize-based foods. Detoxification of fumonisin-contaminated, whole kernel corn by nixtamalization was for the first time demonstrated using a well-characterized rat feeding bioassay. The study combined cooking and rinsing (three times) under conditions that are relevant to households and industry. Unlike earlier in vitro experiments, the in vivo approach unequivocally demonstrated the absence of any significant toxicity that could be attributed to unknown fumonisin by-products remaining in the cooked corn.
Voss, K.A., Riley, R.T., Jackson, L.S., Jabloski, J.E., Bianchini, A., Bullerman, L.B., Hanna, M.A., Ryu, D. 2011. Extrusion cooking with glucose supplementation of fumonsin-contaminated corn grits protects against nephrotoxicity and disrupted sphingolipid metabolism in rats. Molecular Nutrition and Food Research. 55:S312-S320. DOI: 10.1002/mnfr.201100067.