Transcriptome and metabolome analysis of liver and kidneys of rats chronically fed NK603 Roundup-tolerant genetically modified maize

Abstract

Background: A previous 2-year rat feeding trial assessing potential toxicity of NK603 Roundup-tolerant genetically modified maize revealed blood and urine biochemical changes indicative of liver and kidney pathology. In an effort to obtain deeper insight into these findings, molecular profiling of the liver and kidneys from the same animals was undertaken.

Results: Transcriptomics showed no segregation of NK603 maize and control feed groups with false discovery rates ranging from 43 to 83% at a cut-off p value of 1%. Changes in gene expression were not reflective of liver and kidney toxic effects. Metabolomics identified 692 and 673 metabolites in kidney and liver, respectively. None of the statistically significant disturbances detected (12-56 for different test groups) survived a false discovery rate analysis. Differences in these metabolites between individual animals within a group were greater than the effect of test diets, which prevents a definitive conclusion on either pathology or safety.

Conclusions: Even if the biological relevance of the statistical differences presented in this study is unclear, our results are made available for scrutiny by the scientific community and for comparison in future studies investigating potential toxicological properties of the NK603 corn.

Keywords: GMO; Glyphosate; Metabolome; Roundup; Toxicity; Transcriptome.

Fig. 1

Wide-scale transcriptome profiles in liver and kidneys of NK603-fed rats. Liver and kidneys from control rats and animals fed NK603 GM maize either with or without Roundup application during the cultivation cycle were subjected to a full microarray transcriptome analysis. a PCA analysis of transcript cluster expression profiles shows no distinct separation into groups of treated (orange and green) and control (red) rats in both kidney and liver samples. By comparison, rats administered with Roundup (blue) in drinking water from the same experiment and subjected to the same transcriptome analysis clearly separate from the control and NK603 maize-fed groups. Each sphere represents the result of a single animal. b Volcano plots of the liver and kidney transcriptome profiles. Transcript cluster expression derived from the transcriptome profile data of liver and kidneys of control and NK603 maize-fed groups, either with or without Roundup (R) application, was plotted as log 2 fold change against −log10 p values. Each dot represents a single transcript cluster

Fig. 2

Venn diagram showing numbers of genes commonly disturbed in liver and kidney. Data were selected at p < 0.01 and fold changes >1.1

Fig. 3

Toxicity ontology analysis of genes disturbed in liver and kidneys of NK603 fed rats. List of toxicity process networks as revealed by MetaCore analysis of transcriptome profiles of liver and kidney from female rats fed NK603 GM maize either with or without Roundup application during the cultivation cycle or receiving 0.1 ppb of Roundup (50 ng/L glyphosate) in drinking water (p < 0.01, fold changes >1.1). The p values are determined by hypergeometric calculation

Fig. 4

PCA analysis of metabolome profiles of liver and kidneys. Liver and kidneys from control rats and animals fed NK603 GM maize either with or without Roundup application during the cultivation cycle were subjected to a metabolome analysis. PCA analysis of metabolite levels show no distinct separation into groups of treated (orange and green) and control (red) rats in both kidney and liver samples. Each sphere represents the result of a single animal

Fig. 5

Score plots of the significantly altered metabolites in liver and kidneys of NK603-fed rats. Levels of each metabolite from the metabolomics of liver and kidneys from female rats fed NK603 GM maize either with (NK603+R) or without (NK603) Roundup application during the cultivation cycle were subjected to a statistical analysis by comparison to controls using a Mann–Whitney U test. All metabolites showing a statistically significant change are shown. *p < 0.05; **p < 0.01, ***p < 0.001

Fig. 6

Scatter plots of the major significantly altered metabolites in liver of rats fed NK603 GM maize. Levels of the 7 metabolites commonly deregulated in livers from rats fed NK603 GM maize either with (NK603+R) or without (NK603) Roundup application during the cultivation cycle are shown. *p < 0.05; **p < 0.01