MicroRNAs in porcine uterus and serum are affected by zearalenone and represent a new target for mycotoxin biomarker discovery

Abstract

The mycotoxin zearalenone (ZEN) poses a risk to animal health because of its estrogenic effects. Diagnosis of ZEN-induced disorders remains challenging due to the lack of appropriate biomarkers. In this regard, circulating microRNAs (small non-coding RNAs) have remarkable potential, as they can serve as indicators for pathological processes in tissue. Thus, we combined untargeted and targeted transcriptomics approaches to investigate the effects of ZEN on the microRNA expression in porcine uterus, jejunum and serum, respectively. To this end, twenty-four piglets received uncontaminated feed (Control) or feed containing 0.17 mg/kg ZEN (ZEN low), 1.46 mg/kg ZEN (ZEN medium) and 4.58 mg/kg ZEN (ZEN high). After 28 days, the microRNA expression in the jejunum remained unaffected, while significant changes in the uterine microRNA profile were observed. Importantly, 14 microRNAs were commonly and dose-dependently affected in both the ZEN medium and ZEN high group, including microRNAs from the miR-503 cluster (i.e. ssc-miR-424-5p, ssc-miR-450a, ssc-miR-450b-5p, ssc-miR-450c-5p, ssc-miR-503 and ssc-miR-542-3p). Predicted target genes for those microRNAs are associated with regulation of gene expression and signal transduction (e.g. cell cycle). Although the effects in serum were less pronounced, receiver operating characteristic analysis revealed that several microRNA ratios were able to discriminate properly between non-exposed and ZEN-exposed pigs (e.g. ssc-miR-135a-5p/ssc-miR-432-5p, ssc-miR-542-3p/ssc-miR-493-3p). This work sheds new light on the molecular mechanisms of ZEN, and fosters biomarker discovery.

Biotransformation of the Mycotoxin Zearalenone to its Metabolites Hydrolyzed Zearalenone (HZEN) and Decarboxylated Hydrolyzed Zearalenone (DHZEN) Diminishes its Estrogenicity In Vitro and In Vivo

“…Yet, the opposite effects seen between the piglets fed ZEN and DHZEN deserve further investigations. As briefly discussed in Grenier et al [11], the lack of effects observed on the ERα is in agreement with Oliver et al [54], who analyzed the gene and protein expression of ERs in the uterus of piglets exposed to 1.5 mg/kg of ZEN for four weeks. However, Oliver et al reported a two-fold increase in the mRNA level of ERβ in those animals, which could not be confirmed in the present experiment.…”

“…None of the treatments significantly affected the body weight of animals (Table 1). While ZEN caused a time-dependent increase of the vulva size, reaching an enlargement by a factor of 3.3 compared to the control group on day 27 [11], neither HZEN nor DHZEN influenced the vulva size during the experimental period. Similarly, the reproductive tract weight at the end of the experiment was markedly increased in the ZEN group, whereas the ZEN metabolites showed no effect on this parameter (Table 1).…”

“…Finally, we evaluated the impact of ZEN and its metabolites on microRNA expression in uterus. To this end, a subset of 15 microRNAs was selected for qPCR analysis based on previous data generated by Grenier et al [11] (see Section 5.2.3 for details). Figure 4 displays the results of the hierarchical clustering analysis.…”

“…Alteration of hematological and biochemical measures, gene expression and subpopulations of immunocompetent cells in tissues have been observed after ZEN exposure in piglets and prepubertal gilts, albeit with limited reproducibility between studies [7]. Recently, the effect of ZEN on the expression of microRNAs has caught scientific interest [8,9,10,11]. MicroRNAs represent a class of small non-coding RNAs, that negatively regulate gene expression.…”

“…They can serve as indicators for pathological processes in organs, and possess considerable potential as biomarkers [12]. Very recently it has been shown that ZEN affects uterine expression of certain microRNAs in pigs, among them ssc-miR-424-5p, ssc-miR-450a, ssc-miR-450b-5p, ssc-miR-450c-5p, ssc-miR-503, and ssc-miR-542-3p [11]. This effect was partly attributed to estrogen receptor (ER) activation, but further molecular targets of this mycotoxin, such as G protein-coupled estrogen receptor 1 [13] or the pregnane X receptor [14], need to be considered in this context.…”