ABSTRACT
The Neuro-2a cell assay has been a promising in vitro alternative for the detection of saxitoxin (STX)-like toxins. However, its application is problematic in samples with complex matrices containing different toxins, whose mechanisms of action could be antagonistic. In the search of alternative procedures that reduce or avoid this interference, we evaluated the transcriptional modulation produced by a 24-h exposure to STX in Neuro-2a cells under three conditions: exposure to STX (33 nM), a mussel meat matrix (12.5 mg meat/mL) and a fortified sample (STX-fortified matrix). Differential gene expression was evaluated by RNA-seq after Illumina high-throughput sequencing, and data were analyzed to identify genes differentially expressed regardless of the matrix. From the 9487 identified genes, 213 were differentially expressed of these, 10 genes were identified as candidate markers for STX detection due to their regulation by STX regardless of the matrix interference. Expression dynamics of 7 of these candidate genes (Fgf-1, Adgrb2, Tfpt, Zfr2, Nup 35, Fam195a, and Dusp7) was further evaluated by qRT-PCR analysis of cells exposed to different concentrations of STX for up to 24 h. A downregulation of some markers expression was observed, namely Nup35 (involved in nucleo-cytoplasmic transporter activity) and Zfr-2 (involved in nucleic acids binding), whereas Fgf-1 (apoptosis signaling) was significantly upregulated. Markers' expression was not influenced by the matrix, suggesting that gene expression variations are directly related to STX response. These results support the potential of these genes as biomarkers for the development of an alternative STX-like toxins screening method.
