Abstract
BackgroundDioxins and related compounds are suspected of causing neurological disruption in human and experimental animal offspring following perinatal exposure during development and growth. The molecular mechanism(s) of the actions in the brain, however, have not been fully investigated. A major participant in the process of the dioxin-toxicity is the dioxin receptor, namely the aryl hydrocarbon receptor (AhR). AhR regulates the transcription of diverse genes through binding to the xenobiotic-responsive element (XRE). Since the AhR has also been detected in various regions of the brain, the AhR may play a key role in the developmental neurotoxicity of dioxins. This study focused on the effect of AhR activation in the developing neuron.MethodsThe influence of the AhR on the developing neuron was assessed using the Neuro2a-AhR transfectant. The undifferentiated murine neuroblastoma Neuro2a cell line (ATCC) was stably transfected with AhR cDNA and the established cell line was named N2a-Rα. The activation of exogenous AhR in N2a-Rα cells was confirmed using RNAi, with si-AhR suppressing the expression of exogenous AhR. The neurological properties of N2a-Rα based on AhR activation were evaluated by immunohistochemical analysis of cytoskeletal molecules and by RT-PCR analysis of mRNA expression of neurotransmitter-production related molecules, such as tyrosine hydroxylase (TH).ResultsN2a-Rα cells exhibited constant activation of the exogenous AhR. CYP1A1, a typical XRE-regulated gene, mRNA was induced without the application of ligand to the culture medium. N2a-Rα cells exhibited two significant functional features. Morphologically, N2a-Rα cells bore spontaneous neurites exhibiting axon-like properties with the localization of NF-H. In addition, cdc42 expression was increased in comparison to the control cell line. The other is the catecholaminergic neuron-like property. N2a-Rα cells expressed tyrosine hydroxylase (TH) mRNA as a functional marker of catecholaminergic neurotransmitter production. Thus, exogenous AhR induced catecholaminergic differentiation in N2a-Rα cells.ConclusionThe excessive activation of AhR resulted in neural differentiation of Neuro2a cells. This result revealed that dioxins may affect the nervous system through the AhR-signaling pathway. Activated AhR may disrupt the strictly regulated brain formation with irregular differentiation occurring rather than cell death.
Highlights
Dioxins and related compounds are suspected of causing neurological disruption in human and experimental animal offspring following perinatal exposure during development and growth
We have shown that over-expression of aryl hydrocarbon receptor (AhR) caused neural differentiation of Neuro2a cells
Neruo2a cells transfected with AhR, named N2a-Rα, may mimic the neurons forming the neural network during the perinatal stages, expressing AhR in the presence of the ligand
Summary
Dioxins and related compounds are suspected of causing neurological disruption in human and experimental animal offspring following perinatal exposure during development and growth. AhR regulates the transcription of diverse genes through binding to the xenobiotic-responsive element (XRE). The AhR is activated by binding to ligands, including dioxins, translocates from the cytoplasm to the nucleus and binds to the consensus sequence known as XRE (xenobiotic responsive element) [7]. Several phase I and phase II xenobiotic metabolizing enzymes, including cytochrome P-450 (CYP) 1A1, CYP1B1 and glutathione Stransferase (GST) possess multiple XRE sequences in the promoter region, and are the target genes for AhR following binding to the AhR ligands [8,9,10,11]. In addition to the xenobiotic or detoxification processes, the ligands of AhR, such as TCDD, affect the expression of a number of genes involved in cell proliferation (TGF-β, IL-1β and PAI2), cell cycle regulation (p27 and jun-B) [5,12,13,14] and inflammation. The AhR pathway may participate in the developmental toxication of dioxins during ontogenesis
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