Abstract
Many “hot spot” geographic areas across the world with drinking water co-contaminated with inorganic arsenic (iAs) and fluoride (F–), two of the most common natural contaminants in drinking water. Both iAs and F– are known neurotoxins and affect neurodevelopment of children. However, very few studies have investigated the neurodevelopmental effects of concurrent exposure to iAs and F–, which could potentially pose a greater risk than iAs or F– exposure alone. Further, perturbations of gut microbiota, which plays a regulatory role in neurodevelopment, resulting from iAs and F– exposure has been reported in numerous studies. There is lacking of information regarding to the relationship among concurrent iAs and F– exposure, microbiome disruption, and neurodevelopmental impacts. To fill these gaps, we treated offspring rats to iAs (50 mg/L NaAsO2) and F– (100 mg/L NaF), alone or combined from early life (in utero and childhood) to puberty. We applied Morris water maze test to assess spatial learning and memory of these rats and generated gut microbiome profiles using 16S rRNA gene sequencing. We showed that concurrent iAs and F– exposure caused more prominent neurodevelopmental effects in rats than either iAs or F– exposure alone. Moreover, Unsupervised Principal Coordinates Analysis (PCoA) and Linear Discriminant Analysis Effect Size (LEfSe) analysis of gut microbiome sequencing results separated concurrent exposure group from others, indicating a more sophisticated change of gut microbial communities occurred under the concurrent exposure condition. Further, a correlation analysis between indices of the water maze test and microbial composition at the genus level identified featured genera that were clearly associated with neurobehavioral performance of rats. 75% (9 out of 12) genera, which had a remarkable difference in relative abundance between the control and combined iAs and F– exposure groups, showed significantly strong correlations (r = 0.70–0.90) with the water maze performance indicators. Collectively, these results suggest that concurrent iAs and F– exposure led to more prominent effects on neurodevelopment and gut microbiome composition structures in rats, and the strong correlation between them indicates a high potential for the development of novel microbiome-based biomarkers of iAs and/or F– associated neurodevelopmental deficits.
*Abstract online at https://doi.org/10.1016/j.envint.2020.105763
Excerpt:
3.3. Effects of iAs and F-, alone or in combination, on pathological changesin the hippocampal CA1 region
H&E staining of neuron in the hippocampal CA1 region was observed at different optical microscope magnifications (Bar = 100 um 50 um in Fig. 2B & C respectively). In the control group, hippocampal CA1 neurons were tightly organized and had multiple cell layers, with healthy cell morphology and clearly stained nuclei and nucleoli. Compared with the control group, hippocampal CA1 neuron pathological changes were observed in iAs and/or F-exposure groups, including neurofibrillary degeneration, loosely and irregularly organized structure, fewer cell layers, cytoplasmic cavitation, deeper staining of cytoplasm and nuclear pyknosis. In the iAs exposure group the cytoplasm of the neuronal cells deepened markedly, and the neuronal fibers were swollen. In the F-exposure group, neuronal cells were loosely and disorderly organized, and some swollen neuronal fibers were observed. In the AsF combination group, the structure of neuronal cells was even looser and more irregular. The number of cell layers and cell quantities significantly decreased, along with increased swollen neuronal fibers and cells with cytoplasmic cavitation.
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5. Conclusion
In summary, our present study evaluated neurotoxicity and gutmicrobiome disturbance as a result of exposure to high-dose of iAs andF-, alone or in combination, from early life (in utero and childhood) to puberty in rats. iAs and/or F-exposure led to neurobehavioral deficits in spatial learning and memory, most prominently in offspring rats co-exposed to iAs and F-. The H&E staining results confirmed that iAs and/or F-caused pathological changes in hippocampal neuron. 16S rRNAgene sequencing results demonstrated perturbation of gut microbiomecommunities at different taxonomic levels in response to iAs and/or F-exposure. Again, the effects were more noticeable in the AsF combination group. Furthermore, nine genera, which had significant difference of relative abundance between the control and AsF combination groups, were identified to have significantly strong correlation (r= 0.70–0.90) with spatial learning and memory performance. Collectively, these results suggest that concurrent iAs and F-exposureled to more prominent effects on neurodevelopment and gut micro-biome composition structures in rats, and the strong correlation between them indicates a high potential for the development of novel microbiome-based biomarkers of iAs and/or F-associated neurodevelopmental deficits.
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