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Temporal Proteomic Profiling of SH-SY5Y Differentiation with Retinoic Acid Using FAIMS and Real-Time Searching.

ABSTRACT: The SH-SY5Y cell line is often used as a surrogate for neurons in cell-based studies. This cell line is frequently differentiated with all-trans retinoic acid (ATRA) over a 7-day period, which confers neuron-like properties to the cells. However, no analysis of proteome remodeling has followed the progress of this transition. Here, we quantitatively profiled over 9400 proteins across a 7-day treatment with retinoic acid using state-of-the-art mass spectrometry-based proteomics technologies, including FAIMS, real-time database searching, and TMTpro16 sample multiplexing. Gene ontology analysis revealed that categories with the highest increases in protein abundance were related to the plasma membrane/extracellular space. To showcase our data set, we surveyed the protein abundance profiles linked to neurofilament bundle assembly, neuron projections, and neuronal cell body formation. These proteins exhibited increases in abundance level, yet we observed multiple patterns among the queried proteins. The data presented represent a rich resource for investigating temporal protein abundance changes in SH-SY5Y cells differentiated with retinoic acid. Moreover, the sample preparation and data acquisition strategies used here can be readily applied to any analogous cell line differentiation analysis.

SUBMITTER: Zhang T

PROVIDER: S-EPMC8210949 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundMaleic acid is a multi-functional chemical widely used in the field of industrial chemistry for producing food additives and food contact materials. As maleic acid may contaminate food by the release from food packages or intentional addition, it raises the concern about the effects of excessive dietary exposure to maleic acid on human health. However, the influence of maleic acid on human health has not been thoroughly studied. In silico toxicogenomics approaches have found the association between maleic acid and nervous system disease in human. The aim of this study is to experimentally explore the effects of maleic acid on human neuronal cells.MethodsA microarray-based transcriptome profiling was performed to offer a better understanding of the effects of maleic acid on human health. Gene expression profiles of human neuroblastoma SH-SY5Y cells exposed to three concentrations of maleic acid (10, 50, and 100 μM) for 24 h were analyzed. Genes which were differentially expressed in dose-dependent manners were identified and further analyzed with an enrichment analysis. The expression profile of selected genes related to the inferred functional changes was validated using quantitative polymerase chain reaction (qPCR). Specific fluorescence probes were applied to observe the inferred functional changes in maleic acid-treated neuronal cells.ResultsA total of 316 differentially expressed genes (141 upregulated and 175 downregulated) were identified in response to the treatment of maleic acid. The enrichment analysis showed that DNA binding and metal ion binding were the significant molecular functions (MFs) of the neuronal cells affected by maleic acid. Maleic acid exposure decreased the expression of genes associated with calcium and thiol levels of the cells in a dose-dependent manner. The levels of intracellular calcium and thiol levels were also affected by maleic acid dose-dependent.DiscussionThe exposure to maleic acid is found to decrease the cellular calcium and thiol levels in human neuronal cells at both transcriptional and functional levels. This study reported the first transcriptomic profiling of human neuronal cells treated with maleic acid. It is also the first experimental validation of chemical effects predicted by in silico toxicogenomics approaches. The proposed approach may be useful in understanding the potential effects of other poorly characterized chemicals on human health.

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Temporal Proteomic Profiling of SH-SY5Y Differentiation with Retinoic Acid Using FAIMS and Real-Time Searching.

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