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Down-regulation of S100A9 and S100A10 in manganese-resistant RBL-2H3 cells
Hitomi Fujishiro1, Toshinao Ohashi, Miki Takuma
1Laboratory of Molecular Nutrition and Toxicology, Faculty of Pharmaceutical Sciences, Tokushima Bunri University.
Abstract:
Exposure to excess amounts of manganese causes toxic effects, including neurological symptoms such as Parkinsonism. However, endogenous factors involved in the protection against manganese toxicity remain unclear. Previously, we showed that rat basophilic leukemia RBL-2H3 cells are highly sensitive to MnCl₂ compared with other rat cell lines. To identify the genes involved in resistance to manganese toxicity, two lines of Mn-resistant cells showing resistance to 300 µM MnCl₂ (RBL-Mnr300) and 1200 µM MnCl₂ (RBL-Mnr1200) were developed from RBL-2H3 cells by a stepwise increase in MnCl₂ concentration in the medium. Microarray analyses were carried out to compare gene expression between parental RBL-2H3 cells and RBL-Mnr300 or RBL-Mnr1200 cells. Five genes exhibited more than 10-fold up-regulation in both RBL-Mnr300 and RBL-Mnr1200 cells, and 24 genes exhibited less than 0.1-fold down-regulation in both Mn-resistant cell lines. The S100a9 and S100a10 genes, encoding the calcium-binding S100A9 and S100A10 proteins, respectively, were found among the three most down-regulated genes in both Mn-resistant cell lines. The marked decreases in mRNA levels of S100a9 and S100a10 were confirmed by real-time RT-PCR analyses. Further characterization and comparison of these Mn-resistant cells may enable the identification of novel genes that play important roles in the modification of manganese toxicity.
Insights
Researchers developed manganese-resistant cells to study protection against manganese toxicity. They identified down-regulated S100a9 and S100a10 genes, suggesting a role in manganese neurotoxicity.
Area of Science:
- Neurotoxicology
- Cell Biology
- Genetics
Background:
- Excess manganese exposure causes neurotoxicity, mimicking Parkinsonism.
- Endogenous protective factors against manganese toxicity are not well understood.
- Rat basophilic leukemia RBL-2H3 cells are particularly sensitive to manganese chloride (MnCl₂).
Purpose of the Study:
- To identify genes involved in cellular resistance to manganese toxicity.
- To compare gene expression profiles between manganese-sensitive and manganese-resistant cell lines.
Main Methods:
- Developed two manganese-resistant cell lines (RBL-Mnr300 and RBL-Mnr1200) from RBL-2H3 cells.
- Utilized microarray analysis to compare gene expression between parental and resistant cells.
- Confirmed gene expression changes using real-time RT-PCR.
Main Results:
- Identified five genes with >10-fold up-regulation and 24 genes with <0.1-fold down-regulation in both resistant cell lines.
- Found S100a9 and S100a10 genes, encoding calcium-binding proteins, among the most down-regulated genes.
- Confirmed significant decreases in S100a9 and S100a10 mRNA levels in resistant cells.
Conclusions:
- The study identified novel genes potentially involved in manganese toxicity resistance.
- Down-regulation of S100a9 and S100a10 may play a role in cellular defense against manganese.
- Further research on these Mn-resistant cells could reveal new therapeutic targets for manganese neurotoxicity.
