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Effects of Manganese and Iron, Alone or in Combination, on Apoptosis in BV2 Cells
Yuan-Yuan Fang1,2, Cui-Liu Gan1,2, Jian-Chao Peng1,2
1Department of Toxicology, School of Public Health, Guangxi Medical University, 22 Shuang-Yong Rd., Nanning, 530021, Guangxi, China.
Abstract:
The aim of study was to address the effects of manganese and iron, alone and in combination, on apoptosis of BV2 microglia cells, and to determine if combined exposure to these metals augments their individual toxicity. We used a murine microglial BV2 cell line. Cell cytotoxicity was analyzed by propidium iodide (PI) exclusion assay. Cell ROS production was analyzed by 2', 7'-dichlorofluorescin diacetate (DCFH-DA) probe staining. Pro-inflammatory cytokine production was monitored by ELISA. Cell apoptosis was analyzed by PE Annexin V/7-AAD staining. Mitochondrial membrane integrity was analyzed by flow cytometry. We used immunoblotting to analyze the effect of manganese, iron alone, or their combined exposure on the activation of caspase9, P53, Bax, and Bcl2 apoptosis signaling pathways. Caspase3 activity was determined using a Colorimetric. Manganese, iron, and their combined exposure for 24 h induced the activation of BV2 microglia cells and increased ROS production and the expression of the inflammatory cytokines, IL-1β and TNF-α. And we also found that the apoptosis rate increased, mitochondrial membrane potential decreased, apoptosis-related proteins caspase9, P53, Bax, and Bcl2 expression increased, and caspase3 activity increased. Furthermore, we found that combined manganese-iron cytotoxicity was lower than that induced by manganese exposure alone. Manganese, iron alone, or their combination exposure can induce apoptosis in glial cells. Iron can reduce the toxicity of manganese, and there is an antagonistic effect between manganese and iron.
Insights
Manganese and iron exposure induces apoptosis in BV2 microglia cells. Combined exposure showed lower cytotoxicity than manganese alone, indicating an antagonistic effect between these metals.
Area of Science:
- Neuroscience
- Toxicology
- Cell Biology
Background:
- Microglia play crucial roles in neuroinflammation and immune responses within the central nervous system.
- Exposure to heavy metals like manganese and iron can impact microglial function and contribute to neurodegenerative processes.
Purpose of the Study:
- To investigate the individual and combined effects of manganese and iron on apoptosis in BV2 microglia cells.
- To determine if combined exposure to manganese and iron enhances their toxicity compared to individual exposures.
Main Methods:
- Utilized a murine microglial BV2 cell line for experiments.
- Assessed cell viability, reactive oxygen species (ROS) production, pro-inflammatory cytokine release (IL-1β, TNF-α), and apoptosis using various assays including PI exclusion, DCFH-DA staining, ELISA, and Annexin V/7-AAD staining.
- Analyzed apoptosis signaling pathways (caspase9, P53, Bax, Bcl2) and caspase3 activity via immunoblotting and colorimetric assays, respectively.
Main Results:
- Manganese and iron exposure, individually and combined, activated BV2 microglia, increased ROS production, and elevated pro-inflammatory cytokine expression.
- Apoptosis rates increased, mitochondrial membrane potential decreased, and key apoptosis-related proteins (caspase9, P53, Bax, Bcl2) and caspase3 activity were upregulated.
- Combined manganese-iron exposure exhibited lower cytotoxicity than manganese exposure alone, suggesting an antagonistic interaction.
Conclusions:
- Both manganese and iron, alone or in combination, induce apoptosis in glial cells.
- Iron exhibits a protective or antagonistic effect against manganese-induced toxicity in BV2 microglia.
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