DNA damage-inducible transcript 3-mediated endoplasmic reticulum stress drives manganese-induced apoptosis in bovine
Liwei Huang1, Yu Cheng1, Xiaolong Pan1
1College of Animal Science, Shanxi Agricultural University, Taigu, 030801, China.
Abstract:
Manganese (Mn) is an essential trace element, but it is also an environmental pollutant. Excessive Mn accumulation in animals induces toxic effects, particularly reproductive damage. The mechanism of Mn toxicity in ruminants is unclear. This study investigated the mechanism of Mn toxicity on bovine Leydig cells. Bovine Leydig cells were treated with Mn at a semi-inhibitory concentration of 70 μM for 24 h, and this concentration was subsequently set as the Mn-treated experimental group. Multiple analytical approaches were employed, including the examination of cellular ultrastructure, measurement of oxidative stress indicators, analysis of apoptosis-related genes, and RNA-Seq sequencing for the screening of differentially expressed genes. The results showed that Mn treatment led to abnormal damage to the ultrastructure of mitochondria and endoplasmic reticulum. An increase in reactive oxygen species (ROS) and malondialdehyde (MDA) levels, along with a decrease in glutathione peroxidase (GSH-Px) activity, provided evidence of oxidative stress induction. Additionally, Mn exposure upregulated the expression of apoptosis-related genes Caspase-3 and BAX, while downregulating the expression of BCL-2, indicating the initiation of apoptosis. RNA-Seq analysis revealed that the DDIT3 gene, associated with endoplasmic reticulum stress, exhibited highly significant differential expression. Further experiments showed that knockdown of the DDIT3 gene effectively alleviated Mn-induced apoptosis in bovine Leydig cells. In conclusion, Mn exposure promotes apoptosis in bovine Leydig cells, and interference with DDIT3 can mitigate this apoptotic process, which provides valuable references for the prevention and control of Mn pollution in animal husbandry and the management of the reproductive health of breeding bulls.
Insights
Manganese (Mn) exposure causes reproductive damage in bulls by inducing apoptosis in Leydig cells. Targeting DDIT3 gene expression can mitigate this manganese-induced cell death, offering insights for animal husbandry.
Area of Science:
- Veterinary Toxicology
- Reproductive Biology
- Cellular Biology
Background:
- Manganese (Mn) is essential but toxic at high levels, causing reproductive damage in animals.
- The precise mechanism of Mn toxicity in ruminant reproductive systems remains unclear.
- Bovine Leydig cells are crucial for testosterone production and male fertility.
Purpose of the Study:
- To elucidate the mechanism of manganese toxicity in bovine Leydig cells.
- To investigate the role of oxidative stress, apoptosis, and endoplasmic reticulum stress in Mn toxicity.
- To identify potential therapeutic targets for mitigating Mn-induced reproductive damage.
Main Methods:
- Bovine Leydig cells were exposed to 70μM manganese (Mn) for 24 hours.
- Evaluated cellular ultrastructure, oxidative stress markers (ROS, MDA, GSH-Px), and apoptosis-related genes (Caspase-3, BAX, BCL-2).
- Utilized RNA-Seq to identify differentially expressed genes, focusing on DDIT3, and performed gene knockdown experiments.
Main Results:
- Mn exposure damaged mitochondria and endoplasmic reticulum ultrastructure.
- Induced significant oxidative stress, evidenced by increased ROS/MDA and decreased GSH-Px.
- Upregulated pro-apoptotic genes (Caspase-3, BAX) and downregulated anti-apoptotic gene (BCL-2), confirming apoptosis initiation.
- Identified DDIT3 (endoplasmic reticulum stress marker) as significantly upregulated; its knockdown reduced Mn-induced apoptosis.
Conclusions:
- Manganese exposure induces apoptosis in bovine Leydig cells via oxidative stress and endoplasmic reticulum stress pathways, involving DDIT3.
- Interference with DDIT3 expression can alleviate Mn-induced apoptosis.
- Findings provide a basis for managing Mn pollution in animal husbandry and bull reproductive health.
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