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Manganese exposure: a study on apoptosis and Ferroptosis in mouse Leydig and Sertoli cells
Jiaqi He1, Tongci Li1, Yue Su1
1Department of Histology and Embryology, Zunyi Medical University, NO. 6, xuefu west road, xinpu new district, Zunyi 563000, China.
Abstract:
Manganese (Mn), a vital trace element for biological functions, has raised health concerns due to potential toxicity. Excessive Mn impairs male reproduction by reducing testosterone, inducing oxidative stress, and disrupting spermatogenesis. However, its mechanisms targeting Leydig and Sertoli cells remain unclear. This study investigates Mn's reproductive toxicity by utilizing Leydig cell line TM3 and Sertoli cell line TM4, MTT assays revealed median lethal concentrations of 230 μM (TM3) and 170 μM (TM4), with AO/EB/DAPI staining confirming condensed nuclei and enhanced fluorescence. Apoptosis inhibitor Z-VAD-FMK (20 μM) suppressed cell death in both cell lines, whereas ferroptosis inhibitor Ferrostatin-1 (10 μM) specifically attenuated TM4 cell death. Necrosis inhibitor Necrostatin-1 (10 μM) showed no protective effect. Mn triggered ROS elevation in TM4 cells, accompanied by upregulated Caspase 3, Casp8ap2, GPX4, Gtf3c1, Mtfr1, HMOX1, and SLC7A2, while downregulating SLC7A15. These findings reveal Mn activates apoptosis in TM3 cells and concurrent apoptosis/ferroptosis in TM4 cells through ROS-dependent dysregulation of apoptosis- and ferroptosis-related genes. These findings establish distinct toxic mechanisms in TM4 cells and highlight the SLC7A15/HMOX1 axis as a therapeutic target to mitigate Mn-induced spermatogenic damage.
Insights
Manganese toxicity harms male reproduction by damaging Leydig and Sertoli cells. It induces apoptosis in Leydig cells and both apoptosis and ferroptosis in Sertoli cells, highlighting the SLC7A15/HMOX1 axis for potential therapies.
Area of Science:
- Reproductive Toxicology
- Cellular Toxicology
- Environmental Health
Background:
- Manganese (Mn) is essential but toxic at high levels, impacting male reproduction.
- Mechanisms of Mn toxicity in Leydig and Sertoli cells are not fully understood.
- Reproductive dysfunction linked to Mn includes reduced testosterone and impaired spermatogenesis.
Purpose of the Study:
- Investigate Mn's specific toxicity mechanisms in TM3 (Leydig) and TM4 (Sertoli) cell lines.
- Determine the roles of apoptosis and ferroptosis in Mn-induced reproductive cell death.
- Identify potential therapeutic targets to counteract Mn reproductive toxicity.
Main Methods:
- Utilized TM3 and TM4 cell lines for toxicity assays.
- Determined median lethal concentrations via MTT assays.
- Confirmed cell death pathways using AO/EB/DAPI staining and specific inhibitors (Z-VAD-FMK, Ferrostatin-1, Necrostatin-1).
- Analyzed reactive oxygen species (ROS) levels and gene expression related to apoptosis and ferroptosis.
Main Results:
- Mn exhibited differential toxicity: LC50 of 230 μM (TM3) and 170 μM (TM4).
- Apoptosis was confirmed in both cell types; ferroptosis was specific to TM4 cells.
- Mn increased ROS in TM4 cells, altering expression of key apoptosis/ferroptosis genes (e.g., upregulating *HMOX1*, *GPX4*; downregulating *SLC7A15*).
- Inhibitors confirmed apoptosis in TM3 and both apoptosis/ferroptosis in TM4 cells, while Necrostatin-1 had no effect.
Conclusions:
- Mn induces distinct toxic pathways in Leydig (apoptosis) and Sertoli (apoptosis and ferroptosis) cells.
- ROS-dependent gene dysregulation underlies Mn's reproductive toxicity.
- The *SLC7A15/HMOX1* axis presents a promising therapeutic target for mitigating Mn-induced damage to spermatogenesis.
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