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The influence of ferroptosis on the in vitro OGD/R model in rat microglia
Tao Ye1, Ning Zhang2, Anbang Zhang3
1Department of Rehabilitation, The First Affiliated Hospital of Guizhou University of Traditional Chinese Medicine, Guiyang, Guizhou, China.
Objective:
We aimed to explore the influence of ferroptosis on an oxygen-glucose deprivation/reoxygenation (OGD/R) model in primary rat microglia.
Methods:
Primary microglia were extracted from rats and cultured in vitro. The cells were subjected to a hypoxic environment for 6 h in a glucose-free medium, and then re-oxygenated for 24 h in DMEM/F12. Rat microglia were pretreated with the ferroptosis activator erastin and the ferroptosis inhibitor ferrostatin 1 for 24 h, followed by detection of cell cycle progression and apoptosis by flow cytometry. Intracellular total iron levels were measured. In addition, the relative levels of reactive oxygen species (ROS), malondialdehyde (MDA), and superoxide dismutase (SOD) were determined using enzyme-linked immunosorbent assay. The protein levels of 15-lox2, GPX4, SLC7A11, ACSL4, and TFR1 were examined by western blotting.
Results:
Compared with rat microglia subjected to OGD/R, pretreatment with erastin did not influence cell apoptosis but significantly enhanced total iron levels, MDA, and ROS levels, whereas it reduced SOD levels. Moreover, it upregulated ACSL4, TFR1, and 15-lox2 and downregulated GPX4 and SLC7A11. Pretreatment with ferrostatin 1 significantly inhibited cell apoptosis and cell cycle arrest in the G0/G1 phase. It significantly reduced total iron levels, MDA, and ROS levels and enhanced SOD levels, which also downregulated ACSL4, TFR1, and 15-lox2, and upregulated GPX4 and SLC7A11.
Conclusion:
Our study showed that inhibition of ferroptosis is favorable against potential OGD/R-induced damage in rat microglia.
Insights
Inhibition of ferroptosis protects primary rat microglia from oxygen-glucose deprivation/reoxygenation (OGD/R) injury. Blocking ferroptosis reduced cell damage and apoptosis, indicating a protective role.
Area of Science:
- Neuroscience
- Cell Biology
- Biochemistry
Background:
- Microglia play crucial roles in neuroinflammation and brain injury.
- Oxygen-glucose deprivation/reoxygenation (OGD/R) is a common model to simulate ischemic stroke conditions.
- Ferroptosis, a regulated form of cell death, has emerged as a significant factor in various neuropathologies.
Purpose of the Study:
- To investigate the role of ferroptosis in primary rat microglia subjected to OGD/R.
- To determine the effects of ferroptosis modulation on OGD/R-induced cellular damage.
Main Methods:
- Primary rat microglia were cultured and exposed to OGD/R conditions.
- Cells were pretreated with a ferroptosis activator (erastin) or inhibitor (ferrostatin 1).
- Cell apoptosis, cell cycle, iron levels, reactive oxygen species (ROS), malondialdehyde (MDA), superoxide dismutase (SOD), and key ferroptosis-related proteins were analyzed.
Main Results:
- Erastin (activator) increased iron, ROS, MDA, and upregulated ACSL4, TFR1, 15-lox2, while downregulating GPX4, SLC7A11, without affecting apoptosis.
- Ferrostatin 1 (inhibitor) reduced apoptosis, cell cycle arrest, iron, ROS, MDA, and enhanced SOD.
- Ferrostatin 1 also downregulated ACSL4, TFR1, 15-lox2, and upregulated GPX4, SLC7A11.
Conclusions:
- Ferroptosis plays a significant role in OGD/R-induced damage to primary rat microglia.
- Inhibiting ferroptosis demonstrates a protective effect against OGD/R-induced injury in microglia.
- Targeting ferroptosis may represent a therapeutic strategy for ischemic stroke and related neurological conditions.
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