Targeting Mitochondrial ROS in Ferroptosis: The Roles of Drp1 and VDAC1 in Neuronal Cells
Ferroptosis is an iron-dependent necrotic cell death pathway that has emerged as a central mechanism in neurodegenerative diseases. The inhibition of ferroptosis has a strong potential as effective strategy to prevent cell death that is mediated by oxidative stress. One hallmark of ferroptosis is...
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| Format: | Doctoral Thesis |
| Language: | English |
| Published: |
Philipps-Universität Marburg
2025
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| Online Access: | PDF Full Text |
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| Summary: | Ferroptosis is an iron-dependent necrotic cell death pathway that has emerged as a central
mechanism in neurodegenerative diseases. The inhibition of ferroptosis has a strong potential
as effective strategy to prevent cell death that is mediated by oxidative stress. One hallmark
of ferroptosis is the lipid peroxidation mediated rupture of the cell membrane, in which ROS
play a vital role. Mitochondria are known as the main producer of ROS during oxidative
phosphorylation, a byproduct during energy metabolism, emphasizing a link between
mitochondrial dysfunction and ferroptosis. While mitochondrial ROS and lipid peroxidation are
central to ferroptosis, the specific role of Dynamin-related Protein 1 (Drp1) and the Voltage-
dependent Anion Channel (VDAC1) in mitochondrial pathways of ferroptosis remained
unclear.
The aim of the present study was to investigate how Drp1 and VDAC1 contribute to ferroptosis
by regulating mitochondrial integrity, ROS production and iron metabolism and to explore
whether inhibition or modulation of these two proteins can protect neuronal cells from
ferroptotic damage. Drp1 is the key protein to regulate mitochondrial fission. The study
proposed that inhibition of excessive Drp1-mediated mitochondrial fragmentation would
prevent mitochondrial ROS amplification, preservation of cellular bioenergetics and therefore
abrogate ferroptosis. Dysregulation of calcium and iron homeostasis is implicated as trigger
for mitochondrial dysfunction and therefore aberrant redox balance. VDAC1 transports ions
and metabolites bidirectionally in and out of the mitochondria for physiological mitochondrial
function. Inhibition of VDAC1 was anticipated to prevent mitochondrial ROS amplification
through the decrease of mitochondrial respiration and bioenergetic shift towards glycolysis to
abolish ferroptosis progression.
Mitochondrial dynamics and metabolic regulation are interconnected because mitochondrial
shape and function directly influence cellular energy metabolism and vice versa. Mitochondrial
morphology determines ATP generation, ROS production, and metabolite exchange, which
are critical for maintaining cellular energy demands. With this, the impact of both, Drp1 and
VDAC1, was investigated to elaborate on multiple possibilities to target mitochondrial ROS
during ferroptosis.
The model system used for the ferroptosis studies was the mouse hippocampal cell line HT22.
Drp1 was knocked out using a CRISPR/Cas9-based genome editing approach, while VDAC1
was inhibited pharmacologically using Akos-22. The methodological approach comprised cell
viability assessments and fluorescence-based assays to assess ROS, iron, lipid peroxidation,
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Summary
calcium, as well as mitochondrial integrity and function. The Seahorse XF-analyzer was used
to study cellular bioenergetics.
The findings of the study demonstrate that genetical deletion of Drp1 increased the resilience
against ferroptosis by preventing excessive mitochondrial fragmentation and stabilizing
cellular bioenergetics. Mitochondrial integrity and function which are normally lost through
ferroptosis were preserved. Pharmacological inhibition of VDAC1 abolished ferroptotic cell
death by a metabolic shift towards glycolysis to reduce the mitochondrial contribution to
detrimental ROS signaling. Inhibition of VDAC1 decreased mitochondrial respiration and the
generation of mitochondrial ROS while decreasing cytosolic iron and calcium levels.
Alleviation of ferroptosis-mediated iron dysbalance by iron chelation and cellular iron uptake
inhibition also decreased oxidative phosphorylation, leading to the abrogation of detrimental
mitochondrial ROS signaling.
The present study provides valuable insights into mitochondrial pathways of ferroptosis. By
preserving mitochondrial integrity and alleviating mitochondrial ROS generation, mitochondria
are highlighted as important targets for novel strategies to combat ferroptosis-related
diseases, such as neurodegenerative diseases and conditions involving ischemic injuries. In
therapy-resistant cancers, mitochondria-targeted applications that lead to mitochondrial
dysfunction to increase the susceptibility towards ferroptosis might prove useful. The study
fills a critical gap in understanding how mitochondrial-specific processes contribute to
ferroptosis and how they can be modulated for therapeutic benefit.
These findings open new avenues for targeting mitochondrial dynamics and metabolite
exchange to combat ferroptosis-associated pathologies. Future work will focus on translating
these insights into therapeutic strategies for neurodegenerative diseases and cancer. |
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| DOI: | 10.17192/z2025.0108 |