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Updated: Jun 30, 2025
Author Spotlight: Flow Cytometric Determination of Pyroptosis in Avian Cells
Published on: May 31, 2024
Decitabine combined with cold atmospheric plasma induces pyroptosis via the ROS/Caspase-3/GSDME signaling pathway in
Liang Du1, Huiyun Ming1, Zhuna Yan2
1College of Pharmacy, Anhui Medical University, Hefei 230032, China; Anhui Province Key Laboratory of Medical Physics and Technology, Institute of Health & Medical Technology, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031, China.
Background:
High methylation of the DFNA5 gene results in the absence of GSDME, a key protein that mediates pyroptosis, while decitabine demethylates the DFNA5 gene, resulting in high expression of the GSDME protein. Cold atmospheric plasma (CAP) is a novel anti-cancer method that induces tumor cell death.
Methods:
The pyroptosis induced by decitabine in combination with CAP in Ovcar5 cells was evaluated. In particular, mitochondrial membrane potential was estimated by JC-1 staining, dehydrogenase (LDH) release was assessed by ELISA, Annexin V/PI staining was detected by flow cytometry, the cell cycle changes were evaluated using PI staining followed by detection by flow cytometry, and Caspase-9 cleavage, Caspase-3 cleavage and GSDME expression were evaluated by western blot.
Results:
Decitabine resulted in high expression of the GSDME in Ovcar5 in a concentration-dependent manner and increased tumor cell sensitivity to CAP. CAP induced mitochondrial damage and activated the Caspase-9/Caspase-3 pathway. Therefore, decitabine combined with CAP induced Ovcar5 cell pyroptosis through Caspase-3 mediated GSDME cleavage. Reactive oxygen species (ROS) generated by CAP treatment played an important role in the CAP/decitabine combination-induced production of ROS, activation of Caspase-9/Caspase-3, GSDME cleavage and pyroptosis that ROS scavenger NAC inhibited all these processes.
Conclusions:
CAP combined with decitabine induced Caspase-3 activation, which cleaved decitabine-upregulated GSDME and ediated pyroptosis.
Insights
Decitabine and cold atmospheric plasma (CAP) together induce pyroptosis, a form of programmed cell death, in Ovcar5 cancer cells by activating the Caspase-3 pathway and cleaving GSDME. This combination therapy shows promise for cancer treatment.
Area of Science:
- Oncology
- Cell Biology
- Biochemistry
Background:
- High DFNA5 gene methylation silences GSDME, a protein crucial for pyroptosis.
- Decitabine reverses DFNA5 methylation, increasing GSDME expression.
- Cold atmospheric plasma (CAP) is an emerging anti-cancer modality inducing tumor cell death.
Purpose of the Study:
- To investigate the synergistic effect of decitabine and CAP on inducing pyroptosis in Ovcar5 cells.
- To elucidate the molecular mechanisms underlying decitabine and CAP combination therapy.
Main Methods:
- Ovcar5 cells were treated with decitabine and CAP.
- Pyroptosis was assessed via LDH release, Annexin V/PI staining, and cell cycle analysis.
- Mitochondrial membrane potential was measured using JC-1 staining.
- Caspase-9/Caspase-3 cleavage and GSDME expression were analyzed by Western blot.
Main Results:
- Decitabine upregulated GSDME expression in a dose-dependent manner, enhancing Ovcar5 cell sensitivity to CAP.
- CAP induced mitochondrial damage and activated the Caspase-9/Caspase-3 pathway.
- The combination therapy triggered pyroptosis through Caspase-3-mediated GSDME cleavage, with reactive oxygen species (ROS) playing a key role.
Conclusions:
- Decitabine and CAP synergistically induce pyroptosis in Ovcar5 cells.
- The combination activates Caspase-3, leading to GSDME cleavage and cell death.
- ROS generated by CAP are critical mediators of this combined anti-cancer effect.
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