Plasma-Treated Medium Strikes Ovarian Cancer Cells Through DNA Damage and Mitochondrial Collapse

Ovarian cancer remains one of the most lethal gynecological malignancies in the world, claiming more than 200,000 lives each year out of roughly 320,000 new diagnoses recorded in 2022. The disease is notorious for its late presentation, its tendency to spread across the peritoneal cavity, and its stubborn ability to return after initially successful chemotherapy. Now, a team of researchers at the University of Tübingen and the NMI Natural and Medical Sciences Institute in Germany has reported that a liquid activated by cold physical plasma — known as plasma treated medium, or PTM — can kill ovarian cancer cells in the laboratory through a striking combination of DNA damage, cell cycle disruption, and catastrophic mitochondrial dysfunction. The study, published in Cancer Cell International, adds fresh momentum to the emerging field of plasma medicine, in which ionized gas is harnessed as a weapon against tumors.

The physics behind the approach is as unusual as it is promising. Cold atmospheric plasma is a partially ionized gas generated at room temperature, typically from a carrier gas such as argon. Unlike the hot plasmas found in stars or industrial cutters, medical plasmas remain cool enough to touch living tissue, yet they deliver a potent cocktail of reactive oxygen and nitrogen species (RONS), charged particles, electromagnetic fields, and ultraviolet radiation. Cancer cells, which already live on the edge of oxidative stress because of their altered redox balance and elevated baseline levels of reactive oxygen species, are thought to be particularly vulnerable when an extra oxidative push tips them over their antioxidant threshold. Healthy cells, with more robust buffering capacity, may tolerate the same dose — a therapeutic window that plasma researchers have been chasing for more than a decade.

What makes the Tübingen study distinctive is its indirect delivery strategy. Rather than directing the plasma jet at cells themselves, the researchers used a clinical-grade electrosurgical setup — the VIO 3 generator with an APC 3 argon plasma module from Erbe Elektromedizin — to irradiate cell culture medium for two minutes from a fixed distance of seven millimeters. This treatment loads the liquid with long-lived reactive species, chiefly hydrogen peroxide, nitrite, and nitrate, which persist after the plasma is switched off. The resulting plasma treated medium can be stored, dosed in standardized dilutions, and, crucially, applied to parts of the body that a plasma jet could never reach directly — a decisive advantage for a cancer that seeds microscopic deposits across the abdominal lining.

To test the concept, the team worked with three established ovarian cancer cell lines chosen to represent different therapy response profiles: OVCAR-3, derived from a cisplatin-resistant high-grade adenocarcinoma; OVCAR-29, from an ovarian adenocarcinoma; and HEY, from a papillary cystadenocarcinoma with reported taxane resistance. After 18 hours of exposure to serial dilutions of PTM, cell viability measured by a resazurin assay dropped in a dose-dependent manner across all three lines. The potency varied considerably: HEY and OVCAR-3 cells were highly susceptible, with half-maximal inhibitory concentrations at dilutions of 1:3.6 and 1:4 respectively, while OVCAR-29 cells required a far more concentrated 1:1.1 dilution to reach the same effect.

Perhaps the most eye-catching finding concerned the cisplatin-resistant line. OVCAR-3 cells, which shrug off one of the standard chemotherapy drugs used against ovarian cancer, proved among the most sensitive to the plasma-derived reactive species. The authors suggest that resistance to conventional chemotherapy does not necessarily confer resistance to PTM, implying that plasma treatment exploits vulnerabilities distinct from those targeted by platinum and taxane drugs. If this holds up in further studies, plasma-based approaches could offer a route to attacking tumors that have already outmaneuvered the pharmacological arsenal — one of the most pressing unmet needs in gynecological oncology.

Dissecting the mechanism of cell death revealed a multi-pronged assault. In HEY and OVCAR-29 cells, PTM triggered phosphorylation of the histone variant H2AX, a well-established molecular flag for DNA double-strand breaks, alongside significant activation of caspases 3 and 7, the executioner enzymes of apoptosis. Cell cycle analysis by flow cytometry showed shifts consistent with checkpoint activation in response to oxidative DNA damage, with cells accumulating in the G1 phase and a corresponding decline in the G2 fraction. OVCAR-3 cells told a more nuanced story: caspase activity showed only a rising trend and H2AX phosphorylation was not detectable under the applied conditions, hinting that DNA-damage-independent routes — or simply insufficient dosing within the observation window — may contribute to cytotoxicity in this line.

The metabolic arm of the study employed one of the most elegant tools in modern cell biology: fluorescence lifetime imaging microscopy, or FLIM. Because the respiratory chain coenzymes NADH and FAD are naturally autofluorescent, the duration of their fluorescence after excitation reveals whether they are free in the cytosol or bound to mitochondrial proteins, and how the electron transport chain is coping. After just one hour of PTM exposure, the proportion of free NADH fell significantly in HEY and OVCAR-29 cells, while the fluorescence lifetimes of both free and protein-bound NADH and FAD shifted in patterns consistent with an imbalance between electron carrier formation and oxidation — a signature of impaired oxidative phosphorylation.

The mitochondrial damage extended to structure and electrical function. Staining with tetramethylrhodamine (TMRM), a dye that accumulates in mitochondria in proportion to membrane potential, showed significantly prolonged fluorescence lifetimes in HEY and OVCAR-29 cells after treatment, indicating depolarization of the mitochondrial membrane. Meanwhile, the physiological filamentous, thread-like mitochondrial networks of untreated cells collapsed into punctate, partly vesicular fragments — a morphological hallmark of mitochondrial injury frequently seen on the road to apoptosis. Together, the FLIM data and the TMRM measurements sketch a coherent picture: plasma-derived reactive species strike the bioenergetic core of the cancer cell, crippling its power supply and dismantling the organelles that decide its fate.

Crucially, the team did not stop at immortalized cell lines. They isolated primary tumor cells from the ascites — the fluid accumulating in the abdomen — of eight consenting ovarian cancer patients, using repeated partial trypsinization and immunofluorescence staining for the ovarian carcinoma marker PAX8 and the mesothelial marker calretinin to confirm that roughly 90 percent of the recovered cells were indeed malignant. When these patient-derived high-grade serous carcinoma cultures were exposed to PTM, viability fell by about 40 percent at a 1:9 dilution, with an IC50 at 1:5. Yet the response pattern differed markedly from the cell lines: caspase activation and H2AX phosphorylation were minimal, and instead the cells shifted from the G1 phase into S phase, with the S-phase fraction rising from 5 to 12 percent. The mitochondria of these primary cells also resisted depolarization and fragmentation.

This attenuated response, the authors argue, reflects the adaptive resilience of tumor cells conditioned to survive in the hostile, oxidative environment of the peritoneal cavity — cells equipped with robust DNA repair capacity, metabolic plasticity, and possibly stem-like survival programs. It is a sobering reminder that laboratory cell lines can overstate therapeutic efficacy, and it underscores why the inclusion of patient-derived material is becoming the gold standard in preclinical cancer research. The study has clear limitations: all experiments were conducted in vitro, no comparison with standard chemotherapeutics was performed, non-malignant ovarian cells were not tested, and mechanistic validation by Western blotting or ROS-scavenging interventions remains to be done. Even so, the clinical logic is compelling. Because complete cytoreductive surgery is the single strongest prognostic factor in ovarian cancer — yet occult microscopic deposits persist even after macroscopically perfect resections — a storable, doseable liquid that kills residual tumor cells could one day complement surgery in both complete and incomplete cytoreduction settings. The next steps, the researchers say, are validation in animal models and combination studies with standard chemotherapy regimens, as the field of plasma medicine edges closer to the operating theater.

Subject of Research: Antitumor effects of cold atmospheric plasma treated medium in human ovarian cancer cells

Article Title: Cold atmospheric plasma treated medium (PTM) induce antitumor effects in human ovarian cancer cells

Article References: Keßler, F., Oelkrug, F., Oberlechner, E., Andress, J., Brucker, S. Y., Koch, A., Berrio, D. A. C., Schenke-Layland, K., & Weiss, M. (2026). Cold atmospheric plasma treated medium (PTM) induce antitumor effects in human ovarian cancer cells. Cancer Cell International, 26(1), Article 330. https://doi.org/10.1186/s12935-026-04461-6

Image Credits: AI Generated

DOI: 10.1186/s12935-026-04461-6

Keywords: ovarian cancer, cold atmospheric plasma, plasma treated medium, reactive oxygen and nitrogen species, DNA damage, mitochondrial dysfunction, apoptosis, fluorescence lifetime imaging microscopy, ascites-derived tumor cells, plasma medicine, cancer metabolism, cell cycle