Real‐Time Monitoring of Tissue Oxygenation During Hyperbaric Oxygen Exposure Using In Vivo EPR Oximetry

ABSTRACT Purpose Hyperbaric oxygen (HBO) therapy increases blood oxygen levels by exposing patients to 100% oxygen under elevated pressure and is used clinically for conditions such as decompression sickness, carbon monoxide poisoning, chronic wounds, and impaired tissue healing. However, real‐time precise and quantitative measurement of tissue oxygen tension (pO 2 ) under hyperbaric conditions remains technically very challenging. Methods We developed a specialized small‐animal hyperbaric chamber compatible with in vivo electron paramagnetic resonance (EPR) oximetry using customized OxyTrack oxygen detectors. Up to two detectors were implanted simultaneously to measure pO 2 in skeletal muscle and tumor tissue in mice. Animals were exposed to air (21% O 2 ) and 100% O 2 at both normobaric pressure and 2 atmospheres absolute (ATA) in a stepwise protocol. Results The chamber operated safely and stably up to 2 ATA, enabling continuous real‐time pO 2 monitoring without adverse events and issues with the hardware for up to 90 min. In B16‐F10 melanoma, tumor pO 2 increased from ∼9 mmHg at baseline to ∼26 mmHg at 100% O 2 (1 ATA) and ∼50 mmHg at 2 ATA, with partial retention after decompression. In contrast, SCC7 tumors showed minimal responsiveness (∼9–10 mmHg), whereas skeletal muscle demonstrated marked pressure‐dependent increases (up to ∼50 mmHg). Conclusion The small‐animal HBO chamber enables real‐time, tissue‐specific assessment of oxygen dynamics under clinically relevant hyperbaric conditions. The approach could facilitate optimization of HBO protocols and support translational investigation of oxygen‐modulated therapies, including radiation and drug responses in solid tumors.