Nine-grid segmentation strategy for photoacoustic imaging of microcirculation oxygenation dynamics in the thenar muscle

Microcirculatory dysfunction is central to the pathophysiology of diseases such as sepsis, diabetes, and cardiovascular disorders. While clinical tools like near-infrared spectroscopy (NIRS) and transcutaneous oxygen monitoring (TcPO₂) offer global oxygenation insights, they lack the spatial resolution for localized microvascular assessment. Photoacoustic computed tomography provides sub-millimeter oxygenation imaging, yet resolving ultra-fine structures for morphology-based functional analysis remains challenging. To address this, we developed the Nine-Grid Segmentation Strategy (NGSS), a significance-analysis framework integrated with a non-invasive vascular occlusion test (VOT). Using the microcirculation-rich thenar muscle as the imaging site, NGSS characterizes each pixel across two physiological dimensions: occlusion response and reperfusion efficiency. The NGSS framework categorizes pixels based on their oxygenation trajectories during occlusion and reperfusion, assigning each to one of three functional states: significant increase, non-significant change, or significant decrease. This two-dimensional classification generates a nine-type map capturing the spatial heterogeneity of tissue microregions. Our results demonstrate that even without discerning specific microvascular morphologies, NGSS evaluates tissue oxygenation on a hundred-micron scale. The analysis reveals distinct oxygenation patterns and perfusion efficiencies across phenotypes, providing quantitative volumetric distributions. NGSS offers a novel, high-resolution, non-invasive approach for microcirculatory assessment, showing significant promise for both clinical monitoring and fundamental vascular research.