P38 Spatial transcriptomic mapping of the wound microenvironment: unravelling the gene expression correlates of senescence and wound repair

Abstract Introduction and aims Chronic skin wounds affect 2.2 million patients who are elderly and/or diabetic (UK), yet current treatments remain inadequate due to limited understanding of the molecular and cellular correlates of poor healing. Indeed, our group first demonstrated that cellular senescence contributes to delayed wound repair in vivo. Thus, clearance of senescent cells could present a targeted therapeutic strategy to promote skin repair. In this study, we aimed to spatially characterize immunosenescence within human chronic wounds and in vivo models of delayed wound repair to discern which cell types drive pathological senescence. We also synthesized a senescence radiopharmaceutical, [18F]FPyGal, to enable live tracking of tissue senescence across the wound healing continuum. Methods Flow cytometry and spatial proteomic imaging (MACSima platform) characterized immunosenescence in selected tissues (including skin and wounds) in vivo [young/aged (C57BL/6J) and diabetic (db/db) mice] and in patient chronic wounds. High-performance liquid chromatography validated synthesis of [18F]FPyGal, which was injected (IV) into mice to assess biodistribution via positron emission tomography/computed tomography imaging. Results We show that both lymphoid and myeloid populations are highly senescent in aged and diabetic mouse wounds. This profiling also revealed that diabetic unwounded skin retains highly senescent populations, indicating intrinsic skin senescence in diabetes. Interestingly, we identified a highly senescent CD4+CD8+ T-cell subset, providing functional insight into undercharacterized wound-lymphoid populations. These findings extend to human chronic wounds, where increased immunosenescence is linked to clinical outcome. Optimization of [18F]FPyGal synthesis generated a radiochemical purity of > 98%. Initial in vivo testing confirmed limited adverse effects of [18F]FPyGal, enabling future wound validation Conclusions These data reveal a previously unappreciated role for immunosenescence in delayed wound repair, presenting an exciting avenue for therapeutic targeting. Successful synthesis of the [18F]FPyGal radiotracer will enable future temporospatial studies elucidating how local and systemic tissue senescence contribute to cutaneous wound pathology.