S100A8/A9 Inhibition Reduces Splenic Myelopoiesis and Improves Outcomes After Stroke

Kim, H. A.; Al-Sharea, A.; Chu, H.; Tang, S.-C.; Rupasinghe, S. A.; Zhang, S. R.; Nagareddy, P. R.; Drummond, G. R.; Arumugam, T. V.; Murphy, A. J.; Sobey, C. G.; Lee, M. K.

doi:10.64898/2025.12.03.692236

bioRxiv · 10.64898/2025.12.03.692236

S100A8/A9 Inhibition Reduces Splenic Myelopoiesis and Improves Outcomes After Stroke

Kim, H. A.·Al-Sharea, A.·Chu, H.·Tang, S.-C.·Rupasinghe, S. A.·Zhang, S. R.·Nagareddy, P. R.·Drummond, G. R.·Arumugam, T. V.·Murphy, A. J.·Sobey, C. G.·Lee, M. K.

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Abstract

BackgroundNeutrophils are among the earliest immune cells to infiltrate the ischemic brain and contribute to secondary neuronal damage. The alarmin S100 calcium-binding protein A8/A9 (S100A8/A9), predominantly released by neutrophils, is upregulated during this process. Although the bone marrow is recognised as the principal site of neutrophil production via myelopoiesis, the role of the spleen as an immune-responsive organ remains incompletely understood. MethodsIn this study, we employed a transient middle cerebral artery occlusion (MCAO) model in male C57Bl/6 mice and examined immune responses 24 hours post-stroke in the blood, bone marrow and spleen using flow cytometry. To understand the role of S100A8/A9 in modulating stroke-induced myelopoiesis, we administered a small molecule inhibitor of S100A8/A9, ABR-215757, before and after stroke. ResultsNeutrophils and S100A8/A9 were found in the infarcted brain tissue. Interestingly, we observed a marked increase in splenic neutrophils, accompanied by an expansion of myeloid progenitors, indicating activation of extramedullary myelopoiesis. Given our previous work showing that S100A8/A9 promotes myelopoiesis, we pharmacologically inhibited S100A8/A9 to determine if this would modulate stroke-induced myelopoiesis. Treatment with ABR-215757 at 24 hours post-stroke led to reduced splenic myelopoiesis, reversed neutrophilia, enhanced forelimb grip strength, and a one-third reduction in infarct size. ConclusionThese findings identify the spleen as a key contributor to neutrophil production following stroke and suggest that targeting S100A8/A9 may attenuate post-stroke inflammation and improve neurological recovery. HighlightsO_LIStroke induces extramedullary myelopoiesis in the spleen, not femoral bone marrow. C_LIO_LINeutrophil-derived S100A8/A9 drives splenic myelopoiesis after ischemic stroke. C_LIO_LIPharmacological blockade of S100A8/A9 with ABR-215757 reduces neutrophilia. C_LIO_LIInhibition of S100A8/A9 lessens infarct size and improves neurological recovery. C_LIO_LIHuman stroke tissue confirms S100A8/A9 accumulation with neutrophil infiltration. C_LI

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Kim, H. A., Al-Sharea, A., Chu, H., Tang, S.-C., Rupasinghe, S. A., Zhang, S. R., Nagareddy, P. R., Drummond, G. R., Arumugam, T. V., Murphy, A. J., Sobey, C. G., Lee, M. K.. 2025-12-07. S100A8/A9 Inhibition Reduces Splenic Myelopoiesis and Improves Outcomes After Stroke. https://doi.org/10.64898/2025.12.03.692236

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