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Background <p>γδ T cells boost inflammatory responses and exacerbate tissue damage after ischemic stroke. However, the origin, dynamics, and tissue adaptation of γδ T cells in the ischemic brain and its border regions remain poorly understood. A systematic integration of large-scale datasets is urgently needed. Here, we investigated the impact of ischemic stroke on the state of meningeal and brain-infiltrating γδ T cells and explored their potential contributions to post-stroke inflammation.</p> Methods <p>We conducted an integrated analysis of publicly available single-cell RNA sequencing (scRNA-seq) datasets, which included meningeal and brain-infiltrating <i>Ptprc</i><sup>+</sup> (CD45<sup>+</sup>) immune cells following experimental stroke. γδ T cells were identified and subsequently classified into distinct subtypes through data integration and reference mapping. Subtype-specific functions, tissue residency signatures, migratory programs, and the cellular interactions between γδ T cells and endothelial cells or fibroblasts in the dura and brain were investigated, respectively. Key findings were validated by flow cytometry and immunofluorescence assays in vivo.</p> Results <p>On day 2 post-experimental stroke, the number of parenchymal γδ T cells significantly increased while dural γδ T cells decreased. The majority of γδ T cells residing in the meninges and infiltrating the brain, both under homeostatic conditions and following stroke, were <i>Rorc</i>⁺ and belonged to the Vγ6⁺ γδ17 cell subset. Compared to dural γδ T cells, brain-infiltrating γδ T cells showed reduced tissue residency capacities, higher migratory pathway activation, and lower Ki‑67 positivity, indicating acute recruitment. In contrast, dural γδ T cells exhibited greater IL‑17-producing capacities on day 3. Redistributions of dural γδ T cells were analyzed, and immunofluorescence revealed a close spatial association between dural γδ T cells and CD31<sup>+</sup> cells. Cell–cell communication analysis predicted increased interactions between γδ T cells and CD45<sup>−</sup> cells in both the dura and the brain.</p> Conclusion <p>Our data indicate that most meningeal and brain-infiltrating γδ T cells after stroke share an activated γδ17 phenotype but display compartmentalized dynamics in activation, proliferation, and migration. These results establish a foundation for further studies on the spatially distinct roles of γδ T cells in post-stroke immunity.</p>

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Single-cell omics and flow cytometry identify distinct immune states of dural and brain-infiltrating IL-17-producing γδ T cells after experimental stroke

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年4月9日

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  • Mingming Zha,
  • Alina Jander,
  • Haodi Cai,
  • Marius Piepke,
  • Karoline Degenhardt,
  • Leo Winter,
  • Tim Magnus,
  • Mathias Gelderblom

摘要

Background

γδ T cells boost inflammatory responses and exacerbate tissue damage after ischemic stroke. However, the origin, dynamics, and tissue adaptation of γδ T cells in the ischemic brain and its border regions remain poorly understood. A systematic integration of large-scale datasets is urgently needed. Here, we investigated the impact of ischemic stroke on the state of meningeal and brain-infiltrating γδ T cells and explored their potential contributions to post-stroke inflammation.

Methods

We conducted an integrated analysis of publicly available single-cell RNA sequencing (scRNA-seq) datasets, which included meningeal and brain-infiltrating Ptprc+ (CD45+) immune cells following experimental stroke. γδ T cells were identified and subsequently classified into distinct subtypes through data integration and reference mapping. Subtype-specific functions, tissue residency signatures, migratory programs, and the cellular interactions between γδ T cells and endothelial cells or fibroblasts in the dura and brain were investigated, respectively. Key findings were validated by flow cytometry and immunofluorescence assays in vivo.

Results

On day 2 post-experimental stroke, the number of parenchymal γδ T cells significantly increased while dural γδ T cells decreased. The majority of γδ T cells residing in the meninges and infiltrating the brain, both under homeostatic conditions and following stroke, were Rorc⁺ and belonged to the Vγ6⁺ γδ17 cell subset. Compared to dural γδ T cells, brain-infiltrating γδ T cells showed reduced tissue residency capacities, higher migratory pathway activation, and lower Ki‑67 positivity, indicating acute recruitment. In contrast, dural γδ T cells exhibited greater IL‑17-producing capacities on day 3. Redistributions of dural γδ T cells were analyzed, and immunofluorescence revealed a close spatial association between dural γδ T cells and CD31+ cells. Cell–cell communication analysis predicted increased interactions between γδ T cells and CD45− cells in both the dura and the brain.

Conclusion

Our data indicate that most meningeal and brain-infiltrating γδ T cells after stroke share an activated γδ17 phenotype but display compartmentalized dynamics in activation, proliferation, and migration. These results establish a foundation for further studies on the spatially distinct roles of γδ T cells in post-stroke immunity.