Chemotaxis overrides the killing response in alloreactive CTLs, providing vascular immune privilege during cellular rejection
Thomas Barba, … , Faddi G. Lakkis, Olivier Thaunat
Thomas Barba, … , Faddi G. Lakkis, Olivier Thaunat
Published May 28, 2025
Citation Information: J Clin Invest. 2025;135(14):e155191. https://doi.org/10.1172/JCI155191.
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Research Article Immunology Nephrology

Chemotaxis overrides the killing response in alloreactive CTLs, providing vascular immune privilege during cellular rejection

Abstract

Graft endothelial cells (ECs) express donor alloantigens and encounter cytotoxic T lymphocytes (CTLs) but are generally spared during T cell–mediated rejection (TCMR), which predominantly affects epithelial structures. The mechanisms underlying this vascular immune privilege are unclear. Transcriptomics analyses and endothelial-mesenchymal transition assessments confirmed that the graft endothelium was preserved during TCMR. Coculture experiments revealed that endothelial and epithelial cells were equally susceptible to CTL-mediated lysis, ruling out cell-intrinsic protection. Intravital microscopy of murine kidney grafts and single-cell RNA-Seq of human renal allografts demonstrated that CTL interactions with ECs were transient compared with epithelial cells. This disparity was mediated by a chemotactic gradient produced by graft stromal cells, guiding CTLs away from ECs toward epithelial targets. In vitro, chemotaxis overrode T cell receptor–induced cytotoxicity, preventing endothelial damage. Finally, analysis of TCMR biopsies revealed that disruption of the chemotactic gradient correlated with endothelialitis lesions, linking its loss to vascular damage. These findings challenge the traditional view of cell-intrinsic immune privilege, proposing a cell-extrinsic mechanism, in which chemotaxis preserves graft vasculature during TCMR. This mechanism may have implications beyond transplantation, highlighting its role in maintaining vascular integrity across pathological conditions.

Authors

Thomas Barba, Martin Oberbarnscheidt, Gregory Franck, Chantal Gao, Sebastien This, Maud Rabeyrin, Candice Roufosse, Linda Moran, Alice Koenig, Virginie Mathias, Carole Saison, Valérie Dubois, Nicolas Pallet, Dany Anglicheau, Baptiste Lamarthée, Alexandre Hertig, Emmanuel Morelon, Arnaud Hot, Helena Paidassi, Thierry Defrance, Antonio Nicoletti, Jean-Paul Duong Van Huyen, Yi-Chung Xu-Dubois, Faddi G. Lakkis, Olivier Thaunat

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Figure 4

Chemotaxis protects graft endothelium against alloreactive T cell–mediated cytotoxicity in vitro.

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Chemotaxis protects graft endothelium against alloreactive T cell–mediat...
(A) UMAP plot of 4 cell clusters (ECs, epithelial cells, stromal cells, and infiltrating T cells) identified by scRNA-Seq analysis of a rejected kidney allograft. (B) Bubble plots comparing the expression of 18 cytokines genes by ECs (end), epithelial cells (tub), and stromal (str) cells of the rejected graft. Bubble size is proportional to the percentage of cells in a cluster expressing the gene, and color intensity is proportional to the average scaled gene expression level (avg. expr). (C) Relative expression of CXCL12 (left panel) and CXCR4 (right panel) overlaid on UMAP plot. Color intensity is proportional to average scaled gene expression. (D–H) In vitro modeling of dynamic and static CTL–target cell interactions. (D) Schematic representation of the dynamic and static assays. chemok., chemokine. (E) ciGENC target cells were cultured with 80 ng/mL CXCL12 in static (sta) or dynamic (dyn) assay, with nonspecific (left column) or allospecific (right column) CTLs (CTV stained, blue). Target cell destruction was monitored by the decrease in live cell area (calcein, green) and acquisition of the apoptosis marker (ethidium bromide, red) measured by fluorescence microscopy. Representative images at the end of cocultures are shown (scale bars: 50 μm). (F) Quantification of ciGENC target cell destruction. Results (mean ± SEM) of 2 independent experiments are shown. ****P < 0.0001, by unpaired, 2-sample Wilcoxon test. (G) The same experiments were conducted with increasing concentrations (0–320 ng/mL) of CXCL12 and other chemokines (CXCL9 and CCL2). Where indicated, the chemokine was placed in the upper (top, 320 ng/mL) chamber of the assay. Each symbol shape represents an individual experiment (CXCL12, n = 4; CXCL9, n = 3; CCL2, n = 3). Small symbols are replicates, and larger symbols indicate the mean. (H) The same experiments (n = 2) were performed using the tubular epithelial cell line HK2 as target cells. ****P < 0.0001, by two-sided Student’s t test. Data are presented as mean ± SEM (G and H).