this is reminiscent of molecularly distinc t and immune - interacting lec subsets in the nasal mucosa ( 111, 112 ) and dermis ( 113 ). this heterogeneity may arise, in part, from microenvironment signals, such as ifn - γ, which drive context - dependent reprogramming of lecs in inflammation or cancer ( 114 – 116 ). we show that lecs upregulate pvr and lgals9 in response to ifn - γ, echoing responses in the blood endothelium ( 87, 88 ) and supporting a paradigm in which the behavior of lymphatics is actively shaped by their surrounding milieu. in kidney transplantation, lymphatics have been associated with improved graft survival, possibly through increased leukocyte clearance ( 27 – 29 ), but also with immune activation and fibrosis ( 23, 25, 31, 117, 118 ). our findings challenge the notion that lymphangiogenesis is uniformly pathogenic. although we observed lymphangiogenesis and proximity of these vessels to tlss in rejecting allografts, we showed that allograft lecs acquire a tolerogenic transcriptional program driven by ifn - γ. lec - derived immune - inhibitory ligands dampen effector t cell function in cancer ( 119, 120 ), neuroinflammation ( 121 ), and infection ( 69 ), and we confirmed the expression of 2 exemplar molecular candidates, pvr and lgals9, at both the transcript and protein level. however, this tolerogenic molecular program coincides with structural perturbations to allograft lymphatics. in rejection, lymphatics exhibited loss of hierarchical organization, infiltration into the medulla, and transformation of cell - cell junctions from button - to zipper - like morphology, changes known to impair fluid and cell transport ( 68 – 70 ). building on previous studies in kidney ( 26, 27 ) and other inflammatory contexts ( 80, 122, 123 ), we identified tlss of varying maturity positioned along lymphatic networks. given the potential for in situ antigen presentation and t cell activation within the tls ( 75, 77, 78, 124 – 127 ), and given the observed altered localization of cd4 t cells within and around lymphatic vessels, it is tempting to speculate that lymphatic perturbation may contribute to cd4 t cell retention within allografts, heralding the formation and maintenance of the tls in chronic rejection. additionally, we demonstrate that allograft lecs express hla class ii and show c4d deposition in patients with de novo donor - specific antibod ies, consistent with allo antibod y targeting and complement activation. analogous injury to the blood vasculature ( 19 ) is well - characterized in transplant pathology ( 24 ), and donor lymphatics may thus represent a previously underappreciated target of alloimmune responses. this study has several limitations. first, our 3d imaging was cross - sectional and included a small number of fixed samples, restricting inference of dynamic events during transplant rejection. second, and common to all scrna - seq studies of human tissues, our control tissues were derived from nontransplanted kidneys and tumour nephrectomies and are thus likely subject to inflammatory changes. we attempted to mitigate this by using samples with histological evidence of minimal chronic damage. third, although we identified expression of coinhibitory ligands and evidence of allo antibod y binding of kidney lymphatics, the downstream consequences on alloimmunity and graft function require further mechanistic study, which is challenging given the absence of an animal model that mimics the long - term sequalae of chronic mixed rejection, which occurred in our cohort of patients over decades to years, while enabling simultaneous genetic or pharmacological manipulation of lecs in a targeted manner. together, our data provide a comprehensive and multimodal view of the lymphatic vasculature in human kidney health and rejection. we propose that lymphatics acquire a tolerogenic, ifn - γ – driven phenotype during chronic rejection, but this is accompanied by structural disor ganization and immune-associated perturbations. these findings point to a potentially new perspective on the role of lymphatic remodeling in transplantation, featuring a tolerogenic profile yet subject to alloimmune injury. this work lays the foundation for future studies exploring kidneys in health and disease and opens new avenues for therapeutic targeting of the lymphatic vasculature to improve the longevity of kidney transplants. given the exploratory nature of 3d imaging and scrna - seq performed in this study and the limited kidneys available for 3d imaging analysis, sex was not considered as a biological variable. human kidney tissue was fixed in 4 % paraformaldehyde in pbs at 4°c overnight and stored in pbs with 0. 02 % sodium azide. a modified shanel protocol ( 128 ) was used for whole - mount immunolabeling, followed by optical clearing in benzyl alcohol / benzyl benzoate ( 1: 2 ). imaging was performed using an lsm880 upright confocal microscope ( zeiss ) or custom - built mesoscale selective plane illumination microscope ( mesospim ) ( 129 ). image segmentation and 3d reconstruction were carried out in imaris and amira. binarized lymphatic networks were skeletonized in fiji using bonej ( 130 ). cd4 t cell and cd20 b cell counts, centroids, and areas were obtained using 3d objects counter with no further preprocessing ( 131 ). the mean distance of each cell from the nearest point of the lymphatic network ( d ) was calculated using the cross - product 3d point - line distance: source paper: pmc12435838 where x 1 and x 2 are the 2 closest adjacent nodes from the lymphatic 3d skeleton, found by minimizing cross – nearest neighbor distances, and x 0 is the centroid of the cell of interest. to evaluate whether the cell distances were different from what would be expected by chance, within each region of interest, the cd4 t cell and cd20 b cell populations were randomly redistributed under complete spatial randomness for 20 simulations. a comparison was then made as to whether the measured mean cell - lymphatic distances fell within the 95 % cis obtained through the simulations under complete spatial randomness. single - cell suspensions from fresh kidney explants were processed using the 10x genomics chromium 5 ′ v2 kit and sequenced on an illumina novaseq. data were mapped to grch38 and processed using scanpy and seurat, using scvi ( 132 ) or harmony ( 133 ) for integration. cell identity was assigned via marker gene expression and assisted by celltypist prediction. differential expression was assessed using wilcoxon rank - sum tests and go term enrichment using panther. to infer putative cell - cell interactions in scrna - seq data, the cellphonedb resource ( 85 ) was used. to generate the human lymphatic cell atlas, lecs were extracted from publicly available single - cell datasets across multiple organs and integrated using harmony. scenic ( 134 ) was used to infer transcription factor activity across clusters. the nephroseq database ( v5, rrid: scr _ 019050 ) was used to examine candidate genes by pulling data from its online browser. adult human dermal lecs ( promocell, c - 12217 ) were cultured in mv2 medium and treated with recombinant human ifn - γ ( 50 ng / ml ) or unstimulated control medium for 24, 48, or 72 hours. lgals9 transcript levels were quantified by qrt - pcr and normalized to hprt using the 2 method. secreted lgals9 protein in conditioned media was measured by elisa ( r & d systems ). data are shown as fold - change relative to untreated controls. assays were performed across 2 independent cell lines in triplic ate. statistical analyses were performed using graphpad prism unless otherwise specified. data normality and variance were assessed using shapiro - wilk and brown - forsythe tests, respectively. for normally distributed data, comparisons between 2 groups used 2 - tailed student ’ s t test and 1 - way anova with bonferroni ’ s post hoc tests for multiple groups. a p value less than 0. 05 was considered significant.