Single-Cell Atlas Reveals Macrophages That May Drive Chronic Kidney Transplant Rejection

Kidney transplantation is often a lifesaving procedure, but the long-term success of a transplanted organ remains stubbornly limited by a process that clinicians and researchers still do not fully understand: chronic rejection. While acute rejection episodes can often be managed with immunosuppressive drugs, chronic rejection unfolds slowly and insidiously over months and years, ultimately causing the transplanted kidney to scar, stiffen, and fail. Now, a team of researchers in China has produced one of the most detailed maps to date of the immune cells operating inside a rejecting kidney graft, and their findings point to a specific population of macrophages, and a specific molecular regulator inside them, as potential architects of the fibrotic damage that destroys long-term graft function. The study, published in the Journal of Translational Medicine, combines two powerful single-cell technologies to trace how the immune environment of a transplanted kidney changes from the earliest inflammatory crisis to the late, scar-dominated phase of chronic rejection.

The research team, led by Bingxuan Zheng, Junbo Li, Qi He, and senior author Chenguang Ding, established a mouse model of kidney transplantation in which donor kidneys were transplanted into unrelated recipients, recreating the immunological conflict that occurs in human transplantation. Rather than sampling the graft at a single moment, the investigators collected immune cells at multiple post-transplant stages, capturing the transition from acute rejection, when the immune assault is at its most violent, to the chronic phase, when fibrosis and gradual functional decline take hold. This longitudinal design is critical, because chronic rejection is not a static condition but a dynamic process in which the cellular players and their relationships shift over time. By sampling across that timeline, the researchers could watch the immune landscape evolve rather than merely snapshot its endpoint.

The technological core of the study is the pairing of single-cell RNA sequencing with single-cell ATAC sequencing. Single-cell RNA sequencing, or scRNA-seq, measures which genes are actively being transcribed in each individual cell, revealing the identity and functional state of thousands of cells simultaneously. Single-cell ATAC sequencing, or scATAC-seq, measures which regions of chromatin are physically accessible in each cell’s genome, exposing the regulatory landscape that determines which genes a cell could potentially activate. By analyzing both layers of information in the same immune cell populations, the researchers gained not just a census of which cells were present, but insight into the transcription factor programs driving their behavior. The team supplemented these core assays with computational tools including CellChat for mapping communication between cell types, RNA velocity and Monocle3 for inferring developmental trajectories, UMAP for visualization, and Weighted Gene Co-expression Network Analysis for identifying gene modules correlated with disease states.

Out of this multi-omic atlas emerged a striking finding: a late-stage, terminal-like state of macrophages defined by expression of the gene SPP1, which encodes secreted phosphoprotein 1, also known as osteopontin. These SPP1-positive macrophages were markedly enriched in chronically rejecting allografts compared with earlier stages. Macrophages are the tissue-resident scavengers and sentinels of the immune system, capable of enormous functional plasticity, and the study’s trajectory analyses suggest that the SPP1-positive population represents a terminal differentiation state that macrophages in the graft progressively adopt as rejection moves from acute inflammation toward chronic fibrosis. In other words, these cells appear to be not transient visitors but an end-stage product of immune evolution within the failing graft.

Position matters in tissue immunology, and the SPP1-positive macrophages were not merely abundant; they were central. When the researchers constructed ligand-receptor interaction networks to map how different immune cell populations communicate with one another, the SPP1-positive macrophages occupied a hub position in a macrophage-centered communication network. This means that many of the signaling conversations shaping the graft’s immune environment either originated from or passed through these cells. A cell at the center of such a network has outsized influence: it can recruit other immune cells, shape their differentiation, and coordinate collective behaviors such as tissue remodeling. The implication is that SPP1-positive macrophages may act as orchestrators of the fibrotic microenvironment rather than passive bystanders in a deteriorating graft.

The chromatin accessibility data added a crucial mechanistic dimension. Motif enrichment analysis of the scATAC-seq data revealed that the open regulatory regions in SPP1-positive macrophages were characterized by binding sites for CEBPβ, a transcription factor known as CCAAT/enhancer-binding protein beta. This finding identifies a CEBPβ-centered regulatory program as a candidate driver of the SPP1-positive macrophage state. Even more tellingly, the genes under this regulatory program were associated with sustained activation of extracellular matrix remodeling pathways. The extracellular matrix is the protein scaffold that gives tissue its structure, and its excessive deposition and remodeling is the defining feature of fibrosis. A regulatory program that keeps matrix-remodeling genes switched on in macrophages provides a plausible molecular link between chronic inflammation and the scarring that ultimately destroys graft function.

Perhaps the most intriguing cell-cell interaction identified in the study was a regulatory axis connecting SPP1-positive macrophages with CD8-positive tissue-resident memory T cells, abbreviated Trm cells. Tissue-resident memory T cells are a specialized population of T lymphocytes that take up long-term residence in tissues rather than circulating through blood and lymph nodes. In transplanted organs, they are increasingly recognized as persistent local actors that can sustain immune attack independently of the systemic immune system. The integrative ligand-receptor and pathway analyses in this study suggest that SPP1-positive macrophages and CD8-positive Trm cells engage in a predicted interaction network that is associated with the fibrosis-related immune remodeling characteristic of chronic rejection. This macrophage-T cell axis offers a concrete cellular partnership around which future mechanistic studies can be designed.

The significance of this work lies partly in its methodology and partly in its framing. Most studies of transplant rejection have relied on bulk tissue analysis, which averages signals across millions of cells and can mask the critical contributions of rare or state-specific populations. By resolving the graft immune environment at single-cell resolution across time, and by integrating gene expression with chromatin accessibility, the researchers have produced what they describe as a high-resolution longitudinal immune atlas of kidney allograft rejection. Within that atlas, SPP1-positive macrophages and the CEBPβ regulatory program stand out as candidate contributors to the fibrotic process. If validated, these targets could open therapeutic avenues that current immunosuppression does not address, since standard drugs broadly dampen lymphocyte activation but are not designed to reprogram macrophage differentiation or block matrix remodeling programs.

The authors are careful to emphasize the limits of what their study establishes. The work was performed in a mouse transplantation model, and the findings are framed as candidate regulators and predicted interaction networks rather than proven causal mechanisms. Functional validation, in which the activity of SPP1 or CEBPβ is experimentally manipulated to test whether fibrosis is altered, remains necessary. Human translation is an additional hurdle, since the immune environment of human transplanted kidneys may differ in important ways from the mouse model, and sampling human graft tissue at multiple time points presents practical and ethical challenges. The study was conducted under approved animal protocols at Xi’an Jiaotong University, and the researchers state that their findings offer a framework for future mechanistic and therapeutic investigations rather than immediate clinical application.

Even with those caveats, the study adds an important piece to the puzzle of why transplanted kidneys fail over the long term. Chronic rejection has long been viewed as an intractable, slowly accumulating process, but single-cell multi-omics is beginning to reveal that it has an identifiable cellular logic: specific immune cell states, governed by specific transcription factors, communicating through specific ligand-receptor pairs, drive the tissue toward scarring. By naming SPP1-positive macrophages, CEBPβ, and the CD8-positive tissue-resident memory T cell axis as central players, this atlas gives transplant immunologists a concrete set of hypotheses to test and, potentially, a new generation of therapeutic targets to pursue. For the hundreds of thousands of transplant recipients worldwide whose grafts slowly fail despite the best available immunosuppression, that kind of molecular specificity is exactly what the field has been waiting for.

Subject of Research: Single-cell multi-omics of immune cell dynamics during chronic rejection of transplanted kidneys

Article Title: Single-cell multi-omics characterization of rejection in transplanted kidneys reveals key cell types and candidate regulators of chronic rejection

Article References: Single-cell multi-omics characterization of rejection in transplanted kidneys reveals key cell types and candidate regulators of chronic rejection. (n.d.). https://doi.org/10.1186/s12967-026-09002-3

Image Credits: AI Generated

DOI: 10.1186/s12967-026-09002-3

Keywords: kidney transplantation, chronic rejection, single-cell RNA sequencing, single-cell ATAC sequencing, macrophages, SPP1, CEBPβ, CD8-positive tissue-resident memory T cells, fibrosis, extracellular matrix remodeling, transplant immunology, multi-omics