Introduction: Immune cell infiltration is a hallmark of renal allograft injury, yet standardized markers to assess its clinical relevance in pediatric kidney transplantation (KTx) are lacking. Children have unique immunological characteristics, including increased tolerance and adaptive responses. This study investigates the immune cell composition (focusing on mononuclear phagocytes: monocytes, macrophages, dendritic cells) in pediatric kidney allografts and its association with histologic phenotypes, retransplantation, fibrosis, and graft function.
Methods: We retrospectively analyzed 202 renal biopsies from 59 pediatric recipients (63% male, mean age 10 years) transplanted at MHH between 2000 and 2017. Samples were re-evaluated according to the Banff 2022 criteria. Immunohistochemical staining was performed for CD68, CD206, CD163L1 (macrophages), CD209 (dendritic cells), CD20 (B cells) and CD3 (T cells). Whole slide images were analyzed using QuPath software to quantify immune cell density in cortex, medulla, and extrarenal tissue. mRNA expression of >700 immune-related genes from FFPE tissue was assessed using the NanoString® Human Organ Transplant Panel (B-HOT). Additionally, correlation with clinical data was performed.
Results: Macrophage infiltration was significantly increased in T cell-mediated, antibody-mediated and combined rejection biopsies compared to non-rejection biopsies (p<0.05). B cells predominated in TCMR, whereas dendritic cells were enriched in samples with i-IFTA ≥1 (p<0.05). Retransplanted kidneys showed higher macrophage densities than initial allografts (p<0.05). Protocol biopsies showed significantly fewer immune cells than indication biopsies (p<0.01). Macrophage infiltration correlated positively with fibrosis (IFTA; r>0.4, p<0.001) and serum creatinine. B-HOT data identified ADAMDEC1, AOAH, APOL1 and B2M (mainly expressed by mononuclear phagocytes) as the most significant transcripts and differentiated rejection types by principal component analysis (PCA). Multiplex staining revealed detailed spatial information for the tissue biomarkers.
Conclusion: Mononuclear phagocytes are key cellular players in the pathology of pediatric renal allografts. Antibody-based staining and molecular profiling confirm mononuclear phagocytes as biomarkers of rejection, complement histologic diagnosis, and may improve future graft risk stratification. Our findings highlight the potential of quantitative immune cell profiling to improve diagnostic accuracy and personalized monitoring in pediatric KTx.