NOVEL THERAPEUTIC APPROACHES TARGETING TERTIARY LYMPHOID STRUCTURES, AN INFLAMMATORY MICROENVIRONMENT ACCELERATING KIDNEY INJURIES

Tertiary lymphoid structures (TLSs) are ectopic lymphoid aggregates in non-lymphoid organs. We demonstrated that TLSs develop in aged injured kidneys and promote inflammation with maladaptive repair, contributing to AKI to CKD progression. Our previous study identified two types of age associated lymphocytes, CD153+PD-1+CD4+ senescence-associated T (SA-T) cells and CD30+ age-associated B cells (ABCs), driving TLS expansion and kidney injury via the CD153-CD30 pathway. However, whether therapies targeting CD153-expressing cells can reduce TLSs and ameliorate kidney injury remains unknown.

CD153 knock-in (KI) mice expressing enhanced green fluorescent protein (EGFP) and human diphtheria toxin receptor (hDTR) under the CD153 promoter were generated. In aged CD153KI mice after renal ischemia-reperfusion injury (IRI), CD153+ SA-T cells were traced by EGFP expression and depleted by diphtheria toxin (DT) after TLS formation. As a second model, anti-CD153 antibodies were administered to aged C57BL/6J mice post-IRI. In both models, immune cells in kidneys and spleens were analyzed by flow cytometry and immunostaining. TLS size, fibrosis, tubular injury, proliferation, and chemokine/cytokine expression were evaluated by PAS and Sirius-red staining, immunostaining, in situ hybridization (ISH), Edu proliferation assay, and qPCR. Human kidney samples containing TLSs were analyzed by single-cell RNA-seq (scRNA-seq) and validated histologically.

In injured kidneys of aged CD153KI mice, EGFP was exclusively expressed in most SA-T cells, but not in other hematopoietic cells, or parenchymal cells such as fibroblasts and tubular cells. hDTR expression was confined to EGFP+CD4+ T cells within TLSs. The administration of DT depleted SA-T cells in injured kidneys with TLSs, leading to TLS disappearance within 7 days and rapid reduction of Il21 and Ifng expression, key SA-T-derived cytokines for B cell differentiation. These changes were followed by decreased expression of Cxcl9, Ccl19, and Tnfsf13b (encoding BAFF) by fibroblasts within TLSs and reduction of GCB in injured kidneys. Edu assay showed reduced lymphocyte proliferation in residual TLSs. Similarly, anti-CD153 antibodies reduced SA-T cells, diminished TLSs, and ameliorated fibrosis and tubular injury in aged injured kidneys. scRNA-seq of human kidneys with TLSs to investigate counterparts of murine SA-T cells revealed follicular helper T (Tfh)-like cells with high TNFSF8 (encoding CD153) expression. Immunostaining and ISH identified TNFSF8+ T cells within human TLSs, which were considered potential counterparts of murine SA-T cells.

These findings demonstrated that SA-T cells within TLSs are essential for TLS maintenance, potentially via driving chemokine/cytokine production by proinflammatory fibroblasts, and their interactions may sustain lymphocyte proliferation. Renoprotective effects of anti-CD153 antibodies via reduction of SA-T cells and TLS size and the presence of TNFSF8+ Tfh-like cells in human renal TLSs suggested that CD153-directed therapy may be a novel strategy to prevent AKI to CKD progression in the elderly patients.

This work was first presented at ASN Kidney Week 2024 and re-submission is permitted by ASN.