Single-cell sequencing reveals increased renal lymphatic density alters renal inflammatory pathway activation in kidney injury

Heidi A. Creed, Joseph M. Rutkowski

Division of Lymphatic Biology Department of Medical Physiology Texas A&M University College of Medicine Bryan, TX 77807

In the United States an estimated 50% of ICU patients develop Acute Kidney Injury (AKI) which is associated with a 50% mortality rate. A single occurrence of AKI predisposes a patient to develop chronic conditions such as Chronic Kidney Disease (CKD). The mechanism of the AKI to CKD transition is still unknown but is likely mediated by the extent and length of the inflammatory response following the initial injury. Lymphatic vessels help to maintain tissue homeostasis through fluid, macromolecule, and immune modulation. Increased lymphatic growth, or lymphangiogenesis, often occurs during inflammation and plays a role in acute and chronic disease processes. The primary objective of this study was to understand what roles renal lymphatic endothelial cells (LECs) play in AKI recovery and CKD progression, a role which remains largely unknown. To determine if LECs have protective immunomodulatory protective roles in kidney injury, we utilized uninjured C57/BL6 mice and a single 20 mg/kg dose Cisplatin to induce AKI and isolated renal LECs 72hrs post-injury. We have previously demonstrated that an expanded lymphatic network in KidVD mice increases the CD4:CD8 Tcell ratio and that this may afford protection from an AKI-to-CKD transition. Using scRNA sequencing of renal tissues we defined and identified differences in Cisplatin injured renal inflammatory activation. Differences in gene expression revealed alterations in signaling pathway enrichment between KidVD and WT mice for (1) Vessel permeability; (2) T regulatory cell development; and (3) T cell maturation. These findings illustrate specific differences in immune pathway activation between uninjured and injured renal LECs in regulating the inflammatory response in AKI. Together, this data indicates increased renal lymphatic density may alter immunological outcomes in injury