Kidney Week

Abstract: FR-PO556

Differentiated Cell Types in the Kidney's Thick Ascending Limb: Functional Insights on TAL α and TAL β

Session Information

• Fluid, Electrolyte, and Acid-Base Disorders: Basic
  October 25, 2024 | Location: Exhibit Hall, Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Fluid, Electrolytes, and Acid-Base Disorders

• 1101 Fluid, Electrolyte, and Acid-Base Disorders: Basic

Authors

• Bahena-López, Jessica Paola, Oregon Health & Science University, Portland, Oregon, United States

Jessica Paola Bahena-López, MD

• Nelson, Jonathan W., University of Southern California, Los Angeles, California, United States

Jonathan W. Nelson, PhD

• Su, Xiao-Tong, Oregon Health & Science University, Portland, Oregon, United States

Xiao-Tong Su, PhD

• Demirci, Hasan, Charite Universitatsmedizin Berlin, Berlin, Berlin, Germany

Hasan Demirci, MS, MSc

• Curry, Joshua N., Oregon Health & Science University, Portland, Oregon, United States

Joshua N. Curry, MD, PhD

• Bleich, Markus, Christian-Albrechts-Universitat zu Kiel Islamwissenschaft, Kiel, Schleswig-Holstein, Germany

Markus Bleich, MD, PhD

• Bachmann, Sebastian, Charite Universitatsmedizin Berlin, Berlin, Berlin, Germany

Sebastian Bachmann, PhD

• Ellison, David H., Oregon Health & Science University, Portland, Oregon, United States

David H. Ellison, MD, FASN

Background

The Thick Ascending Limb (TAL) plays a key role in reabsorbing NaCl and Ca⁺⁺ and Mg⁺⁺, driven ultimately by NKCC2. Spanning from the outer medulla to the cortex, the TAL has traditionally been viewed as a single functional cell type, but recent transcriptomic findings indicate the existence of two TAL cell clusters, with high Claudin 10 and Claudin 16 expression (PMID 28028216). Beyond mosaic expression of claudins, however, the specifc functions of these two cell types are unknown. Here, to better understand the heterogeneity within the TAL population, we enriched for TAL nuclei and performed single-nucleus transcriptomics.

Methods

Kidneys from male NKCC2-Cre-INTACT (Isolation of Nuclei TAgged in specific Cell Types, which fluorescently labels nuclei) mice were harvested for single-nucleus RNA-seq to create an enriched TAL dataset. Data were compared with re-analyzed datasets from mouse (PMID 31689386), rat (PMID 37023747) , and human (PMID 37468583). IHC of kidney from the same species was performed.

Results

Enriched TAL sn-RNA sequencing identified two cell clusters: TAL α and TAL β. Differential Gene Expression analysis showed distinct gene profiles. TAL α cells express Cldn10 and factors regulating sodium transport, including Wnk4, Stk39, and Avpr2, and display arachidonic acid signaling genes like Ptger3. TAL β express Cldn16 and genes related to calcium homeostasis,such a Casr, Pth1r and Vdr, as well as higher levels of Kcnj10 and Wnk1. TAL clustering also shows segregation following a gradient-like pattern across kidney zones, revealing TAL subtypes or states unique to each zone. This transcriptional heterogeneity was similar among mouse, rat, and human. Immunofluorescence confirmed two cell types. In the cortex and OSOM, Kir4.1 was coexpressed with CaSR and Claudin-16 (TAL β), while apical ROMK was observed in TAL α cells, expressing Claudin-10. In the ISOM, Kir4.1 and ROMK maintained their exclusive patterns, with Claudin-10 present in both cell types.

Conclusion

Combined morphological and transcriptomic analyses identified two distinct TAL cell types, each linked to specific transport pathways, presenting a novel model of interconnected ion transport in the TAL.

Funding

• NIDDK Support