Proceedings of The Physiological Society

Europhysiology 2018 (London, UK) (2018) Proc Physiol Soc 41, PCB138

Poster Communications

mCCDcl1 cells show Plasticity Consistent with the Ability to Transition between Principal and Intercalated Cells

A. Assmus1, M. K. Mansley1, L. Mullins1, A. Peter1, J. J. Mullins1

1. CVS, University of Edinburgh, Edinburgh, Grande-Bretagne, United Kingdom.

The cortical collecting duct of the mammalian kidney plays a critical role in the regulation of body volume, sodium pH and osmolarity and is composed of two distinct cells types, principal cells and intercalated cells. Each cell type is detectable in the kidney by the localization of specific transport proteins such as Aqp2 and ENaC in principal cells and V-ATPase B1 and Cx30 in intercalated cells. The inter-relationship between PC and IC cells, both during development and in the adult, is complex and is yet to be fully elucidated. The ratio of the number of principal cells to intercalated cells (typically 70:30 in healthy mice) has been shown to be influenced by multiple factors involved in Notch signalling [1], [2]. mCCDcl1 cells have been widely used as a mouse principal cell line on the basis of their physiological characteristics. In this study, the mCCDcl1 parental cell line and three sub-lines cloned from isolated single cells (Ed1, Ed2, and Ed3) were grown on filters to assess their transepithelial resistance, transepithelial voltage, equivalent short circuit current and expression of the cell-specific markers Aqp2, ENaC, V-ATPaseB1 and Cx30, as well as precursor markers p63 and IC-specific ΔNp63. The parental mCCDcl1 cell line presented typical [3] amiloride-sensitive electrogenic sodium transport indicative of principal cell function, however immunocytochemistry and RT-PCR showed that a subset of cells expressed the intercalated cell-specific markers V-ATPase B1 and Cx30, including cells also positive for Aqp2 and ENaC (intermediate cells). Intermediate cells represented ~42% of the mCCDcl1 cell population and were present in different proportions in the clonal sub-lines. Parental and clonal sub-lines all exhibited expression of collecting duct precursor markers p63 and ΔNp63, and RNA sequencing showed the expression of a large range of other PC and IC specific markers such as ROMK or NKCC1. The vertical transmission of both principal and intercalated cell characteristics via single cell cloning reveals the plasticity of mCCDcl1 cells, and a direct lineage relationship between these two physiologically important cell types, and is consistent with mCCDcl1 cells being precursor cells.

Where applicable, experiments conform with Society ethical requirements