Investigating the identity and role of the collecting duct intermediate cell

Abstract

The classical model of the collecting duct (CD) defines two functionally distinct epithelial cell types - principal cells (PC) and intercalated cells (IC). A third, intermediate cell type, which expresses both PC and IC markers, has also been described in the literature but its exact role is yet to be determined. Plasticity within the CD has also been noted, both in vivo and in mCCDcl1 cells leading to questions regarding the origin of these cells and the mechanism underlying their differentiation.

This work details the investigation of the intermediate cell type in the mCCDcl1 and mpkCCDcl4 cell lines. Using immunocytochemistry, RT-PCR and scRNA-Seq, both cell lines were determined to widely express classical PC, IC and recently identified intermediate cell markers. Heterogeneity of mpkCCDcl4 cell populations was sustained through single cell cloning indicating plasticity within the cell line. In line with previous work on the mCCDcl1 cell line, it is concluded that the mpkCCDcl4 cells is heterogenous and not a PC cell line as once thought. As these cell lines mirror what is observed in vivo, this suggests that the mCCDcl1 and mpkCCDcl4 cell lines are appropriate for studying CD cellular composition and plasticity, in particular the intermediate cell type that is still poorly understood.

Despite this, functional differences were noted in the mCCDcl1 and mpkCCDcl4 cells lines in terms of sodium transport, with scRNA-Seq data leading to the hypothesis that up- and down-regulation of certain genes upstream of the epithelial sodium channel (ENaC) and those related to intracellular cholesterol storage, may be in part, be responsible.

Understanding the mechanisms of plasticity within the CD will improve understanding of the kidney both under physiological and pathophysiological conditions. This work reports on one such condition where CD plasticity is observed – in mouse and rat models of the syndrome of apparent mineralocorticoid excess (SAME), conferred by an HSD11B2 knockout. A shift in cell fate from PC to intermediate cell was observed in knockout animals which was not recovered at maturity. Similar studies in mice on a high salt diet and under the ACTH-induced model of Cushing’s syndrome displayed no alteration in cell type composition. The results in the SAME study indicate the importance of cell plasticity activation under certain genetic diseases and pose further questions about the role and function of the intermediate cell type.