Considering hypertonicity in the interpretation and analysis of cell type-specific gene expression pattern in the collecting duct

The different parts of the renal nephron have different functions that are related to reabsorption or secretion processes. This is mediated by genes that show a distinct expression pattern along the renal nephron segments (1). The majority of the segments contain only one epithelial cell type. However, the collecting duct is built up by principal cells (PCs), type A intercalated cells (A-ICs), and type B intercalated cells (B-ICs). Chen et al. (2) applied RNA sequencing on the single-cell level using enriched PCs, A-ICs, and B-ICs. These data clearly show major differences in the transcriptomes of the different cell types. Besides known genes, several other transcripts are identified, which allows discrimination of PCs from A-ICs, etc. The authors highlight the importance of this study because the transcriptomes of these cell types were missing. Since the collecting duct is responsible for regulation of blood pressure and body fluid composition, this information could help for a better understanding of physiological and pathological conditions.

However, the underlying mechanisms that control these specific gene expression patterns are not fully understood. One aspect that Chen et al. should also consider is the environmental hypertonicity, especially the cells in the inner medullary collecting duct which are exposed to varying and markedly increasing hypertonicity. Hypertonicity itself alters several intracellular processes including cell volume regulation, cell cycle, intracellular ion homeostasis, and macromolecular as well as nucleic acid stability, and can induce apoptosis (3). A general response to increasing osmolality is the activation of the transcription factor tonicity enhancer binding protein (TonEBP) leading to increased transcription of genes involved in transport or synthesis of organic osmolytes (4–7). In a recent study, we have performed global gene expression profiling to identify changes that were induced by hypertonicity in primary cultured rat inner medullary collecting duct cells (8). These cells still express the collecting duct-specific genes like Aqp2 to -4, TonEBP, epithelial sodium channel, or UTA2 (9, 10). In this cell model, we observed massive changes in gene expression upon changes in tonicity. We identified several novel and so-far-undescribed genes that were affected by hypertonicity. Comparison with other available public databases and the described renal segment-specific gene expression pattern showed that hypertonicity can control the kidney and even segment-specific gene expression pattern. The hypertonicity-induced genes included Aqp2, Avpr2, Fxyd4, or Cdo1, which are also selective PC markers. However, for example, Fstl1, also a specific PC marker, is down-regulated in expression by hypertonicity. This kind of regulation can also be observed for selective IC markers. For example, Fxyd2 expression is induced by hypertonicity and expression of Slc26a4 is down-regulated.

Therefore, the hypertonicity should be considered in the interpretation and analysis of the data by Chen et al. The integration of the hypertonicity-affected changes in gene expression into data in the analysis of nephron segment, IC or PC specific, would be of great benefit under physiological and pathophysiological conditions since in this case the control in gene expression is partly known.