Renal specific expression of the novel second peptide ligand for the apelin receptor, Elabela/Toddler, in human and mouse tissues

D. Nyimanu¹, C. Read¹, T. Williams¹, P. Yang¹, R. E. Kuc¹, A. Hadjantonakis1², K. M. O\'Shaughnessy¹, J. J. Maguire¹, A. P. Davenport¹. ¹Experimental Medicine & Immunotherapeutics,, University of Cambridge,, Cambridge, United Kingdom, ²Developmental Biology Program, Sloan Kettering Institute, New York.

Introduction: The apelin signalling pathway is important for the regulation of cardiovascular function and fluid homeostasis(1). Elabela (ELA) was recently discovered as a second apelin receptor ligand pivotal to normal cardiac development. Our aim was to investigate expression of the apelinergic system in human kidney and compare this to tissues obtained from apela knockout (KO), heterozygous (Het) and wildtype (WT) mice.

Methods: Human kidneys were obtained with informed consent and ethical approval. Five major organs (heart, kidney, spleen, liver and brain) were obtained from adult (≥12 weeks old) CD-1 WT,Het and KO mice(2). mRNA expression of the apelin system in mouse and human tissues were assessed using RT-qPCR. Autoradiography (using 0.5nM [I¹²⁵]apelin-13 with non-specific binding defined using 2μM [Pyr¹]apelin-13) in human and mouse kidney; and dual-labelling immunofluorescence staining using specific markers of the renal nephron in human kidney were used to characterize the distribution of apelin, ELA and apelin receptor protein. Data are mean±sem (n=6 patients;n=4 animals/group) and were analysed using One-Way ANOVA with Tukey’s multiple comparisons test.

Results: The mRNA levels of ELA, apelin or apelin receptor in the cortex compared to their level in the medulla were similar (n=6, p>0.05 in each case). In WT, Het and KO mice, apelin receptor mRNA was expressed in all organs investigated. High levels of apelin were detected in heart, kidney and brain with much lower levels in spleen and liver. In contrast, ELA was highly expressed in kidney with lower levels in other organs. Autoradiography revealed a dense distribution of apelin binding sites in cortex and medulla of human kidneys, but in mouse kidneys dense expression was observed only in the inner medulla. Immunohistochemistry demonstrated apelin receptor, apelin and ELA were all localised to the endothelium of renal blood vessels, epithelial cells of the proximal convoluted tubules and descending limb of the loop of Henle. Additionally, apelin receptor was localised to macula densa cells, epithelial cells of the thick ascending limb of the loop of Henle and distal convoluted tubule.

Conclusions: We report for the first time cell-specific expression of apelin receptor, apelin and ELA in human kidney and confirm the renal specific expression of ELA in mouse. These data suggest that the apelinergic system may have yet unidentified roles in the regulation of renal physiology.

References:

(1) Yang P et al. (2016). Trends Pharmacol Sci 36: 560-567.

(2) Freyer et al. (2017) Cell Rep 20: 2116-2130