LGR5 Activates Noncanonical Wnt Signaling and Inhibits Aldosterone Production in the Human Adrenal

Abstract

Context:Aldosterone synthesis and cellularity in the human adrenal zona glomerulosa (ZG) is sparse and patchy, presumably due to salt excess. The frequency of somatic mutations causing aldosterone-producing adenomas (APAs) may be a consequence of protection from cell loss by constitutive aldosterone production.Objective:The objective of the study was to delineate a process in human ZG, which may regulate both aldosterone production and cell turnover.Design:This study included a comparison of 20 pairs of ZG and zona fasciculata transcriptomes from adrenals adjacent to an APA (n = 13) or a pheochromocytoma (n = 7).Interventions:Interventions included an overexpression of the top ZG gene (LGR5) or stimulation by its ligand (R-spondin-3).Main Outcome Measures:A transcriptome profile of ZG and zona fasciculata and aldosterone production, cell kinetic measurements, and Wnt signaling activity of LGR5 transfected or R-spondin-3-stimulated cells were measured.Results:LGR5 was the top gene up-regulated in ZG (25-fold). The gene for its cognate ligand R-spondin-3, RSPO3, was 5-fold up-regulated. In total, 18 genes associated with the Wnt pathway were greater than 2-fold up-regulated. ZG selectivity of LGR5, and its absence in most APAs, were confirmed by quantitative PCR and immunohistochemistry. Both R-spondin-3 stimulation and LGR5 transfection of human adrenal cells suppressed aldosterone production. There was reduced proliferation and increased apoptosis of transfected cells, and the noncanonical activator protein-1/Jun pathway was stimulated more than the canonical Wnt pathway (3-fold vs 1.3-fold). ZG of adrenal sections stained positive for apoptosis markers.Conclusion:LGR5 is the most selectively expressed gene in human ZG and reduces aldosterone production and cell number. Such conditions may favor cells whose somatic mutation reverses aldosterone inhibition and cell loss.

The adrenal capsule is a signaling center controlling cell renewal and zonation through Rspo3

“…This signaling appeared to inhibit aldosterone production and cell renewal and induce apoptosis (Shaikh et al 2015). These findings are in stark contrast to the data presented in this study.…”

NEFM (Neurofilament Medium) Polypeptide, a Marker for Zona Glomerulosa Cells in Human Adrenal, Inhibits D1R (Dopamine D1 Receptor)–Mediated Secretion of Aldosterone

“…This surprising finding, together with their absence from upregulated genes in rodent ZG 26 and patchiness of aldosterone synthase in human adrenal, was interpreted as an adaptive mechanism to high-salt intake in Western diet. 20,21 NEFM encodes one of the subunits forming the neurofilament, the type IV intermediate filament of mature neurons, one of the most abundant proteins in the nervous system. Neurofilament subunits perform extracytoskeletal roles in the neuronal synapses as receptor-interacting proteins.…”

“…20,21 NEFM is also a biomarker for the wild-type, ZG-like subtype of APAs; compared with these, it was 4-fold downregulated in KCNJ5 mutant, ZF-like APAs. 22 We hypothesized that NEFM plays a role in regulating human ZG endocrine function through a similar relationship with D1R as described in the nervous system.…”

Review of Markers of Zona Glomerulosa and Aldosterone-Producing Adenoma Cells

“…A transcriptome analysis of laser capture microdissected ZG and ZF cells from adrenal cortex adjacent to APA or pheochromocytoma showed that expression of LGR5 was 25-fold greater in ZG than in ZF and was the ‘top hit’ among 213 over-expressed genes in ZG compared to ZF. 50 A radial streaking of LGR5 into ZF was observed in some serial sections, presumably reflecting the centripetal migration of ZG cells during development. 50 Expression of the gene coding for the LGR5 cognate ligand R-spondin-3 (RSPO3) was also up-regulated 5-fold in ZG compared to ZF.…”