Cell Atlas at Single-Nuclei Resolution of the Adult Human Adrenal Gland and Adrenocortical Adenomas

Abstract

The human adrenal gland is a complex endocrine tissue. Developmental studies on this tissue have been limited to animal models or human foetus. Here, we present a cell atlas analysis of the adult human normal adrenal gland, combining single-nuclei RNA sequencing and spatial transcriptome data to reconstruct adrenal gland development and tumourigenesis. We identified two populations of potential progenitor cells resident within the adrenal cortex: adrenocortical progenitors NR2F2+-ID1+ cells, located within and underneath the capsule, and medullary progenitors SYT1+-CHGA- cells, located in islets in the subcapsular region. Using pseudotime analyses, we provided evidence of the centripetal nature of adrenocortical cell development and of the essential role played by the Wnt/β-catenin pathway in the adrenocortical self-renewal. By comparing transcriptional profiles of cells of normal adrenal glands and adrenocortical adenomas we revealed a high heterogeneity with six adenoma-specific clusters. Overall, our results give insights into adrenal plasticity and mechanisms underlying adrenocortical tumourigenesis.

Cellular landscape of adrenocortical carcinoma at single-nuclei resolution

“…Using the rationale that a higher log ROP is indicative of a higher proportion of cells derived from cancer samples, we identified eight adrenocortical cell types that appear to be specific to ACC. Conversely, clusters with a lower log ROP include infiltrating cell types, medullary cells and two adrenocortical cell types identified with the ZG and ZR, based on expression of known markers such as DACH1 and SULT2A1 , respectively (Table S1) [28]. Four remaining adrenocortical cell types, which contained approximately equal proportions of cells from ACC and NAG samples (Figure 1B), are identified as representing a progressive continuum of cell types from the ZF based on expression of cytochrome P450 CYP17A1 (ZF 1) and non-canonical NOTCH ligand DLK1 (ZF 2 and 3) (number of ZF cells expressing DLK1 in the normal adult adrenal gland has recently been shown to increase with patient age) [28,32], and a comparatively undifferentiated adrenocortical zone (UZ) expressing very few known marker genes (Table S1).…”

“…Our understanding of ACC pathogenesis has increased considerably over the past decade following in-depth characterisations of the molecular processes involved in development, homeostasis and disease of the adult adrenal gland [2,3,14,15,19–21,27,28]. In the present study, we contribute to this emerging field by providing a cellular transcriptome atlas of ACC at single-nuclei resolution that complements existing data and reveals a mixture of cell types present within the malignant adrenal cortex.…”

“…Among these top marker genes, expression of the RSPO3 receptor LGR4 was also recently detected in the developing adrenal cortex [27] and its mutation is known to drive aberrant adrenocortical zonation [60]. RSPO3 itself is a WNT signalling coactivator that is proposed to play a key role in maintaining the adrenocortical stem cell niche near the adrenal capsule [20,21,24,25,27,28]. This model is once again analogous to the mechanism for maintenance of the LGR5 -positive (homologue RSPO3 receptor) stem cell compartment in colon or intestine, whose perturbation during carcinogenesis involves aberrant differentiation of TACs into absorptive or secretory cell types based on levels of NOTCH signalling [59,61–63].…”

“…Nuclei were isolated from snap-frozen samples derived from ACC tissue dissected by an expert pathologist and obtained during tumour resection using the protocol previously described in [28,69]. Quality and purity of isolated nuclei were confirmed by microscopy and yield was quantified using a Neubauer chamber.…”

“…Although great progress has been made towards delineating the molecular mechanisms of adrenocortical development, renewal and pathogenesis in humans, very little remains known about the landscape of cell types involved in these processes. Notable exceptions include recent single-cell and single-nuclei studies on the foetal [27] and adult [28] human adrenal gland, respectively. While the latter also includes data from a cohort of adrenocortical adenoma (ACA) samples, equivalent data on ACC have hitherto been lacking.…”