SUN-223 Role of the Stress-responsive Transcription Factor NR4A2 in the Development of Aldosterone-producing Cell Clusters

Abstract Disclosure: N. Iwahashi: None. H. Umakoshi: None. Y. Kitamura: None. F. Masamichi: None. T. Ogasawara: None. T. Fukumoto: None. M. Yokomoto-Umakoshi: None. K. Nishimoto: None. S. Ogawa: None. Background: Aldosterone-producing cell clusters (APCCs), which share some transcriptomic features with aldosterone-producing adenomas (APAs), are frequently observed in the adrenocortical tissues of aged individuals and those with primary aldosteronism (PA). However, the molecular mechanisms of how APCCs are formed remains poorly understood. Methods: We performed integrated spatial transcriptomics and single-cell RNA sequencing analysis of a human adrenocortical tissue containing both APA and APCCs. The findings thus obtained were validated by in vitro studies with an adrenocortical carcinoma cell line (H295R) and analysis of adrenal venous sampling (AVS) data. Results: APCC cells, unlike APA cells, exhibited a distinct cellular population with a gene expression profile, which are more similar to that of the zona glomerulosa (ZG) cells than that of APA cells. Through the machine learning approach, we identified LUZP2, CYB5B, CYP21A2, CACNB2, and HOPX as novel APCC-specific marker genes. Gene set enrichment analysis revealed enrichment of stress response pathways in APCCs, with significant activation of the stress-responsive transcription factor NR4A2. In silico perturbation analysis revealed that NR4A2 promotes the formation of APCC cells from ZG cells. In vitro studies showed that knockdown of NR4A2 in H295R cells resulted in reduced expression of the above APCC marker genes and those associated with aldosterone synthesis. Analysis of clinical AVS data showed the enhanced ACTH responsiveness in patients with APCCs, supporting the stress-responsive nature of APCCs. Conclusions: Our data suggest that NR4A2 plays a crucial role in APCC development by linking cellular stress response to enhanced aldosterone production. This study provides novel insight into the molecular mechanisms underlying stress-induced formation of APCCs with enhanced aldosterone production. Presentation: Sunday, July 13, 2025