Endocrine Abstracts

Introduction: Transcriptional regulation of gene expression by miRNAs is critical for the fine-tuning of adrenal stress response. However, its role in hypercortisolism has not been explored well. The study addresses this gap using adrenal samples of 3 patient groups from the German Cushing’s registry: Cortisol-Producing-Adenoma (CPA), Primary Bilateral Adrenal Hyperplasia (PBMAH) and controls (adrenal samples of patients with pheochromocytoma).

Methods: Next generation sequencing based miRNA profiling and associated target analyses by QPCR were performed. Transcriptomic data of RNA-Seq were analysed and validated by QPCR. For pathway mapping bioinformatic tools (R, String, KEGG, Gprofiler) were used.

Results: miRNA based NGS revealed 23 miRNAs to be differentially expressed between Cushing (PBMAH and CPA) and Controls. Of these, significantly upregulated miRNAs (n=6) were used for validation. Upregulated expression of hsa-miR-139-3p (l2fc>1.4), hsa-miR-1247-5p (l2fc>2.5) and hsa-miR-150-5p (l2fc>1.9) in PBMAH and CPA (vs Controls) could be confirmed by QPCR (P<0.05). Next, the experimentally validated targets of the individual miRNAs were selected from miRWALK and majority of the selected genes were found to be involved in steroid biosynthesis (Alox15, Cyp2b6, Cybrd1). In-vitro and QPCR analyses of the targets are in process. In case of RNA seq, PBMAH was found to have the most dysregulated genes compared to Controls and CPA (n=1248). Pathway mapping using the significantly altered genes in PBMAH gave neuronal synaptic signalling as top hits. Specifically, there was an increased expression (l2fc >5; P<0.05) of dopamine (Drd2) and glutamate receptors (Gria4, Grin2a) in PBMAH. Validation of the pathway analysis is on-going.

Conclusion: This study identifies a miRNA-target gene network in possible steroid biosynthesis dysfunctions in adrenals of patients with Cushing’s syndrome. Additionally, potential changes in neuronal synaptic pathways in PBMAH were identified.