Endocrine Abstracts

Context: Transcriptomic based characterization of signaling pathways in the adrenals of different subtypes of Cushing’s syndrome.

Patients: For preliminary NGS analyses, a total of 27 adrenal samples were used. The cohort consisted of the following patient groups: Cortisol producing adenoma (CPA, n=9), primary bilateral macronodular adrenal hyperplasia (PBMAH, n=10). The adjacent normal adrenal tissue from pheochromocytoma patients (n=8) served as controls. For QPCR validation, the patient groups included: Cushing’s disease (CD, n=8), ectopic Cushing’s syndrome (n=3). Controls included adrenal tissues of aldosterone producing adenoma (n=10) and normal adrenals from patients who underwent kidney surgery (n=10).

Methods: Next-generation sequencing was performed in the 27 adrenal samples (Illumina HiSeq). Bioinformatic analyses was done by R to identify significantly differently expressed genes between the groups. For pathway mapping bioinformatic tools (ShinyGO and Gprofiler) were used. The significant genes related to the respective pathways were validated by real-time reverse transcription-qPCR.

Results: With reference to transcriptomic data PBMAH was found to have the most dysregulated genes compared to Controls (n=5394) and CPA (n=248). Pathway mapping using the significantly altered genes gave neuronal synaptic signalling pathways and PPARG (peroxisome proliferator-activated receptor- gamma) signalling pathway as top hits in the groups of PBMAH and CPA. Validation of the pathway genes identified PPARG (l2fc < -1.5) and its related genes - FABP4 (l2fc < -5.5), PCK1 (l2fc < -2.1), PLIN1 (l2fc < -4.1) and ADIPOQ (l2fc < -3.3) to be significantly downregulated (P<0.005) in CS subtypes - CPA, CD and PBMAH in comparison to the controls. The in vitro mechanistic characterization of this pathway in cortisol production using adrenal cell lines is in process.

Conclusion: This study investigated for the first time PPARG pathway, which plays a critical physiological role in lipid and glucose metabolism, in cortisol regulation and found a significant downregulation of the pathway in the adrenals of CS patients.