ISSN 1470-3947 (print) | ISSN 1479-6848 (online)

Endocrine Abstracts (2019) 66 OC4.8 | DOI: 10.1530/endoabs.66.OC4.8

SGPL1 deficiency leads to accumulation of sphingolipid species and downregulation of key enzymes within the steroidogenic pathway

Avinaash Maharaj1, Jack Williams1, Tülay Güran2, Debora Braslavsky3, Josefina Casas4, Louise Metherell1 & Rathi Prasad1


1Centre for Endocrinology, William Harvey Research Institute, John Vane Science Centre, Queen Mary, University of London, Charterhouse Square, London, UK; 2Marmara University, School of Medicine, Department of Paediatric Endocrinology and Diabetes, Istanbul, Turkey; 3Centro de Investigaciones Endocrinológicas ‘Dr. Cesar Bergadá’ (CEDIE) – CONICET – FEI – División de Endocrinología, Hospital de Niños ‘Ricardo Gutiérrez,’, Buenos Aires, Argentina; 4Research Unit on BioActive Molecules (RUBAM), Department of Biomedicinal Chemistry, IQAC-CSIC, Jordi Girona 18–26, Barcelona, Spain


Background: SGPL1 carries out the final degradative step of the sphingolipid pathway, irreversible cleavage of sphingosine-1-phopshate. SGPL1 deficiency is associated with a pathological accumulation of sphingolipid species and a multi-systemic condition incorporating primary adrenal insufficiency (PAI). Sphingolipid intermediates, ceramide and sphingosine are postulated to act as modulators of the steroidogenic pathway, acting as second messengers altering downstream expression of steroid responsive transcriptional elements. Ceramide, sphingomyelin and sphingosine are reported inhibitors of steroidogenesis. Pathological accumulation of these sphingolipid species in SGPL1 deficiency may therefore have negative implications for the steroidogenic cascade.

Objective and hypotheses: Investigating the impact of SGPL1 deficiency on sphingolipid profile and steroidogenesis using patient derived dermal fibroblasts and RNA-seq interrogation of the differential expression of steroidogenic genes in an SGPL1-KD adrenocortical cell line (H295R).

Methods: (1) Primary cultures of dermal fibroblasts were established from skin biopsies of two patients with SGPL1 mutations (Patient1 – p.F545del; Patient 2 – p.S65Rfs*6G) and PAI. The steroidogenic capacity of fibroblasts was explored using a precursor substrate, progesterone, as a stimulator of cortisol production. Culture media from treated/untreated cells were subjected to cortisol measurement (ELISA). (2) Mass spectrometric analysis of sphingolipid intermediates in control and patient fibroblasts 2. Lentiviral shRNA mediated KD of SGPL1 in H295R cell line with subsequent RNA-seq interrogation.

Results: Control fibroblasts showed a significant cortisol response after progesterone stimulation (P<0.001). In comparison Patient 1 fibroblasts were less responsive (P<0.05) and Patient 2 cells unresponsive to stimulation (P<0.001). Mass spectrometry revealed significantly increased ceramide and sphingomyelin levels in both patient cell lines compared to control (P<0.01). Differential gene expression data from RNA-seq in the SGPL1-KD H295R cell line revealed functional enrichment of genes for the metabolism of steroids (FDR 0.045) and cortisol synthesis and secretion (FDR 0.0496). Further interrogation revealed significant downregulation of steroidogenic genes, with reduced transcript levels of STAR, CYP21A2 and CYP11B1 in the SGPL1-KD H295R.

Conclusion: Our results are in keeping with a prominent role for sphingolipids in modulating the acute phase of steroidogenesis, suggesting that alterations in sphingolipid metabolism due to SGPL1 deficiency negatively impact the expression of genes responsible for steroid hormone biosynthesis.