ESPE Abstracts

Loss of function mutations in SGPL1 (sphingosine-1-phosphate lyase) are associated with a syndromic form of primary adrenal insufficiency. A third of affected male individuals also have evidence of early primary gonadal insufficiency, with an undervirilised phenotype at birth. SGPL1 carries out irreversible breakdown of the signalling molecule sphingosine-1-phosphate (S1P) and deficiency leads to accumulation of S1P and other upstream sphingolipid intermediates to varying degrees in affected tissues. In our in vitro model of the gonadal disease, CRISPR-engineered knock-out (KO) of Sgpl1 in the MA10 immortalised Leydig cell line, we have previously demonstrated reduced expression of key steroidogenic enzymes and reduced progesterone production in response to forskolin stimulation. To further interrogate the mechanism underpinning this reduced steroidogenesis we carried out RNASequencing analysis, sphingolipid and wider lipidomic analysis by LCMS. Sphingolipid analysis revealed significant accumulation of S1P and sphingosine in our Sgpl1 - KO MA-10 cells as compared to WT. There was no significant difference in total ceramides or more complex sphingolipids such as sphingomyelin which form a major component of the sphingolipid metabolome in cells. This delicate imbalance in sphingolipid intermediates in SGPL1 deficient Leydig cells results in dysregulation of genes involved in cholesterol metabolism and we see reduced protein expression of enzymes involved in de novo cholesterol metabolism. On lipidomic analysis, we find reduced total cholesterol and furthermore reduced triaclyglycerol and cholesterol esters, key components of intracellular lipid droplets required for acute steroidogenesis. The interplay between disrupted sphingolipid and cholesterol metabolism further impedes steroid hormone synthesis in SGPL1 deficiency. Sphingosine is purported to attenuate the activity of steroidogenic factor 1 (SF-1), which governs transcriptional regulation of multiple processes required for steroidogenesis, including cholesterogenesis. Work is currently underway to determine whether this plays a role in Leydig cell dysfunction in SGPL1 deficiency. Ongoing evaluation of gonadal function would be recommended in all affected male patients, including those who do not present with disease at birth as there may be a risk of evolving insufficiency.