Endocrine Abstracts

Sphingosine-1-phosphate lyase 1 insufficiency syndrome (SPLIS) is a multisystemic syndrome in which primary adrenal insufficiency (PAI) and steroid resistant nephrotic syndrome predominate, secondary to loss-of-function mutations in SGPL1 (sphingosine-1-phosphate lyase). SGPL1 carries out the irreversible breakdown of sphingosine-1-phosphate, a bioactive sphingolipid intermediate, with implicated roles in various cellular processes. Wider endocrinopathy including gonadal insufficiency and hypothyroidism are described. We aimed to conduct a retrospective analysis to determine the extent of gonadal insufficiency in our SPLIS patient cohort and the wider literature and develop an in vitro model to study the potential impact of SGPL1 on gonadal steroidogenesis. A third of male patients presented with primary gonadal insufficiency (all with concomitant adrenal disease), with microphallus and bilateral cryptorchidism, suggestive of reduced androgen exposure in utero. Where tested, individuals showed an exaggerated response during LHRH stimulation and poor androgen response to HCG stimulation. All, excepting 1 individual, died in early infancy with multi-systemic disease. Mortality in the condition is high (approximately 50% in childhood) and in those surviving male patients pubertal delay has yet to be reported. No impact on gonadal function has been reported in girls with the condition. Accordingly, we generated CRISPR-engineered knock-out (KO) of Sgpl1 in the MA10 immortalised Leydig cell line. Sanger sequencing confirmed a single base ‘A’ insertion in Exon 7, predicting a frameshift mutation and premature stop codon in the KO clone. This was further validated by western blotting demonstrating loss of SGPL1 expression in the KO as compared to wild type (WT). WT and KO cell lines were stimulated with forskolin for 6 hours, with significantly reduced progesterone production seen in the KO. This was associated with decreased steroidogenic enzyme STAR and CYP11A1 expression by western blotting, both in unstimulated and forskolin stimulated conditions in the KO Leydig cells. MTT assays also demonstrated decreased cell proliferation in the KO cell line.

Conclusion: SGPL1 deficiency should be considered in the differential diagnosis of 46XY infants with differences in sex development (DSD) and PAI. SGPL1 deficiency impairs steroidogenesis in Leydig cells and clinicians need to be mindful of evolving gonadal disease in patients with SPLIS.