Introduction: LXRs are transcription factors that regulate cholesterol homeostasis and likely modulate other aspects of lipid metabolism. In the testis, tightly regulated lipid metabolism is crucial to maintain fertility. Testicular LXRs are highly expressed but their role in regulating lipid homeostasis is not fully understood. Lxrα/β double knockout male mice (Lxrα/β DKO) are sterile by 7 months of age, with aberrations in lipid metabolism.
Aim: To identify specific disrupted cellular lipids and candidate target genes in the testes of Lxrα/β DKO mice using integrated wide platform studies.
Methods: RNA-seq, quantitative mass spectrometry and mass spectrometry imaging (MSI) were combined to study whole testicular tissues from Lxrα/β DKO mice compared to age and strain matched controls. cDNA libraries were prepared for sequencing using NextSeq-500 and lipid extracts for LC-MS analysis with SONAR acquisition, based on an m/z isolation range of the quadrupole. Results were analysed using LipidMaps and ProgenesisQI for normalised quantitation. For MSI, the MALDI SYNAPT G2-Si was equipped with an ion mobility cell and experiments performed using Waters High Definition Imaging (HDI) 1.4 and MassLynx.
Results: Histological assessment confirmed abnormal seminiferous tubules, germ-cell loss and lipid deposition in Lxrα/β DKO mice. Quantitative lipidomics analysis confirmed statistically significant differences in lipids compared to controls. Retrieved curated targets were mapped with KEGG pathway analysis. Alterations in cholesterol, triglyceride, sphingomyelin and ceramide metabolism were identified. From RNA-seq, 1161 genes (log2 FC −3.49 to +2.17, P<0.01) were differentially expressed in the Lxrα/β DKO with genes relevant to pathways identified from lipidomic data. Finally MSI confirmed deposition of specific lipid species and a visualisation of their location within the testis.
Discussion: An integrative approach using lipidomic analysis with mRNA transcript studies provides data implicating LXRs in novel lipid pathways critical for male reproductive function.
19 Nov 2018 - 21 Nov 2018