Bioactive sphingolipid profile in a xenograft mouse model of head and neck squamous cell carcinoma

Abstract

The purpose of this study was to determine the profile of bioactive sphingolipids in xenograft mouse tissues of head and neck squamous cell carcinoma. We utilized UHPLC-MS/MS to determine the profile of full set of ceramides, sphingosine, and sphingosine 1-phosphate in this xenograft mouse model. The tissues isolated and investigated were from brain, lung, heart, liver, spleen, kidney, bladder, tumors and blood. With the exception of equal volume of blood plasma (100ul), all tissues were studied with the same amount of protein (800ug). Results demonstrated that brain contained the highest level of ceramide and kidney had the highest level of sphingosine, whereas sphingosine 1-phosphate and dihydrosphingosine 1-phosphate were heavily presented in the blood. Brain also comprised the highest level of phospholipids. As for the species, several ceramides, usually present in very low amounts in cultured tumor cells, showed relatively high levels in certain tissues. This study highlights levels of bioactive sphingolipids profiles in xenograft mouse model of head and neck squamous cell carcinoma, and provides resources to investigate potential therapeutic targets and biomarkers that target bioactive sphingolipids metabolism pathways.

Fig 1. Levels of total Cer, Sph, dhC16-Cer, and dhSph present in various tissues.

Tissues were homogenized in lysis buffer containing protease-inhibitor cocktail before centrifugation to get protein. Then equal amounts of protein (800ug) were provided for analysis of SL. Results are presented as pmol SL/mg protein with means ± st dev. of 6x replicates. A.Total Cer, * p<0.05 (vs brain), ** p<0.05 (vs liver); B.dhC16-Cer, * p<0.05 (vs tumor), ** p<0.05 (vs liver); C. Sph, * p<0.05 (vs kidney), ** p<0.05 (vs heart); D. dhSph, * p<0.05 (vs brain), ** p<0.05 (vs bladder).

Fig 2. Predominant Cn-Cer species in eight tissues.

Results are presented as pmol Cn-Cer/mg protein with means ± st dev. of 6x replicates. A. C18-Cer; B.C18:1-Cer; C. C20-Cer; D. C20:1-Cer; E. C26-Cer; F. C26:1-Cer.

Fig 3. Cn-Cer’s profile in various tissues.

Results are presented as pmol Cn-Cer/mg protein with means ± st dev. of 6x replicates. A. Brain; B. Heart; C. Kidney; D. Lung; E. Bladder; F. Spleen; G. Liver; H. Blood plasma; I. Tumor xenograft.

Fig 4. Levels of total phosphate in various tissues.

Results are presented as nmol/mg protein with means ± st dev. of 6x replicates. * p<0.05 (vs brain), ** p<0.05 (vs bladder).