Analysis of urinary sphingolipids using liquid chromatography-tandem mass spectrometry in diabetic nephropathy.

AIMS/INTRODUCTION: Sphingolipids, such as ceramides and sphingosine, are involved in the pathogenesis of diabetes; however, the modulation of urinary sphingolipids in diabetic nephropathy has not been fully elucidated. Therefore, we aimed to develop a simultaneous measurement system for urinary sphingolipids using liquid chromatography-tandem mass spectrometry and to elucidate the modulation of urinary sphingolipids in diabetic nephropathy. MATERIALS AND METHODS: We established a simultaneous measurement system for the urinary sphingosine, dihydrosphingosine, and six ceramide species (Cer d18:1/16:0, Cer d18:1/18:0, Cer d18:1/18:1, Cer d18:1/20:0, Cer d18:1/22:0 and Cer d18:1/24:0), and we examined the urinary sphingolipids in 64 type 2 diabetes patients and 15 control participants. RESULTS: The established measurement system for the urinary sphingolipids showed good precision for Cer d18:1/16:0, Cer d18:1/20:0, Cer d18:1/22:0 and Cer d18:1/24:0. We observed that the urinary levels of Cer d18:1/16:0, Cer d18:1/18:0, Cer d18:1/20:0, Cer d18:1/22:0 and Cer d18:1/24:0 were elevated in patients with stage 3 of diabetic nephropathy, and were correlated with urinary biomarkers, such as albumin and N-acetyl-ß-d-glucosaminidase, and sediment score. CONCLUSIONS: Our method is useful for the measurement of ceramide in urine specimens, and urinary ceramides might be associated with the pathological condition of diabetic nephropathy, such as renal tubular injury.

Distribution in skin of ceramide after oral administration to rats.

In this study, the absorption, distribution and excretion of ceramide were examined in rats. After a single oral administration of (3)H-ceramide, mean plasma concentration of radioactivity reached maximum at approximately 10.67 hr and decreased with a T(1/2) of 67.12 hr. The mean cumulative excretion of radioactivity in urine and feces accounted for approximately 4.79% and 87.44%, respectively, of the dose. At 96 hr after dosing, 1.67% and 3.67%, respectively, of the dose were still present in the skin and carcass. The radioactivity in the skin at 12 hr was lower than that in plasma and the ratio of skin to plasma concentration was 0.7. However, at 120 hr after dosing, the ratio of skin to plasma concentration increased to 4. A detailed analysis of the distribution of radioactivity in a section of skin showed that radioactivity was located in the dermis and epidermis. At 168 hr, the radioactivity in the epidermis was 8.0% of the radioactivity in skin. The results of the present study clearly demonstrate that some ceramide orally administered is distributed gradually in the dermis after gastrointestinal absorption, followed by transfer from the dermis to the epidermis.

Above-normal urinary excretion of urinary ceramides in Farber's disease, and characterization of their components by high-performance liquid chromatography.

We compared the sphingolipid content of urine from a patient with Farber's disease with that of control urine. The ceramides were measured by high-performance liquid chromatography. The patient's urine contained 1.2 mug of ceramides per milligram of creatinine, more than 200-fold the normal amount. The urinary ceramides were isolated by high-performance liquid chromatography for further identification. They contained mainly nonhydroxy fatty acids and only a small quantity of those with 2-hydroxy fatty acids. This contrasts with the previously described composition of the patient's renal and cerebellar tissue. The fatty acid and long-chain base compositions of the urinary ceramides containing nonhydroxy fatty acids were nearly identical to those of the patient's kidney.