Sphingolipids have emerged as key signaling molecules involved in the regulation of a variety of cellular functions. Most research investigating sphingolipids has focused on the intracellular signaling functions of two bioactive sphingolipids - ceramide and S1P, but little attention has been given to the source of delivery of these molecules to cells. The proposed studies will focus on the extracellular transport of not only these sphingolipids but also that of sphingomyelin and the complex hexosyl- and lactosylceramide molecular species in plasma, urine, and in isolated lipoproteins and additionally, determine the changes in their transport which are associated with diabetes and nephropathy. The lipidomics profile of plasma obtained from 633 type 1 diabetic patients at their entry into the Diabetes Control and Complications Trial (DCCT), demonstrated that low plasma concentrations of several ceramide species predicted the development of macroalbuminuria. In contrast, the levels of sphingosine-1-phosphate (S1P), although 2.6-fold higher in diabetic patients compared to matched non-diabetic subjects, did not. Plasma concentrations of apolipoprotein M (ApoM), a physiological carrier of S1P, were significantly increased in Type 1 diabetic patients with albuminuria and ApoM was found in urine from these patients but not in normoalbuminuric patients. It is not known how changes in plasma sphingolipid metabolism contribute to nephropathy in diabetes and what role the individual lipoprotein classes play in this metabolism. We will test the hypothesis that plasma lipoproteins from type 1 diabetes patients, but especially HDL, are markedly enriched in S1P but poor in selected ceramides. Additionally, the decreased content of selected ceramides in HDL leads to their decreased concentrations both in plasma and urine. Lastly, measuring the levels of these ceramides in type 1 diabetes will identify patients at high risk to develop nephropathy at different stages of the disease. To evaluate these hypotheses, we will conduct studies with three Specific Aims: 1) Determine whether the levels of sphingolipids and ApoM measured in the plasma collected at the beginning and at the end of the DCCT trial in a subgroup of patients are associated with the development and progression of diabetic nephropathy; 2) Compare the sphingolipid and ApoM concentrations in plasma and urine from matched Type 1 diabetes patients with normoalbuminuria with those with macroalbuminuria, with non-diabetic subjects with macroalbuminuria, and with healthy controls; 3) Determine if the observed differences in plasma total sphingolipid and ApoM concentrations which are associated with diabetes and albuminuria are localized to VLDL, LDL or HDL. Knowledge of plasma sphingolipid transport in diabetes and in nephropathy will direct our future studies investigating the potential for lipoprotein sphingoliid to alter cell metabolism, especially cells associated with diabetic kidney disease.
Sphingolipids are complex molecules that control a variety of cellular functions. Knowledge of the transport of these lipids in blood is limited. We will measure the changes in the amounts of these lipids that occur in blood from patients with diabetes by comparing the levels with those of non-diabetic subjects and we will determine if the kidney disease that frequently accompanies diabetes further alters the blood levels of these lipids. Also little is known regarding the appearance of these sphingolipids in urine. We will measure the changes in the amounts of these lipids in urine that occur in patients with diabetes and determine if diabetic kidney disease further alters the urine levels of these lipids. We will measure the levels of these sphingolipids in blood and urine samples obtained from diabetic patients before they developed kidney disease to determine whether they are able to identify patients at high risk to develop renal disease and we will also measure them in samples collected from the same patients after they developed the disease to better understand the effects of diabetic kidney disease on these sphingolipids. These studies will expand the limited information available regarding sphingolipid transport in blood and urine in humans and, for the first time, will investigate sphingolipids in the blood and urine from patients with diabetes and their possible role in the development of diabetic nephropathy.
