Acyl ethanolamides in Diabetes and Diabetic Nephropathy: Novel targets from untargeted plasma metabolomic profiles of South Asian Indian men

Abstract

The pathophysiology of diabetic nephropathy (DN) in type 2 diabetes (T2D) patients is minimally understood. We compared untargeted high-resolution accurate mass (HRAM) orbitrap-based plasma metabolomic profiles of 31 T2D-DN (with estimated glomerular filtration rate ≤80 mL/min/1.73 m²), 29 T2D and 30 normal glucose tolerance (NGT) Indian men. Of the 939 plasma metabolites that were differentially abundant amongst the NGT, T2D and T2D-DN (ANOVA, False Discovery Rate - FDR adjusted p-value < 0.05), 48 were associated with T2D irrespective of the renal function of the subjects. Acyl ethanolamides and acetylcholine were decreased while monoacylglycerols (MAGs) and cortisol were elevated in both T2D and T2D-DN. Sixteen metabolites, including amino acid metabolites Imidazolelactate and N-Acetylornithine, changed significantly between NGT, T2D and T2D-DN. 192 metabolites were specifically dysregulated in T2D-DN (ratio ≥2 or ≤0.5 between T2D-DN and T2D, similar abundance in NGT and T2D). These included increased levels of multiple acylcarnitine and amino acid metabolites. We observed a significant dysregulation of amino acid and fatty acid metabolism in South Asian Indian male T2D-DN subjects. Unique to this study, we report a reduction in acyl ethanolamide levels in both T2D and T2D-DN males. Those with dysregulation in acyl ethanolamides, which are endogenous agonists of GPR119, are likely to exhibit improved glycemic control with GPR119 agonists.

Figure 1

Multivariate analysis of untargeted metabolomics data using 5781 metabolites. Principal component analysis (PCA) plot of principal component (PC)1 and PC2 for each of the 90 plasma samples from subjects of normal glucose tolerance (NGT: dark blue circles), type 2 diabetes mellitus (T2D: orange circles) and T2D-Diabetic nephropathy (T2D-DN: light blue circles) groups.

Figure 2

Biochemical pathway and chemical relationships network of (a) the 16 metabolites significantly dysregulated between NGT and T2D, T2D and T2D-DN as well as NGT and T2D-DN (FDR adjusted p-value <0.05) and (b) the 48 metabolites significantly associated with T2D irrespective of renal function of the subjects [significantly dysregulated (FDR adjusted p-value <0.05) between NGT and T2D and between NGT and T2D-DN but not between T2D and T2D-DN]. Blue represents downregulated and red represents upregulated metabolites in the diabetic groups (T2D and T2D-DN) compared to NGT. (NGT: normal glucose tolerance, T2D: type 2 diabetes mellitus, T2D-DN: type 2 diabetes mellitus-diabetic nephropathy).

Figure 3

Box plots depicting levels of (a) Hexadecanamide, (b) Lauramide (c) Linoleamide, (d) Oleamide, (e) Palmitoleoyl ethanolamide and (f) Stearamide in normal glucose tolerance (dark blue), type 2 diabetes mellitus (orange) and type 2 diabetes mellitus-diabetic nephropathy (light blue) subjects.

Figure 4

Framework for understanding the role of fatty acyl ethanolamides (FAEs) in precipitating T2D-associated dysglycemia. Lower plasma N-APE levels could lead to lower hypothalamic N-APE levels, upregulating the hypothalamic-pituitary-adrenal (HPA) axis, leading to increased plasma cortisol levels, central adiposity and development of dysglycemia associated with T2D. Proposed steps stemming from the findings of the current study are highlighted in bold while those lacking available evidence are highlighted with question marks. N-APE: N-acylphosphatidylethanolamine, N-APE-PLD: N-acylphosphatidylethanolamine phospholipase D, CRH: Corticotropin-releasing Hormone, ACTH: Adrenocorticotropic Hormone.