Metabolomic comparison followed by cross-validation of enzyme-linked immunosorbent assay to reveal potential biomarkers of diabetic retinopathy in Chinese with type 2 diabetes

Abstract

PurposeTo identify the biomarkers in the critical period of development in diabetic retinopathy (DR) in Chinese with type 2 diabetes using targeted and untargeted metabolomics, and to explore the feasibility of their clinical applicationMethodsThis case-control study described the differential metabolites between 83 Chinese type 2 diabetes mellitus (T2DM) samples with disease duration ≥ 10 years and 27 controls matched cases. Targeted metabolomics using high-resolution mass spectrometry with liquid chromatography was performed on plasma samples of subjects. The results were compared to our previous untargeted metabolomics study and ELISA was performed to validate the mutual differential metabolites of targeted and untargeted metabolomics on plasma. Multiple linear regression analyses were performed to adjust for the significance of different metabolites between groups.ResultMean age of the subjects was 66.3 years and mean T2DM duration was 16.5 years. By cross-validating with results from previous untargeted metabolomic assays, we found that L-Citrulline (Cit), indoleacetic acid (IAA), 1-methylhistidine (1-MH), phosphatidylcholines (PCs), hexanoylcarnitine, chenodeoxycholic acid (CDCA) and eicosapentaenoic acid (EPA) were the most distinctive metabolites biomarkers to distinguish the severity of DR for two different metabolomic approaches in our study. We mainly found that samples in the DR stage showed lower serum level of Cit and higher serum level of IAA compared with samples in the T2DM stage, while during the progression of diabetic retinopathy, the serum levels of CDCA and EPA in PDR stage were significantly lower than NPDR stage. Among them, 4 differential key metabolites including Cit, IAA, CDCA and EPA were confirmed with ELISA.ConclusionThis is the first study to compare the results of targeted and untargeted metabolomics via liquid chromatography-mass spectrometry to find the serum biomarkers which could reflect the metabolic variations among different stages of DR in Chinese. The progression of DR in Chinese at different critical stages was related to the serum levels of specific differential metabolites, of which there is a significant correlation between DR progression and increased IAA and decreased Cit, hexanoylcarnitine, CDCA, and EPA. However, larger studies are needed to confirm our results. Based on this study, it could be inferred that the accuracy of targeted metabolomics for metabolite expression in serum is to some extent higher than that of untargeted metabolomics.

Causal relationships between blood metabolites and diabetic retinopathy: a two-sample Mendelian randomization study

“…Recent studies revealed that the pathological processes of DR were associated with long-term metabolic disturbances ( 9 , 27 , 28 ). With the advancement of metabolomics, metabolites and metabolic pathways associated with DR are continuously being discovered ( 10 , 27 , 29 ). These crucial metabolites are often considered potential biomarkers for disease prediction.…”

Untargeted Multiomics Approach Coupling Lipidomics and Metabolomics Profiling Reveals New Insights in Diabetic Retinopathy

“…To date, a substantial amount of research has reported potential novel biomarkers for DR in human plasma in samples from patients with T1D or T2D with different disease durations and DR stages, including citrulline, indoleacetic acid, 1-methylhistidine, chenodeoxycholic acid, eicosapentaenoic acid, glutamine and glutamic acid, 1,5-gluconolactone, 2deoxyribonic acid, gluconic acid, fumaric acid, uridine, acetic acid, and cytidine [13,18,[20][21][22][23][24][25][26][27][28][29]. Several of these, such as 3,4-dihydroxybutyric acid (3,4-DHBA), were validated as independent risk markers for DR progression in a study of 648 individuals with T1D [27].…”

Recent Advances and Perspectives in Relation to the Metabolomics-Based Study of Diabetic Retinopathy

“…However, this association lost significance after adjusting for creatinine values. Notably, a comparison between targeted and non-targeted metabolomics results unveiled the ability of citrulline and carnitine to differentiate the severity of DR using both approaches [ 55 ]. Combined with proteomics research, it has been discovered that tryptophan metabolism also plays a crucial role in the development of DR.…”

“…NO is a crucial vasodilator that plays a vital role in maintaining vascular endothelial health [ 49 ]. Insufficient NO levels, combined with elevated levels of polyamines and proline, can lead to endothelial cell dysfunction and impaired vasodilation, and can also stimulate cell proliferation and fibrosis [ 55 ]. However, it should be noted that while arginine can stimulate insulin release in pancreatic β cells, poorly regulated arginine and citrulline levels can also contribute to retinal endothelial cell dysfunction [ 55 , 103 ].…”

“…Insufficient NO levels, combined with elevated levels of polyamines and proline, can lead to endothelial cell dysfunction and impaired vasodilation, and can also stimulate cell proliferation and fibrosis [ 55 ]. However, it should be noted that while arginine can stimulate insulin release in pancreatic β cells, poorly regulated arginine and citrulline levels can also contribute to retinal endothelial cell dysfunction [ 55 , 103 ]. Moreover, the metabolism of tryptophan, glutamic acid, and alanine has also been associated with the development of DR, although their precise mechanisms remain unclear [ 63 , 85 ].…”