| Author: | Sham, Tung Ting |
| Title: | Mass spectrometry-based untargeted metabolomics studies of selected chronic diseases : hyperlipidemia, non-alcoholic fatty liver disease and chronic kidney diseases in diabetes |
| Advisors: | Mok, Kam-wah Daniel (ABCT) |
| Degree: | Ph.D. |
| Year: | 2020 |
| Subject: | Metabolism -- Disorders Mass spectrometry Biochemical markers Drugs -- Metabolism -- Analysis Hong Kong Polytechnic University -- Dissertations |
| Department: | Department of Applied Biology and Chemical Technology |
| Pages: | xxvii, 303 pages : color illustrations |
| Language: | English |
| Abstract: | Chronic metabolic disorders are prevalent worldwide and become the leading health hazard and economic burden in the model world, especially three prevalent disorders commonly in modern society: hyperlipidemia, non-alcoholic fatty liver disease and diabetes. Herein, the objective is to apply untargeted metabolomics approaches for discovery of biomarkers, evaluation of potential drug therapy, and metabolic changes of these three prevalent chronic metabolic diseases in biological samples using high-resolution mass-spectrometry. Two animal studies, using piceatannol alone and an aqueous extract of a Chinese herbal medicine, Polygoni Cuspidati Rhizoma et Radix (PCRR), and a clinical observation study were conducted to demonstrate the importance of untargeted metabolomics to unveil the underlying mechanisms of diseases and treatments. The study design, analytical methods and data treatment were optimized with untargeted metabolomics workflows to fit different sample conditions. The first research is the study is on the investigation of therapeutic effects of piceatannol on high fat diet (HFD)-induced hypercholesterolemic rats using UPLC-QTOF-MS and GC-MS. Untargeted serum metabolomics analysis of normal, hypercholesterolemic and piceatannoltreated SD rats revealed a series of bile acids and fatty acids that showed significant changes among different groups. The significant changes in the ratios of fatty acids suggested down-regulation of stearoyl-CoA desaturase and up-regulation of Δ5-desaturase activities. Quantitative analysis further validated the piceatannol's action in reducing the serum levels of primary and secondary bile acids and upregulating those of most conjugated bile acids. Secondary bile acids are gut-biotransformed metabolites from primary bile acids. The reductions of primary and secondary bile acid levels implied that the gut microbiota may play a role in the treatment of hypercholesterolemia using piceatannol. More reduction of CYP7A1 protein expression and increased levels of most conjugated bile acids in piceatannol-treated rats compared with HFD-fed group suggested that the cholesterol-lowering effect of piceatannol supplementation did not rely on conversion of cholesterol into bile acids by CYP7A1 in the bile acid biosynthesis. The cholesterol-lowering effect might be a result of inhibition of cholesterol absorption from the intestine by the binding effect of piceatannol on the dietary cholesterols with bile acid micelles. The second study is to evaluate the therapeutic effect of polyphenol-rich water extract from PCRR using serum metabolomics plus liver lipidomics of SD rat that were affected by HFD-induced non-alcoholic fatty liver diseases (NAFLD). Chemical analysis showed that PCRR water extract is rich in organic acids, stilbenes, flavonoids and other phenolic glucosides. Serum untargeted metabolomics revealed that bile acid synthesis is the major disturbed pathway after intake of PCRR water extract. Elevation of CYP7A1 protein expression indicated the role of PCRR in removal of cholesterols via facilitation of bile acid biosynthesis. Liver lipidomics also showed dramatic difference of lipid profile change between the HFD-fed animal group and the PCRR-treated group. Putatively identified resveratrol and/or polydatinderived metabolites via microbial transformation were solely detected in the serum of the PCRR-treated group, indicating the action of gut microbiota had participated in the biotransformation. PCRR is likely to have action upon multiple targets to bring out hepato-protective effect. The third study aims to identify metabolites associated with the development and risk of chronic kidney diseases (CKD), in Chinese diabetic patients. Two independent cohorts of normal and diabetic patients covering five stages of CKD were recruited at different period to cross-check the results. Over 30 identified metabolites showed a positive relationship with glomerular filtration rate (GFR) and an inverse correlation to urinary albumin to creatinine ratio (UACR). Robust correlations of serum succinyladenosine, pseudouridine and 2-(α-Dmannopyranosyl)-L-tryptophan and L,L-TMAP with MDRD GFR in type 2 diabetic patients were observed, less dependent to sex than serum creatinine. Their associations were early observed when GFR > 60. Furthermore, prediction of MDRD GFR and CKD stages using these biomarkers alone were comparable with serum creatinine. Stepwise linear regression selected pseudourdine and L,L-TMAP as significant and independent predictors adding in the regression equation of MS-detected creatinine and gender in prediction of MDRD GFR. Combination of these biomarkers with serum creatinine improved the diagnostic ability of the serum creatinine alone in differentiation of early CKD stages. Increased production of many uremic retention solutes detected by UPLC-MS and their reduction of kidney clearance by renal degeneration accounted for the progression of CKD in diabetes. Such impairment also might be partly attributed to the increased inflammatory stimuli of diabetic redox imbalance and dysregulation of amino acid and related pathways. Our finding shows the biomarkers are potential supplement to existing clinical markers of early CKD development in diabetic patients and/or better than serum creatinine. In combination with existing biochemical technique and other platforms, these three studies of chronic diseases demonstrated that an untargeted metabolomics approach is a useful and advanced technique for an overview of the metabolic status and discovery of the biological pathways associated to the disease pathogenesis and exploration of biological actions of drug treatments. The action of gut microbiota that interacted with diet, phytochemicals and endogenous metabolites in the three studies should be further examined. Future prospects on more organs, tissues and biofluids with integration of metabolomics and other omics tools would broaden the spectra of metabolites and give a comprehensive view of the molecular processes associated to the chronic diseases. |
| Rights: | All rights reserved |
| Access: | open access |
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