In this thesis, metabolic profiling (MP) platforms were utilised to interrogate the manifestation of cardiovascular disease and provide candidate biomarkers. A number of LC-MS and NMR methodologies were employed. Data processing was followed by assessment using multivariate (MVDA) and univariate (UV) statistics. MP is applied under three cardiovascular disease themes: 1) plaque rupture, 2) plaque formation, and 3) arterial ectopic calcification. Statistically significant features were structurally assigned. Identified metabolites were mapped to their corresponding biochemical pathways.
For MP of ruptured plaque, tissue from symptomatic and asymptomatic patients for stroke was used. After detection of statistically significant features and structural assignment, two biochemical pathways showed dysregulations: the arachidonic acid pathway, indicating increased levels of inflammation, and the β-oxidation pathway with increased levels of three acyl-carnitines.
Tissue extracts were used to investigate plaque formation. Arterial intima tissue, incorporating plaque lesions (carotid and femoral), was compared to intimal thickening tissue. Intima thickening demonstrated distinct MP compared to plaques. Plaques from different anatomical locations also demonstrated altered MP. After metabolite assignment, pathway mapping revealed dysregulations common to both anatomical locations. These were cholesterol, ceramide, purine, pyrimidine and β-oxidation pathways. These pathways are related to inflammation and apoptosis. A metabolite previously unassociated to atherogenesis was detected with strong statistical significance (t-test; p≥9.8x10-12), namely phosphatidylethanolamine-ceramide. It also demonstrated high correlations to cholesterol, a well-established risk-factor of atherosclerosis.
The third theme of the project explores ectopic cardiovascular calcification. Experiments were conducted on blood serum. Patients with coronary artery and aortic valve calcification were compared with non-calcified controls. Phosphatidylcholine moieties and sphingomyelins were the major discriminating metabolites between cases and controls. These are involved in inflammation and apoptosis. The two diseases manifested different profiles with only three commonly dysregulated metabolites.
A number of experiments using additional samples and bottom-up approaches will follow to provide validation of results.