Alteration in the liver metabolome of rats with metabolic syndrome after treatment with Hydroxytyrosol. A Mass Spectrometry and Nuclear Magnetic Resonance - based metabolomics study

Metabolic syndrome (MetS) represents a group of abnormalities that enhances the risk for cardiovascular disease, diabetes and stroke. The Mediterranean diet seems to be an important dietary pattern, which reduces the incidence of MetS. Hydroxytyrosol (HT) - a simple phenol found in olive oil - has received increased attention for its antioxidant activity. Recently, the European Foods Safety Authority (EFSA) claimed that dietary consumption of HT exhibits a protective role against cardiovascular disease. In this study, an experimental protocol has been setup, including isolated HT administration in a diet induced model of MetS in young Wistar rats, in order to find out whether HT has a protective effect against MetS. Rats were randomly divided into two groups nurtured by high-carbohydrate high-fat (H) (MetS inducing diet) and high-carbohydrate high-fat + HT (HHT). HT (20 mg/kg/d oral gavage, water vehicle) was administered for 8 weeks on the basal diet. Previous pharmacological evaluation of HT showed that hepatic steatosis was reduced and the inflammatory cells into the liver were infiltrated. These indicate that HT shows bioactivity against metabolic syndrome. Therefore, the metabolomics evaluation of liver extracts would indicate the putative biochemical mechanisms of HT activity. Thus, the extracts of liver tissues were analyzed using Ultra Performance Liquid Chromatography – High Resolution Mass Spectrometry (UPLC-HRMS, Orbitrap Discovery) and Nuclear Magnetic Resonance (NMR) spectroscopy (Bruker Avance III 600 MHz). Multivariate analysis was performed in order to gain insight on the metabolic effects of HT administration on the liver metabolome. Normalization employing multiple internal standards and Quality Control–based Robust LOESS (LOcally Estimated Scatterplot Smoothing) Signal Correction algorithm (QC-RLSC) was added in the processing pipeline to enhance the reliability of metabolomic analysis by reducing unwanted information. Experimentally, HHT rats were clearly distinguished from H in PLS-DA, showing differences in the liver metabolome between the groups and specific biomarkers were determined supporting the pharmacological findings. More specifically, HT has shown to be effective towards the mobilization of lipids as various lipid classes being differentially regulated between the H and HHT groups. Interestingly branched fatty acid esters of hydroxy oleic acids (OAHSA) lipids have been shown to be up regulated to the HHT group, denoting the alleviation of the MetS to the animals administered with HT