Serum lysophospholipidome of dietary origin as a suitable susceptibility/risk biomarker of human hypercholesterolemia: A cross-sectional study.

BACKGROUND & AIMS: Whether bioactive lysophospholipids (lyso-PLs) and trimethylamine-N-oxide (TMAO) serve as non-invasive biomarkers in early human hypercholesterolemia (HC) is unknown. This study aimed to assess whether serum lyso-PLs and plasma TMAO may be suitable susceptibility/risk biomarkers of HC in humans. Secondarily, we aimed to evaluate the relationships between targeted metabolites, diet composition and circulating liver transaminases, and verify these results in hamsters. METHODS: A targeted metabolomics and lipidomics approach determined plasma TMAO and serum lysophosphatidylcholines (lyso-PCs) and lysophosphatidylethanolamines (lyso-PEs) in low (L-LDL-c) and moderate to high (MH-LDL-c) LDL-cholesterol subjects. Additionally, the relationships between targeted metabolites, liver transaminases and diet, particularly fatty acid intake, were tested. In parallel, plasma and liver lyso-PL profiles were studied in 16 hamsters fed a moderate high-fat (HFD) or low-fat (LFD) diet for 30 days. RESULTS: Predictive models identified lyso-PC15:0 and lyso-PE18:2 as the most discriminant lyso-PLs among groups. In MH-LDL-c (n = 48), LDL-cholesterol and saturated FAs were positively associated with lyso-PC15:0, whereas in L-LDL-c (n = 70), LDL-cholesterol and polyunsaturated fatty acids (PUFAs) were negatively and positively related to lyso-PE18:2, respectively. Interestingly, in MH-LDL-c, the lower lyso-PE 18:2 concentrations were indicative of higher LDL-cholesterol levels. Intrahepatic accumulation of lyso-PLs-containing essential n-6 PUFAs, including lyso-PE18:2, were higher in HFD-fed hamsters than LFD-fed hamsters. CONCLUSIONS: Overall, results revealed a possible hepatic adaptive mechanism to counteract diet-induced steatosis in animal and hypercholesterolemia progression in humans. In particular, low serum lyso-PE18:2 suggests a suitable susceptibility/risk biomarker of HC in humans.

Chronic Effect of a Cafeteria Diet and Intensity of Resistance Training on the Circulating Lysophospholipidome in Young Rats.

The daily practice of physical exercise and a balanced diet are recommended to prevent metabolic syndrome (MetS). As MetS is a multifactorial disorder associated with the development of serious diseases, the advancement of comprehensive biomarkers could aid in an accurate diagnosis. In this regard, it is known that gut microbiota is altered in MetS, and especially, lipid metabolites species are highly modified, thus emerging as potential biomarkers. In preliminary studies, we observed that alterations in serum lysoglycerophospholipids (Lyso-PLs) were shared between animals with diet-induced MetS and those performing resistance exercises assiduously. Therefore, our objective was the targeted determination of the lysophospholipidome in young rats fed a standard (ST) or a cafeteria diet (CAF) and submitted to different training intensities to evaluate its potential as a biomarker of a detrimental lifestyle. Targeted metabolomics focused on lysophosphatidylcholines (Lyso-PCs) and lysophosphatidylethanolamines (Lyso-PEs) and multivariate statistics were used to achieve an integral understanding. Chronic intake of CAF altered the serological levels of both lipid subclasses. Twenty-two Lyso-PLs were significantly altered by CAF, from which we selected Lyso-PCs (14:0), (17:1) and (20:2) and Lyso-PEs (18:2) and (18:3) as they were enough to achieve an optimal prediction. The main effect of physical training was decreased Lyso-PEs levels with disparities among training intensities for each diet. We concluded that an examination of the lysophospholipidome reveals the general state of the metabolome in young female rats, especially due to intake of an MetS-inducing diet, thus highlighting the importance of this family of compounds in lipid disorders.

Serum lysophospholipid levels are altered in dyslipidemic hamsters.

Dyslipidemias are common disorders that predispose individuals to severe diseases. It is known that healthy living habits can prevent dyslipidemias if they are diagnosed properly. Therefore, biomarkers that assist in diagnosis are essential. The aim of this study was to identify biomarkers of dyslipidemia progression, which in turn disclose its etiology. These findings will pave the way for examinations of the regulatory mechanisms involved in dyslipidemias. Hamsters were fed either a normal-fat diet (NFD) or a high-fat diet. Some of the NFD-fed animals were further treated with the hyperlipidemic agent Poloxamer 407. Non-targeted metabolomics was used to investigate progressive changes in unknown serum metabolites. The hepatic expression of putative biomarker-related genes was also analyzed. The serum levels of lysophospholipids (Lyso-PLs) and their related enzymes lecithin-cholesterol acyltransferase (LCAT), secreted phospholipase A2 (sPLA2) and paraoxonase-1 were altered in dyslipidemic hamsters. Lysophosphatidylcholine levels were increased in diet-induced dyslipidemic groups, whereas lysophosphatidylethanolamine levels increased in response to the chemical treatment. The liver was significantly involved in regulating the levels of these molecules, based on the modified expression of endothelial lipase (Lipg), sPLA2 (Pla2g2a) and acyltransferases (Lcat and Lpcat3). We concluded that Lyso-PL evaluation could aid in the comprehensive diagnosis and management of lipid disorders.

Development and validation of a UHPLC-ESI-MS/MS method for the simultaneous quantification of mammal lysophosphatidylcholines and lysophosphatidylethanolamines in serum.

Recent investigations based on non-targeted metabolomics have proposed lysophospholipids (Lyso-PLs) as biomarkers of different diseases. In particular, lysophosphatidylcholines (Lyso-PCs) and lysophosphatidylethanolamines (Lyso-PEs) have been associated with serious lipid pathologies. Methods to determine the different molecular species in a biological sample and to quantify even less abundant species are required for the evaluation of the Lyso-PL pattern as a novel comprehensive biomarker of dyslipidemia. This study describes the development and validation of an ultra-high-performance liquid chromatography coupled to tandem mass spectrometry assay for the determination of a large number of Lyso-PCs and Lyso-PEs in biological samples. The method was validated in rat serum using two simple methanol-based extractions with low sample volumes (5-50µL) that covered the wide concentration range of these metabolites. In total, thirty-one Lyso-PLs were separated and quantified with low method limits of detection and quantification, reaching values of 0.2 and 0.8nM, respectively. The method was subsequently applied in the identification of Lyso-PL-related changes produced by the chronic intake of a cafeteria diet. The results showed alterations in the majority of Lyso-PCs and Lyso-PEs in rat serum. Furthermore, multivariate analysis indicated that the comprehensive evaluation of serum Lyso-PLs could be an excellent indicator of the nutritional phenotype associated with an increased risk of lipid disorders.

Impact of a cafeteria diet and daily physical training on the rat serum metabolome.

Regular physical activity and healthy dietary patterns are commonly recommended for the prevention and treatment of metabolic syndrome (MetS), which is diagnosed at an alarmingly increasing rate, especially among adolescents. Nevertheless, little is known regarding the relevance of physical exercise on the modulation of the metabolome in healthy people and those with MetS. We have previously shown that treadmill exercise ameliorated different symptoms of MetS. The aim of this study was to investigate the impact of a MetS-inducing diet and different intensities of aerobic training on the overall serum metabolome of adolescent rats. For 8 weeks, young rats were fed either standard chow (ST) or cafeteria diet (CAF) and were subjected to a daily program of training on a treadmill at different speeds. Non-targeted metabolomics was used to identify changes in circulating metabolites, and a combination of multivariate analysis techniques was implemented to achieve a holistic understanding of the metabolome. Among all the identified circulating metabolites influenced by CAF, lysophosphatidylcholines were the most represented family. Serum sphingolipids, bile acids, acylcarnitines, unsaturated fatty acids and vitamin E and A derivatives also changed significantly in CAF-fed rats. These findings suggest that an enduring systemic inflammatory state is induced by CAF. The impact of physical training on the metabolome was less striking than the impact of diet and mainly altered circulating bile acids and glycerophospholipids. Furthermore, the serum levels of monocyte chemoattractant protein-1 were increased in CAF-fed rats, and C-reactive protein was decreased in trained groups. The leptin/adiponectin ratio, a useful marker of MetS, was increased in CAF groups, but decreased in proportion to training intensity. Multivariate analysis revealed that ST-fed animals were more susceptible to exercise-induced changes in metabolites than animals with MetS, in which moderate-intensity seems more effective than high-intensity training. Our results indicate that CAF has a strong negative impact on the metabolome of animals that is difficult to reverse by daily exercise.