Comparative Evaluation of Different Targeted and Untargeted Analytical Approaches to Assess Greek Extra Virgin Olive Oil Quality and Authentication.

Extra virgin olive oil (EVOO) is a key component of the Mediterranean diet, with several health benefits derived from its consumption. Moreover, due to its eminent market position, EVOO has been thoroughly studied over the last several years, aiming at its authentication, but also to reveal the chemical profile inherent to its beneficial properties. In the present work, a comparative study was conducted to assess Greek EVOOs' quality and authentication utilizing different analytical approaches, both targeted and untargeted. 173 monovarietal EVOOs from three emblematic Greek cultivars (Koroneiki, Kolovi and Adramytiani), obtained during the harvesting years of 2018-2020, were analyzed and quantified as per their fatty acids methyl esters (FAMEs) composition via the official method (EEC) No 2568/91, as well as their bioactive content through liquid chromatography coupled to high resolution mass spectrometry (LC-HRMS) methodology. In addition to FAMEs analysis, EVOO samples were also analyzed via HRMS-untargeted metabolomics and optical spectroscopy techniques (visible absorption, fluorescence and Raman). The data retrieved from all applied techniques were analyzed with Machine Learning methods for the authentication of the EVOOs' variety. The models' predictive performance was calculated through test samples, while for further evaluation 30 commercially available EVOO samples were also examined in terms of variety. To the best of our knowledge, this is the first study where different techniques from the fields of standard analysis, spectrometry and optical spectroscopy are applied to the same EVOO samples, providing strong insight into EVOOs chemical profile and a comparative evaluation through the different platforms.

High-fat feeding influences the endocrine responses of pubertal rats to an acute stress.

BACKGROUND/AIMS: Studies conducted in adult rats have shown that increased fat intake affects brain energy homeostasis and stress response. The neuroendocrine circuits controlling the aforementioned functions continue to mature during puberty. The aim of the present study was to investigate whether post-weaning high-fat consumption can modify the endocrine responses of pubertal rats to an acute stress. METHODS: Weaning male and female Wistar rats on postnatal day (P) 22 were fed either a high-fat (HF; 45% calories from fat) or a control (10% calories from fat) diet and were sacrificed on the individual day of puberty onset (between P35 and P42) under basal conditions or 4 h after swimming stress. Plasma insulin, leptin and corticosterone levels were determined by radioimmunoassay and the respective receptors in the hypothalamus and hippocampus were determined by Western blot analysis. RESULTS: Stressed HF-fed males showed a smaller increase in plasma insulin levels than chow-fed males. Their leptin receptor levels were reduced in the hypothalamus, but not in the hippocampus, and their glucocorticoid receptor levels were increased in the hypothalamus compared to stressed chow-fed males. HF-fed females were nonresponsive to stress-induced alterations in plasma glucose and corticosterone levels, as well as to hippocampal insulin receptors following stress. Several sex differences were also revealed in the endocrine responses of HF-fed animals following stress. CONCLUSIONS: These data show that consumption of high-fat foods during preadolescence can modify the endocrine responses to an acute stress by affecting both stress and metabolic mediators in a sexually dimorphic manner.

Fat feeding of rats during pubertal growth leads to neuroendocrine alterations in adulthood.

Juvenile obesity is a rising epidemic due largely to consumption of caloric dense, fat-enriched foods. Nevertheless, literature on fat-induced neuroendocrine and metabolic disturbances during adolescence, preceding obesity, is limited. This study aimed to examine early events induced by a fat diet (45% calories from saturated fat) in male rats fed the diet during the pre- and post-pubertal period. The neuroendocrine endpoints studied were the levels of circulating leptin, insulin and corticosterone, as well as their receptors in the hypothalamus and hippocampus. Hormonal levels were determined by radioimmunoassay and receptors' levels by western blot analysis. Leptinemia was increased in pubertal rats and in adult rats fed the fat diet from weaning to adulthood, but not in those fed from puberty to adulthood. Modifications in the developmental pattern from puberty to adulthood were observed for most of the brain receptors studied. In adult animals fed the fat diet from weaning onwards, the levels of leptin receptors in the hypothalamus and glucocorticoid receptors in the hippocampus were decreased compared to chow-fed controls. Switching from fat to normal chow at puberty onset restored the diet-induced alterations on circulating leptin, but not on its hypothalamic receptors. These data suggest that when a fat-enriched diet, resembling those consumed by many teenagers, provided in rats during pubertal growth, it can longitudinally influence the actions of leptin and corticosterone in the brain. The observed alterations at a preobese state may constitute early signs of the disturbed energy balance toward overweight and obesity.

Adult consequences of post-weaning high fat feeding on the limbic-HPA axis of female rats.

The peripubertal period is critical for the final maturation of circuits controlling energy homeostasis and stress response. However, the consequence of juvenile fat consumption on adult physiology is not clear. This study analyzed the adult consequences of post-weaning fat feeding on limbic-hypothalamic-pituitary-adrenal (HPA) axis components and on metabolic regulators of female rats. Wistar rats were fed either a high fat (HF) diet or the normal chow from weaning to puberty or to 3 months of age. Additional groups crossed their diets at puberty onset. Plasma leptin, insulin, and corticosterone levels were determined by radioimmunoassay and their brain receptors by western blot analysis. Adult HF-fed animals though not overweight, had higher corticosterone and reduced glucocorticoid receptor levels in the hypothalamus and hippocampus, compared to the controls. The alterations in HPA axis emerged already at puberty onset. Leptin receptor levels in the hypothalamus were reduced only by continuous fat feeding from weaning to adulthood. The pre-pubertal period appeared more vulnerable to diet-induced alterations in adulthood than the post-pubertal one. Switching from fat diet to normal chow at puberty onset restored most of the diet-induced alterations in the HPA axis. The corticosteroid circuit rather than the leptin or insulin system appears as the principal target for the peripubertal fat diet-induced effects in adult female rats.

Fat diet affects leptin receptor levels in the rat cerebellum.

OBJECTIVE: The role of leptin receptors (Ob-Rs) within the hypothalamus in the control of energy expenditure has well been established. However, their role and regulation in other brain areas, including the cerebellum, is largely unexplored. In the present study we examined whether Ob-R levels in the rat cerebellum are influenced by a high-fat diet and if these changes are sexually divergent during adolescence. METHODS: The fat diet (45% energy from fat) was applied from weaning to puberty (postnatal days [P] 22-42), from weaning to adulthood (P22-90), and from puberty to adulthood (P42-90) in female and male Wistar rats. Ob-R levels were detected by western blotting and the data from pubertal and adult rats were analyzed by two-way analysis of variance for the effects of diet and sex. RESULTS: The fat diet affected Ob-R long isoform levels in a sexually dimorphic manner. In the cerebellum of all fat-fed male groups, Ob-R levels were reduced compared with their chow-fed counterparts (P < 0.05). In contrast, in female rat Ob-R levels were reduced only in the adult P22-90 group. CONCLUSION: Our data show for the first time that Ob-R levels in the rat cerebellum are subject to diet-induced alterations and that these changes are sexually dimorphic.

Degree of oxidation of low density lipoprotein affects expression of CD36 and PPARgamma, but not cytokine production, by human monocyte-macrophages.

Oxidized low-density lipoprotein (oxLDL) exhibits many atherogenic effects, including the promotion of monocyte recruitment to the arterial endothelium and the induction of scavenger receptor expression. However, while atherosclerosis involves chronic inflammation within the arterial intima, it is unclear whether oxLDL alone provides a direct inflammatory stimulus for monocyte-macrophages. Furthermore, oxLDL is not a single, well-defined entity, but has structural and physical properties which vary according to the degree of oxidation. We tested the hypothesis that the biological effects of oxLDL will vary according to its degree of oxidation and that some species of oxLDL will have atherogenic properties, while other species may be responsible for its inflammatory activity. The atherogenic and inflammatory properties of LDL oxidized to predetermined degrees (mild, moderate and extensive oxidation) were investigated in a single system using human monocyte-derived macrophages. Expression of CD36 mRNA was up-regulated by mildly- and moderately-oxLDL, but not highly-oxLDL. The expression of the transcription factor, proliferator-activated receptor-gamma (PPARgamma), which has been proposed to positively regulate the expression of CD36, was increased to the greatest degree by highly-oxLDL. However, the DNA binding activity of PPARgamma was increased only by mildly- and moderately-oxLDL. None of the oxLDL species appeared to be pro-inflammatory towards monocytes, either directly or indirectly through mediators derived from lymphocytes, regardless of the degree of oxidation.