Determinación y caracterización de esteroles no colesterol en tejido adiposo

En el mundo occidental, la obesidad, conocida como un peso corporal desproporcionado con respecto a la estatura, se ha convertido en la epidemia del siglo XXI. Como consecuencia del aumento calórico excesivo, se producen una serie de cambios metabólicos en los diferentes territorios adiposos, que podrían actuar como biomarcadores de sobrepeso u obesidad. El objetivo principal de este estudio fue optimizar un método analítico para determinar colesterol y oxiesteroles en tejido adiposo por HPLC-MS/MS para estudiar posteriormente estos biomarcadores en mujeres con sobrepeso y obesidad. Se determinó colesterol y oxiesteroles en tejido adiposo control y tejido adiposo subcutáneo y visceral de 38 mujeres mayores de edad con índices de masa corporal (IMC) comprendidos entre 23 y 40,1 kg/m2. Se obtuvieron unas concentraciones de colesterol y oxiesteroles en tejido adiposo subcutáneo mayores en mujeres con IMC≤31 kg/m2 que en mujeres con IMC>31 kg/m2. Sin embargo, en tejido adiposo visceral las mujeres con IMC>31 kg/m2 tuvieron concentraciones mayores de colesterol y oxiesteroles. En tejido adiposo subcutáneo se encontró una relación inversa entre el colesterol y el IMC, al contrario que en tejido adiposo visceral donde la relación fue directa. Hubo una relación directa entre el colesterol y los oxiesteroles tanto en tejido adiposo subcutáneo como en visceral, siendo estadísticamente significativa con el 27-hidroxicolesterol y con el 24S-hidroxicolesterol, respectivamente. En conclusión, el tejido adiposo subcutáneo y el visceral parecen comportarse de manera diferente a la hora de acumular colesterol y oxiesteroles en función del IMC en las mujeres estudiadas. Los resultados obtenidos tienen una significación estadística muy baja en algunos casos, por lo que sería de interés continuar este trabajo con un mayor tamaño muestral. Futuros estudios de los oxiesteroles en suero y en tejido adiposo, podrían ofrecer un análisis más global del metabolismo del colesterol en la obesidad.<br /

Genetic predictors of weight loss in overweight and obese subjects

The aim of our study was to investigate a large cohort of overweight subjects consuming a homogeneous diet to identify the genetic factors associated with weight loss that could be used as predictive markers in weight loss interventions. We retrospectively recruited subjects (N = 788) aged over 18 years with a Body Mass Index (BMI) between 25 and 40 kg/m(2) who were treated at our lipid unit for at least one year from 2008 to 2016, and we also recruited a control group (168 patients) with normal BMIs. All participants received counselling from a nutritionist that included healthy diet and physical activity recommendations. We genotyped 25 single nucleotide variants (SNVs) in 25 genes that were previously associated with obesity and calculated genetic scores that were derived from 25 SNVs. The risk allele in CADM2 showed a higher frequency in overweight and obese subjects than in controls (p = 0.007). The mean follow-up duration was 5.58 +/- 2.68 years. Subjects with lower genetic scores showed greater weight loss during the follow-up period. The genetic score was the variable that best explained the variations in weight from the baseline. The genetic score explained 2.4% of weight change variance at one year and 1.6% of weight change variance at the end of the follow-up period after adjusting for baseline weight, sex, age and years of follow-up

Effect of bergamot on lipid profile in humans: A systematic review

Dyslipidemia is a well-established modifiable cardiovascular risk. Although statins can reduce LDLc by 50-60%, less than 20% of patients with high risk of CVD achieve LDL targets. The aim of this systematic review is to evaluate the effect of the nutraceutical, bergamot (Citrus bergamia), on lipid parameters in humans. PubMed, Embase, Cochrane Library, and Google Scholar databases were searched for interventional and observational studies investigating the effect of bergamot on lipid profile in humans. This systematic review retrieved a total of 442 studies of which 12 articles fulfilled the eligibility criteria and were included in the qualitative synthesis. Based on data, 75% of studies showed a significant decrease in total cholesterol, triglycerides and LDLc. The decrease in total cholesterol varied from 12.3% to 31.3%, from 7.6% to 40.8% in LDLc and from 11.5% to 39.5% in triglycerides. Eight trials reported HDLc increase after intervention with bergamot. Overall, a dose-dependent and possible synergistic effect when administering with statins can be deducted from these trials. It is essential to point out that studies had heterogeneous designs and scientific quality of studies was quite limited. Promising findings reveal an alternative therapeutic option in dyslipidemia management with bergamot supplementation, especially in subjects with statins intolerance

ABCG5/G8 gene is associated with hypercholesterolemias without mutation in candidate genes and non-cholesterol sterols

Context Approximately 20% to 40% of clinically defined familial hypercholesterolemia (FH) cases do not show a causative mutation in candidate genes (mutation-negative FH), and some of them may have a polygenic origin. Objective The aim of this work was to study the prevalence of ABCG5/G8 genetic variants in mutation-negative FH, as defects in these genes relate to intestinal hyperabsorption of cholesterol and thus ABCG5/G8 variants could explain in part the mechanism of hypercholesterolemia. Design, setting, and patients We sequenced the ABCG5/G8 genes in 214 mutation-negative FH and 97 controls. Surrogate markers of cholesterol absorption (5?-cholestanol, ?-sitosterol, campesterol, stigmasterol, and sitostanol) were quantified by high-performance liquid chromatography?tandem mass spectrometry in both studied groups. Results We found 8 mutation-negative FH patients (3.73%) with a pathogenic mutation in ABCG5/G8 genes. We observed significantly higher concentration of surrogate markers of cholesterol absorption in mutation-negative FH than in controls. In addition, we found significantly higher concentrations of cholesterol absorption markers in mutation-negative FH with ABCG5/G8 defects than in mutation-negative, ABCG5/G8-negative FH. A gene score reflecting the number of common single nucleotide variants associated with hypercholesterolemia was significantly higher in cases than in controls (P = .032). Subjects with a gene score above the mean had significantly higher 5?-cholestanol and stigmasterol than those with a lower gene score. Conclusions Mutation-negative FH subjects accumulate an excess of rare and common gene variations in ABCG5/G8 genes. This variation is associated with increased intestinal absorption of cholesterol, as determined by surrogate makers, suggesting that these loci contribute to hypercholesterolemia by enhancing intestinal cholesterol absorption.This study was supported by grants from the Spanish Ministry of Economy and Competitiveness PI15/01983, PI13/02507, PI12/01321, CIBERCV, CIBEROBN, and Cuenca Villoro Foundation. These projects are co-financed by Instituto de Salud Carlos III and the European Regional Development Fund (ERDF) of the European Union “A way to make Europe.

STARD1 promotes NASH-driven HCC by sustaining the generation of bile acids through the alternative mitochondrial pathway

Background & Aims Besides their physiological role in bile formation and fat digestion, bile acids (BAs) synthesised from cholesterol in hepatocytes act as signalling molecules that modulate hepatocellular carcinoma (HCC). Trafficking of cholesterol to mitochondria through steroidogenic acute regulatory protein 1 (STARD1) is the rate-limiting step in the alternative pathway of BA generation, the physiological relevance of which is not well understood. Moreover, the specific contribution of the STARD1-dependent BA synthesis pathway to HCC has not been previously explored. Methods STARD1 expression was analyzed in a cohort of human non-alcoholic steatohepatitis (NASH)-derived HCC specimens. Experimental NASH-driven HCC models included MUP-uPA mice fed a high-fat high-cholesterol (HFHC) diet and diethylnitrosamine (DEN) treatment in wild-type (WT) mice fed a HFHC diet. Molecular species of BAs and oxysterols were analyzed by mass spectrometry. Effects of NASH-derived BA profiles were investigated in tumour-initiated stem-like cells (TICs) and primary mouse hepatocytes (PMHs). Results Patients with NASH-associated HCC exhibited increased hepatic expression of STARD1 and an enhanced BA pool. Using NASH-driven HCC models, STARD1 overexpression in WT mice increased liver tumour multiplicity, whereas hepatocyte-specific STARD1 deletion (Stard1ΔHep) in WT or MUP-uPA mice reduced tumour burden. These findings mirrored the levels of unconjugated primary BAs, β-muricholic acid and cholic acid, and their tauroconjugates in STARD1-overexpressing and Stard1ΔHep mice. Incubation of TICs or PMHs with a mix of BAs mimicking this profile stimulated expression of genes involved in pluripotency, stemness and inflammation. Conclusions The study reveals a previously unrecognised role of STARD1 in HCC pathogenesis, wherein it promotes the synthesis of primary BAs through the mitochondrial pathway, the products of which act in TICs to stimulate self-renewal, stemness and inflammation. Lay summary Effective therapy for hepatocellular carcinoma (HCC) is limited because of our incomplete understanding of its pathogenesis. The contribution of the alternative pathway of bile acid (BA) synthesis to HCC development is unknown. We uncover a key role for steroidogenic acute regulatory protein 1 (STARD1) in non-alcoholic steatohepatitis-driven HCC, wherein it stimulates the generation of BAs in the mitochondrial acidic pathway, the products of which stimulate hepatocyte pluripotency and self-renewal, as well as inflammation.We acknowledge support from grants PID2019-111669RB-100, SAF2017-85877R and SAF2015-73579-JIN from Plan Nacional de I+D funded by the Agencia Estatal de Investigación (AEI), the Fondo Europeo de Desarrollo Regional (FEDER) and CIBEREHD; the center grant P50AA011999 Southern California Research Center for ALPD and Cirrhosis funded by NIAAA / NIH; as well as support from AGAUR of the Generalitat de Catalunya SGR-2017-1112, European Cooperation in Science & Technology (COST) ACTION CA17112 Prospective European Drug-Induced Liver Injury Network, the ‘ER stress-mitochondrial cholesterol axis in obesity-associated insulin resistance and comorbidities’-Ayudas FUNDACION BBVA and the Red Nacional 2018-102799-T de Enfermedades Metabólicas y Cáncer, and Project 201916/31 "Contribution of mitochondrial oxysterol and bile acid metabolism to liver carcinogenesis" 2019 by Fundació Marato TV3. We also acknowledge the support from the Fondo de Investigaciones Sanitarias, Instituto de Salud Carlos III, Spain (PI16/00598, co-funded by European Regional Development Fund / European Social Fund, ‘Investing in your future’) and Centro Internacional sobre el Envejecimiento (OLD-HEPAMARKER, 0348_CIE_6_E), Spain. We also acknowledge support from R01 CA2344128 and U01 AA022614 grants to M.K.Peer reviewe

ApoA-I Mimetic administration, but not increased apoA-I-containing HDL, inhibits tumour growth in a mouse model of inherited breast cancer.

Low levels of high-density lipoprotein cholesterol (HDLc) have been associated with breast cancer risk, but several epidemiologic studies have reported contradictory results with regard to the relationship between apolipoprotein (apo) A-I and breast cancer. We aimed to determine the effects of human apoA-I overexpression and administration of specific apoA-I mimetic peptide (D-4F) on tumour progression by using mammary tumour virus-polyoma middle T-antigen transgenic (PyMT) mice as a model of inherited breast cancer. Expression of human apoA-I in the mice did not affect tumour onset and growth in PyMT transgenic mice, despite an increase in the HDLc level. In contrast, D-4F treatment significantly increased tumour latency and inhibited the development of tumours. The effects of D-4F on tumour development were independent of 27-hydroxycholesterol. However, D-4F treatment reduced the plasma oxidized low-density lipoprotein (oxLDL) levels in mice and prevented oxLDL-mediated proliferative response in human breast adenocarcinoma MCF-7 cells. In conclusion, our study shows that D-4F, but not apoA-I-containing HDL, hinders tumour growth in mice with inherited breast cancer in association with a higher protection against LDL oxidative modificatio

ApoA-I mimetic administration, but not increased apoA-I-containing HDL, inhibits tumour growth in a mouse model of inherited breast cancer

Low levels of high-density lipoprotein cholesterol (HDLc) have been associated with breast cancer risk, but several epidemiologic studies have reported contradictory results with regard to the relationship between apolipoprotein (apo) A-I and breast cancer. We aimed to determine the effects of human apoA-I overexpression and administration of specific apoA-I mimetic peptide (D-4F) on tumour progression by using mammary tumour virus-polyoma middle T-antigen transgenic (PyMT) mice as a model of inherited breast cancer. Expression of human apoA-I in the mice did not affect tumour onset and growth in PyMT transgenic mice, despite an increase in the HDLc level. In contrast, D-4F treatment significantly increased tumour latency and inhibited the development of tumours. The effects of D-4F on tumour development were independent of 27-hydroxycholesterol. However, D-4F treatment reduced the plasma oxidized low-density lipoprotein (oxLDL) levels in mice and prevented oxLDL-mediated proliferative response in human breast adenocarcinoma MCF-7 cells. In conclusion, our study shows that D-4F, but not apoA-I-containing HDL, hinders tumour growth in mice with inherited breast cancer in association with a higher protection against LDL oxidative modification

International descriptive and interventional survey for oxycholesterol determination by gas- and liquid-chromatographic methods

Increasing numbers of laboratories develop new methods based on gas-liquid and high-performance liquid chromatography to determine serum concentrations of oxygenated cholesterol metabolites such as 7α-, 24(S)-, and 27-hydroxycholesterol. We initiated a first international descriptive oxycholesterol (OCS) survey in 2013 and a second interventional survey 2014 in order to compare levels of OCS reported by different laboratories and to define possible sources of analytical errors. In 2013 a set of two lyophilized serum pools (A and B) was sent to nine laboratories in different countries for OCS measurement utilizing their own standard stock solutions. In 2014 eleven laboratories were requested to determine OCS concentrations in lyophilized pooled sera (C and D) utilizing the same provided standard stock solutions of OCS. The participating laboratories submitted results obtained after capillary gas-liquid chromatography-mass selective detection with either epicoprostanol or deuterium labelled sterols as internal standards and high-performance liquid chromatography with mass selective detection and deuterated OCS as internal standard. Each participant received a clear overview of the results in form of Youden-Plots and basic statistical evaluation in its used unit. The coefficients of variation of the concentrations obtained by all laboratories using their individual methods were 58.5–73.3% (survey 1), 56.8–60.3% (survey 2); 36.2–35.8% (survey 1), 56.6–59.8, (survey 2); 61.1–197.7% (survey 1), 47.2–74.2% (survey 2) for 24(S)-, 27-, and 7α-hydroxycholesterol, respectively. We are surprised by the very great differences between the laboratories, even under conditions when the same standards were used. The values of OCS's must be evaluated in relation to the analytical technique used, the efficiency of the ample separation and the nature of the internal standard used. Quantification of the calibration solution and inappropriate internal standards could be identified as major causes for the high variance in the reported results from the different laboratories. A harmonisation of analytical standard methods is highly needed

First international descriptive and interventional survey for cholesterol and non-cholesterol sterol determination by gas- and liquid- chromatography–Urgent need for harmonisation of analytical methods

Serum concentrations of lathosterol, the plant sterols campesterol and sitosterol and the cholesterol metabolite 5α-cholestanol are widely used as surrogate markers of cholesterol synthesis and absorption, respectively. Increasing numbers of laboratories utilize a broad spectrum of well-established and recently developed methods for the determination of cholesterol and non-cholesterol sterols (NCS). In order to evaluate the quality of these measurements and to identify possible sources of analytical errors our group initiated the first international survey for cholesterol and NCS. The cholesterol and NCS survey was structured as a two-part survey which took place in the years 2013 and 2014. The first survey part was designed as descriptive, providing information about the variation of reported results from different laboratories. A set of two lyophilized pooled sera (A and B) was sent to twenty laboratories specialized in chromatographic lipid analysis. The different sterols were quantified either by gas chromatography-flame ionization detection, gas chromatography- or liquid chromatography-mass selective detection. The participants were requested to determine cholesterol and NCS concentrations in the provided samples as part of their normal laboratory routine. The second part was designed as interventional survey. Twenty-two laboratories agreed to participate and received again two different lyophilized pooled sera (C and D). In contrast to the first international survey, each participant received standard stock solutions with defined concentrations of cholesterol and NCS. The participants were requested to use diluted calibration solutions from the provided standard stock solutions for quantification of cholesterol and NCS. In both surveys, each laboratory used its own internal standard (5α-cholestane, epicoprostanol or deuterium labelled sterols). Main outcome of the survey was, that unacceptably high interlaboratory variations for cholesterol and NCS concentrations are reported, even when the individual laboratories used the same calibration material. We discuss different sources of errors and recommend all laboratories analysing cholesterol and NCS to participate in regular quality control programs