Zara Pons Vila | Universitat Rovira i Virgili (original) (raw)

Papers by Zara Pons Vila

Arteriosclerosis, Thrombosis, and Vascular Biology, 2022

Background: Understanding the processes behind carotid plaque instability is necessary to develop... more Background: Understanding the processes behind carotid plaque instability is necessary to develop methods for identification of patients and lesions with stroke risk. Here, we investigated molecular signatures in human plaques stratified by echogenicity as assessed by duplex ultrasound. Methods: Lesion echogenicity was correlated to microarray gene expression profiles from carotid endarterectomies (n=96). The findings were extended into studies of human and mouse atherosclerotic lesions in situ, followed by functional investigations in vitro in human carotid smooth muscle cells (SMCs). Results: Pathway analyses highlighted muscle differentiation, iron homeostasis, calcification, matrix organization, cell survival balance, and BCLAF1 (BCL2 [B-cell lymphoma 2]-associated transcription factor 1) as the most significant signatures. BCLAF1 was downregulated in echolucent plaques, positively correlated to proliferation and negatively to apoptosis. By immunohistochemistry, BCLAF1 was found...

Els polifenols presents a la dieta s’han estudiat extensament pel seus efectes positius sobre dif... more Els polifenols presents a la dieta s’han estudiat extensament pel seus efectes positius sobre diferents components de la si?ndrome metabo?lica (MetS). En aquesta Tesi, es va proposar avaluar els efecte beneficio?s dels polifenols de pinyol de rai?m sobre la disfuncio? endotelial i la hipertensio? (HTN) associada a la MetS, utilitzant el model de dieta de cafeteria com a model de MetS. Els resultats mostraren que una dosi aguda de 375 mg/Kg d’un extracte de polifenols de baix pes molecular de pinyol de rai?m (LM-GSPE) 6 hores post- administracio? va ser la me?s efectiva per a baixar la pressio? arterial en aquest model de HTN indui?t per dieta. En questa millora va ser veure la implicacio? de la Sirtuina-1 (Sirt-1) aixi? com bloqueig del Sistema Renina-Angiotensina-Aldosterona (SRAA), que com a consequ?e?ncia va millorar la funcio? endotelial. Per una altra banda, es va veure com alguns dels polifenols me?s abundants del LM-GSPE, com l’(-)-epicatequina, la (+)-catequina o l’a?cid ga?...

Metabolic syndrome (MS) is becoming pandemic in developed countries. Industrialized countries hav... more Metabolic syndrome (MS) is becoming pandemic in developed countries. Industrialized countries have the greatest prevalence of MS. Even thought there are genetic factors, the unhealthy lifestyle is the major contributor. Cafeteria-fed rats are commonly used as a model for MS, because it mimics all the symptoms of MS and shares the pathogenesis of the disease. No general pharmacological treatment for the whole pathology exists. Angiotensin converting enzyme (ACE) inhibitors have been widely used for years to control blood pressure of hypertensive patients, but more research is needed to elucidate other beneficial effects. In this study 20 wistar rats were fed for 12 weeks with cafeteria diet and treated daily with vehicle (CAF) or 50 mg/kg captopril (C-CAF), an ACE inhibitor. A standard (ST) fed group and treated with vehicle also was added to the experiment. Measurements of blood pressure, body weight, and waist perimeter were recorded weekly. Plasma cholesterol and triglycerides wer...

La enfermedad cardiovascular es la principal causa de muerte en el mundo y la hipertensión arteri... more La enfermedad cardiovascular es la principal causa de muerte en el mundo y la hipertensión arterial es uno de sus principales factores de riesgo. Varios estudios han demostrado que el consumo de alimentos o ingredientes ricos en flavanoles, como el cacao o extracto de pepita de uva, mejoran la función endotelial y disminuyen la presión arterial. Diferentes mecanismos podrían justificar las propiedades antihipertensivas de los flavanoles. La vasodilatación ocasionada por estos compuestos se ha relacionado con la reducción del estrés oxidativo, con la producción de óxido nítrico (NO) y la inhibición de la enzima convertidora de la angiotensina (ECA), clave para el control de la presión arterial. El cacao y el extracto de pepita de uva son productos ricos en flavanoles del tipo flavan-3-ol y proantocianidinas que han demostrado gran actividad antioxidante. Sin embargo, la concentración real de flavonoides que alcanzan los vasos sanguineos es tan baja que es poco probable que estos comp...

Metabolic syndrome (MS) is a combination of medical disorders such as hypertension, dyslipidaemia... more Metabolic syndrome (MS) is a combination of medical disorders such as hypertension, dyslipidaemia and obesity that, when occurring together, increase the risk of developing cardiovascular and diabetes. Our group, have demonstrated that a grape seed procyanidin extract (GSPE) exhibit different beneficial properties on various parameters related to MS. The aim of this study was to evaluate the effects of GSPE in blood pressure (BP) and to study the potential antihypertensive mechanisms involved. Male spontaneously hypertensive rats (SHR) (17-20-week-old) were administered by gastric intubation water or 250, 375 and 500 mg/Kg GSPE (minimum n=8 per group). Captopril (50 mg/Kg), a known antihypertensive drug, was used as a positive control (n=8). Systolic and diastolic blood pressure (SBP and DBP) were recorded in the rats by the tail cuff method initially and 2, 4, 6, 8, 24, and 48 h post-administration. To determine the mechanisms involved, 2 groups of 15 animals were administered by i...

The Journal of Nutritional Biochemistry, 2015

Physiological concentrations (1 μM) of 15 flavonoids were evaluated in human umbilical vein endot... more Physiological concentrations (1 μM) of 15 flavonoids were evaluated in human umbilical vein endothelial cells in the presence of hydrogen peroxide (H(2)O(2)) for their ability to affect endothelial nitric oxide synthase (eNOS) and endothelin-1 (ET-1) expression in order to establish the structural basis of their bioactivity. Flavonoid effects on eNOS transcription factor Krüpple like factor-2 (KLF-2) expression were also evaluated. All studied flavonoids appeared to be effective compounds for counteracting the oxidative stress-induced effects on vascular gene expression, indicating that flavonoids are an excellent source of functional endothelial regulator products. Notably, the more effective flavonoids for KLF-2 up-regulation resulted in the highest values for eNOS expression, showing that the increment of eNOS expression would take place through KLF-2 induction. Structure-activity relationship studies showed that the combinations of substructures on flavonoid skeleton that regulate eNOS expression are made up of the following elements: glycosylation and hydroxylation of C-ring, double bond C2C3 at C-ring, methoxylation and hydroxylation of B-ring, ketone group in C4 at C-ring and glycosylation in C7 of A-ring, while flavonoid features involved in the reduction of vasoconstrictor ET-1 expression are as follows: double bond C2C3 at C-ring glycosylation in C7 of A-ring and ketone group in C4 of C-ring.

Journal of Functional Foods, 2015

Flavan-3-ols and their oligomeric forms, proanthocyanidins (PAs), are poorly absorbed in the smal... more Flavan-3-ols and their oligomeric forms, proanthocyanidins (PAs), are poorly absorbed in the small intestine and reach the colon where gut bacteria enzymes can hydrolyse them to produce small molecular metabolites, which can reach systemic circulation. However, the microbial metabolism of flavanols has been poorly described in vivo. The aim of this study was to determine the colonic biotransformation pathway and the plasma temporal appearance of grape seed flavanols colonic metabolites in rats. Rat plasma colonic metabolites were analysed by HPLC-MS/MS at 2, 7, 24 and 48 h after 1000 mg/kg of a grape seed PA extract (GSPE) administration. Results indicated that non-metabolised flavanols have peak plasma concentrations 2 h after GSPE administration, whereas the colonic metabolites appeared in plasma later, indicating their gradual colonic biotransformation as valerolactone > phenylpropionic acids ≈ phenylacetic acids > benzoic acids. This study shows how flavanols are biotransformed by gut bacteria in rats over time, facilitating potential bioactive compound identification for particular health effects.

The Journal of Nutritional Biochemistry, 2015

Flavanols are metabolized in the small intestine and the liver to produce their glucuronidated, s... more Flavanols are metabolized in the small intestine and the liver to produce their glucuronidated, sulfated or methylated conjugates that can be body-distributed or excreted in the urine. However, the intake of large amounts of flavanols is not directly related to their bioavailability. This study aims to investigate the administered dose dependence of flavanols' conjugation and body distribution. In this study different doses of a grape seed proanthocyanidin extract (GSPE; 125, 250, 375 and 1000mg/kg) were orally administered to male Wistar rats. Tissues were collected 2h after GSPE administration. Flavanols were quantified by HPLC-MS/MS. Results show that the majority of GSPE metabolites are located in the kidney, followed by the liver. Lower concentrations were found in mesenteric white adipose tissue (MWAT) and the brain. Moreover, flavanol metabolites followed a tissue-specific distribution pattern independent of dosage. In the kidney, glucuronidated metabolites were the most abundant; however, in the liver were mainly methyl-glucuronidated metabolites. In MWAT, free flavanols were dominant, and methylated metabolites were dominant in the brain. Concentration within a tissue was dependent on the administered dose. In conclusion, flavanol metabolites follow a tissue-specific distribution pattern and only the tissue concentration of flavanol metabolites is dependent on the administered dose.

Journal of Agricultural and Food Chemistry, 2014

This study describes the development and validation of a liquid chromatography−mass spectrometry ... more This study describes the development and validation of a liquid chromatography−mass spectrometry method for determination of a large number of flavanol colonic derivatives in biological samples. The method was validated with rat plasma after the intake of grape seed flavanols. The minimum plasma volume necessary to maintain good recovery values within the range of 83−110% for all of the standards was determined by micro solid-phase extraction (μ-SPE). In total, 16 commercial standards were used to measure 30 different phenolic compounds present at low concentration levels (micromolar). The chromatographic method enabled reliable quantification of plasma colonic flavanol derivatives with low limits of detection and quantification, achieving values of 0.03 nM and 0.10 nM, respectively. The developed method can be readily applied to determine all of the flavanol metabolites that are most likely responsible for the majority of biological effects of poorly absorbed flavanols.

Journal of Functional Foods

Abstract Involvement of nitric oxide (NO) in the antihypertensive effect of grape seed flavanols ... more Abstract Involvement of nitric oxide (NO) in the antihypertensive effect of grape seed flavanols was previously demonstrated. This study investigates the endothelial vasoprotective pattern and the role of Sirtuin-1 (Sirt-1) in this antihypertensive effect. Cafeteria diet-fed hypertensive rats (CHR) were administered water or 375 mg/kg of a low-molecular grape seed polyphenol extract (LM-GSPE) rich in flavanols and sacrificed 6 h post-administration. Plasma endothelin-1 (ET-1) and aortic expression of NO pathway genes, NADPH oxidase subunit 4 (NOX4) and ET-1 genes were measured. LM-GPSE decreased plasma ET-1, upregulating eNOS and Sirt-1 and downregulating aortic gene expression of ET-1 and NADPH, the endothelial major producer of free radicals, indicating the vasoprotective effect of grape seed flavanols. Additionally, twenty-four spontaneously hypertensive rats (SHR) were administered water or 375 mg/kg LM-GSPE and treated with 1 mg/kg sirtinol (inhibitor of Sirt-1). The antihypertensive effect of LM-GSPE was completely abolished by sirtinol, indicating that grape seed flavanols decrease BP in a Sirt-1-dependent manner.

Scientific Reports

Circulating microparticles (MPs) are procoagulant due to the surface containing phosphatidylserin... more Circulating microparticles (MPs) are procoagulant due to the surface containing phosphatidylserine (PS), which facilitates coagulation. We investigated if MPs improve hemostasis in HA plasma models. MPs isolated from pooled normal human plasma were added to severe, moderate and mild HA plasma models (0%, 2.5%, 20% FVIII). The MPs’ effect on hemostasis was evaluated by calibrated automated thrombogram (CAT) and overall hemostasis potential (OHP) assays, while fibrin structure was imaged by standard confocal, stimulated emission depletion (STED) microscopy and scanning electron microscopy (SEM). MPs partially restored thrombin generation and fibrin formation in all HA plasma models. The procoagulant effect of MPs requires PS exposure, to a less extent of contact pathway activation, but not tissue factor exposure or in vitro stimulation of MPs. MPs partially normalized the fibrin structure, and using super-resolution STED, MPs attached to fibrin were clearly resolved. In summary, our r...

Nutrients

The antihypertensive properties of different doses of a chicken foot hydrolysate, Hpp11 and the m... more The antihypertensive properties of different doses of a chicken foot hydrolysate, Hpp11 and the mechanisms involved in this effect were investigated. Spontaneously hypertensive rats (SHR) were administered water, Captopril (50 mg/kg) or Hpp11 at different doses (25, 55 and 85 mg/kg), and the systolic blood pressure (SBP) was recorded. The SBP of normotensive Wistar-Kyoto (WKY) rats administered water or Hpp11 was also recorded. Additionally, plasmatic angiotensin-converting enzyme (ACE) activity was determined in the SHR administered Hpp11. Moreover, the relaxation caused by Hpp11 in isolated aortic rings from Sprague-Dawley rats was evaluated. Hpp11 exhibited antihypertensive activity at doses of 55 and 85 mg/kg, with maximum activity 6 h post-administration. At this time, no differences were found between these doses and Captopril. Initial SBP values of 55 and 85 mg/kg were recovered 24 or 8 h post-administration, respectively, 55 mg/kg being the most effective dose. At this dose,...

British Journal of Nutrition

The effects of grape-seed polyphenols against the development of hypertension and other cardiomet... more The effects of grape-seed polyphenols against the development of hypertension and other cardiometabolic conditions associated with the metabolic syndrome (MetS) were studied in rats fed a high-fat, high-carbohydrate diet, known as the cafeteria (CAF) diet. Two groups of Wistar rats were fed standard (STD) or CAF diets for 12 weeks. The CAF diet-fed rats were administered different doses of a low-molecular-weight grape-seed polyphenol extract (LM-GSPE) (25, 100 and 200 mg/kg per d) or vehicle daily, and the STD diet-fed rats were administered LM-GSPE (100 mg/kg per d) or vehicle using ten animals per group. Body weight (BW), waist perimeter (WP) and systolic and diastolic blood pressures (BP) by the tail-cuff method were recorded weekly. The animals were housed in metabolic chambers every 2 weeks to estimate daily food and liquid intakes and to collect faeces and urine samples. The plasma lipid profile was analysed at time 0 and on the 4th, 7th, 10th and 12th weeks of the experiment....

Food Chemistry, 2017

Flavanols, which exert several health benefits, are metabolized after ingestion. Factors such as ... more Flavanols, which exert several health benefits, are metabolized after ingestion. Factors such as the host physiological condition could affect the metabolism and bioavailability of flavanols, influencing their bioactivities. This study aimed to qualitatively evaluate whether a pathological state influenced flavanol plasma bioavailability. Standard and cafeteria (CAF) diet fed rats, a robust model of metabolic syndrome (MeS), were administered 1000mg/kg of flavanol enriched grape seed polyphenol extract (GSPE). Flavanols and their metabolites were quantified by HPLC-MS/MS in plasma before and at 2, 4, 7, 24, and 48h after GSPE ingestion. Results showed that in CAF administered rats the maximum time of plasma flavanol concentration was delayed and these animals presented higher levels of plasma phase-II metabolites as well as altered microbial metabolites. In conclusion, this study demonstrated that MeS pathological state modified flavanol bioavailability, supporting the hypothesis that flavanol metabolism, and therefore flavanol functionality, depend on the organism's state of health.

Molecular Nutrition & Food Research, 2016

Studying the flavanol metabolism is essential to identify bioactive compounds, as beneficial effe... more Studying the flavanol metabolism is essential to identify bioactive compounds, as beneficial effects of flavanols have been attributed to their metabolic products. However, host-related factors, including pathological conditions, may affect flavanol metabolism and, thus, their bioactivity. This study aims to elucidate whether hypertension affects grape seed flavanol metabolism, influencing their bioactivity in relation to hypertension. Grape seed flavanols' effect on blood pressure (BP) was studied in spontaneously hypertensive rats (SHR) and healthy Wistar rats 6 h after grape seed extract administration (375 mg/kg). Animals were then sacrificed, and plasma bioavailability and aorta distribution of flavanol metabolites were studied by HPLC-MS/MS in both the groups. Grape seed flavanols were only able to decrease BP in SHR. Plasma total flavanol metabolites showed similar levels, being the difference noticed in specific metabolites' concentrations. Specifically, microbial metabolites showed quantitative and qualitative differences between both health states. Moreover, aorta total concentrations were found decreased in SHR. Interestingly, flavanol microbial metabolites were specifically increased SHR aortas, showing qualitative differences in small phenolic forms. This study demonstrates important differences in bioactivity and target tissue metabolite levels between healthy and diseased rats, indicating potential metabolites responsible of the anti-hypertensive effect.

diometabolic abnormality was significantly and negatively associated with physical activity level... more diometabolic abnormality was significantly and negatively associated with physical activity level, education level, alcohol intake and olive oil intake and positively associated with BMI. Conclusions: The prevalence of cardiometabolic abnormalities related to obesity is similar to that found in US population. Smoking, physical activity level and alcohol intake contribute to the explanation of the prevalence of metabolic non-healthy obee subjects in the Spanish population, as in the US population. However, in Spain olive oil intake, independently of education level, contributes significantly to the explanation of the variance in the prevalence of metabolic abnormalities.

The Journal of Nutritional Biochemistry, 2013

The regular consumption of flavonoids has been associated with reduced mortality and a decreased ... more The regular consumption of flavonoids has been associated with reduced mortality and a decreased risk of cardiovascular diseases. The proanthocyanidins found in plasma are very different from the original flavonoids in food sources. The use of physiologically appropriate conjugates of proanthocyanidins is essential for the in vitro analysis of flavonoid bioactivity. In this study, the effect of different proanthocyanidin-rich extracts, which were obtained from cocoa (CCX), French maritime pine bark (Pycnogenol extract, PYC) and grape seed (GSPE), on lipid homeostasis was evaluated. Hepatic human cells (HepG2 cells) were treated with 25 mg/L of CCX, PYC or GSPE. We also performed in vitro experiments to assess the effect on lipid synthesis that is induced by the bioactive GSPE proanthocyanidins using the physiological metabolites that are present in the serum of GSPE-administered rats. For this, Wistar rats were administered 1 g/kg of GSPE, and serum was collected after 2 h. The semipurified serum of GSPE-administered rats was fully characterized by liquid chromatography tandem triple quadrupole mass spectrometry (LC-QqQ/MS(2)). The lipids studied in the analyses were free cholesterol (FC), cholesterol ester (CE) and triglycerides (TG). All three proanthocyanidin-rich extracts induced a remarkable decrease in the de novo lipid synthesis in HepG2 cells. Moreover, GSPE rat serum metabolites reduced the total percentage of CE, FC and particularly TG; this reduction was significantly higher than that observed in the cells directly treated with GSPE. In conclusion, the bioactivity of the physiological metabolites that are present in the serum of rats after their ingestion of a proanthocyanidin-rich extract was demonstrated in Hep G2 cells.

Molecular Nutrition & Food Research, 2016

SCOPE Dietary flavanols produce beneficial health effects, and once absorbed, they are recognized... more SCOPE Dietary flavanols produce beneficial health effects, and once absorbed, they are recognized as xenobiotics and undergo phase-II enzymatic detoxification. Flavanols health-promoting properties are mainly attributed to their metabolic products. This work aimed to elucidate whether rats of the opposite sex exhibited differences in the metabolism and distribution of ingested flavanols. METHODS AND RESULTS Acute doses of grape seed polyphenols were administered to male and female rats. After 1, 2 and 4 h, plasma, liver, mesenteric white adipose tissue (MWAT), brain and hypothalamus flavanol metabolites were quantified by HPLC-MS/MS. Results indicated important sex-related quantitative differences in plasma and brain. Moreover, remarkable sex-related differences in the distributions and types of flavanol metabolites were also observed between liver and brain. CONCLUSIONS This study demonstrated that sex differentially influences the metabolism and distribution of flavanols throughout the bodies of rats, which may affect the physiological bioactivities of flavanols between males and females.

Arteriosclerosis, Thrombosis, and Vascular Biology, 2022

Background: Understanding the processes behind carotid plaque instability is necessary to develop... more Background: Understanding the processes behind carotid plaque instability is necessary to develop methods for identification of patients and lesions with stroke risk. Here, we investigated molecular signatures in human plaques stratified by echogenicity as assessed by duplex ultrasound. Methods: Lesion echogenicity was correlated to microarray gene expression profiles from carotid endarterectomies (n=96). The findings were extended into studies of human and mouse atherosclerotic lesions in situ, followed by functional investigations in vitro in human carotid smooth muscle cells (SMCs). Results: Pathway analyses highlighted muscle differentiation, iron homeostasis, calcification, matrix organization, cell survival balance, and BCLAF1 (BCL2 [B-cell lymphoma 2]-associated transcription factor 1) as the most significant signatures. BCLAF1 was downregulated in echolucent plaques, positively correlated to proliferation and negatively to apoptosis. By immunohistochemistry, BCLAF1 was found...

Els polifenols presents a la dieta s’han estudiat extensament pel seus efectes positius sobre dif... more Els polifenols presents a la dieta s’han estudiat extensament pel seus efectes positius sobre diferents components de la si?ndrome metabo?lica (MetS). En aquesta Tesi, es va proposar avaluar els efecte beneficio?s dels polifenols de pinyol de rai?m sobre la disfuncio? endotelial i la hipertensio? (HTN) associada a la MetS, utilitzant el model de dieta de cafeteria com a model de MetS. Els resultats mostraren que una dosi aguda de 375 mg/Kg d’un extracte de polifenols de baix pes molecular de pinyol de rai?m (LM-GSPE) 6 hores post- administracio? va ser la me?s efectiva per a baixar la pressio? arterial en aquest model de HTN indui?t per dieta. En questa millora va ser veure la implicacio? de la Sirtuina-1 (Sirt-1) aixi? com bloqueig del Sistema Renina-Angiotensina-Aldosterona (SRAA), que com a consequ?e?ncia va millorar la funcio? endotelial. Per una altra banda, es va veure com alguns dels polifenols me?s abundants del LM-GSPE, com l’(-)-epicatequina, la (+)-catequina o l’a?cid ga?...

Metabolic syndrome (MS) is becoming pandemic in developed countries. Industrialized countries hav... more Metabolic syndrome (MS) is becoming pandemic in developed countries. Industrialized countries have the greatest prevalence of MS. Even thought there are genetic factors, the unhealthy lifestyle is the major contributor. Cafeteria-fed rats are commonly used as a model for MS, because it mimics all the symptoms of MS and shares the pathogenesis of the disease. No general pharmacological treatment for the whole pathology exists. Angiotensin converting enzyme (ACE) inhibitors have been widely used for years to control blood pressure of hypertensive patients, but more research is needed to elucidate other beneficial effects. In this study 20 wistar rats were fed for 12 weeks with cafeteria diet and treated daily with vehicle (CAF) or 50 mg/kg captopril (C-CAF), an ACE inhibitor. A standard (ST) fed group and treated with vehicle also was added to the experiment. Measurements of blood pressure, body weight, and waist perimeter were recorded weekly. Plasma cholesterol and triglycerides wer...

La enfermedad cardiovascular es la principal causa de muerte en el mundo y la hipertensión arteri... more La enfermedad cardiovascular es la principal causa de muerte en el mundo y la hipertensión arterial es uno de sus principales factores de riesgo. Varios estudios han demostrado que el consumo de alimentos o ingredientes ricos en flavanoles, como el cacao o extracto de pepita de uva, mejoran la función endotelial y disminuyen la presión arterial. Diferentes mecanismos podrían justificar las propiedades antihipertensivas de los flavanoles. La vasodilatación ocasionada por estos compuestos se ha relacionado con la reducción del estrés oxidativo, con la producción de óxido nítrico (NO) y la inhibición de la enzima convertidora de la angiotensina (ECA), clave para el control de la presión arterial. El cacao y el extracto de pepita de uva son productos ricos en flavanoles del tipo flavan-3-ol y proantocianidinas que han demostrado gran actividad antioxidante. Sin embargo, la concentración real de flavonoides que alcanzan los vasos sanguineos es tan baja que es poco probable que estos comp...

Metabolic syndrome (MS) is a combination of medical disorders such as hypertension, dyslipidaemia... more Metabolic syndrome (MS) is a combination of medical disorders such as hypertension, dyslipidaemia and obesity that, when occurring together, increase the risk of developing cardiovascular and diabetes. Our group, have demonstrated that a grape seed procyanidin extract (GSPE) exhibit different beneficial properties on various parameters related to MS. The aim of this study was to evaluate the effects of GSPE in blood pressure (BP) and to study the potential antihypertensive mechanisms involved. Male spontaneously hypertensive rats (SHR) (17-20-week-old) were administered by gastric intubation water or 250, 375 and 500 mg/Kg GSPE (minimum n=8 per group). Captopril (50 mg/Kg), a known antihypertensive drug, was used as a positive control (n=8). Systolic and diastolic blood pressure (SBP and DBP) were recorded in the rats by the tail cuff method initially and 2, 4, 6, 8, 24, and 48 h post-administration. To determine the mechanisms involved, 2 groups of 15 animals were administered by i...

The Journal of Nutritional Biochemistry, 2015

Physiological concentrations (1 μM) of 15 flavonoids were evaluated in human umbilical vein endot... more Physiological concentrations (1 μM) of 15 flavonoids were evaluated in human umbilical vein endothelial cells in the presence of hydrogen peroxide (H(2)O(2)) for their ability to affect endothelial nitric oxide synthase (eNOS) and endothelin-1 (ET-1) expression in order to establish the structural basis of their bioactivity. Flavonoid effects on eNOS transcription factor Krüpple like factor-2 (KLF-2) expression were also evaluated. All studied flavonoids appeared to be effective compounds for counteracting the oxidative stress-induced effects on vascular gene expression, indicating that flavonoids are an excellent source of functional endothelial regulator products. Notably, the more effective flavonoids for KLF-2 up-regulation resulted in the highest values for eNOS expression, showing that the increment of eNOS expression would take place through KLF-2 induction. Structure-activity relationship studies showed that the combinations of substructures on flavonoid skeleton that regulate eNOS expression are made up of the following elements: glycosylation and hydroxylation of C-ring, double bond C2C3 at C-ring, methoxylation and hydroxylation of B-ring, ketone group in C4 at C-ring and glycosylation in C7 of A-ring, while flavonoid features involved in the reduction of vasoconstrictor ET-1 expression are as follows: double bond C2C3 at C-ring glycosylation in C7 of A-ring and ketone group in C4 of C-ring.

Journal of Functional Foods, 2015

Flavan-3-ols and their oligomeric forms, proanthocyanidins (PAs), are poorly absorbed in the smal... more Flavan-3-ols and their oligomeric forms, proanthocyanidins (PAs), are poorly absorbed in the small intestine and reach the colon where gut bacteria enzymes can hydrolyse them to produce small molecular metabolites, which can reach systemic circulation. However, the microbial metabolism of flavanols has been poorly described in vivo. The aim of this study was to determine the colonic biotransformation pathway and the plasma temporal appearance of grape seed flavanols colonic metabolites in rats. Rat plasma colonic metabolites were analysed by HPLC-MS/MS at 2, 7, 24 and 48 h after 1000 mg/kg of a grape seed PA extract (GSPE) administration. Results indicated that non-metabolised flavanols have peak plasma concentrations 2 h after GSPE administration, whereas the colonic metabolites appeared in plasma later, indicating their gradual colonic biotransformation as valerolactone > phenylpropionic acids ≈ phenylacetic acids > benzoic acids. This study shows how flavanols are biotransformed by gut bacteria in rats over time, facilitating potential bioactive compound identification for particular health effects.

The Journal of Nutritional Biochemistry, 2015

Flavanols are metabolized in the small intestine and the liver to produce their glucuronidated, s... more Flavanols are metabolized in the small intestine and the liver to produce their glucuronidated, sulfated or methylated conjugates that can be body-distributed or excreted in the urine. However, the intake of large amounts of flavanols is not directly related to their bioavailability. This study aims to investigate the administered dose dependence of flavanols' conjugation and body distribution. In this study different doses of a grape seed proanthocyanidin extract (GSPE; 125, 250, 375 and 1000mg/kg) were orally administered to male Wistar rats. Tissues were collected 2h after GSPE administration. Flavanols were quantified by HPLC-MS/MS. Results show that the majority of GSPE metabolites are located in the kidney, followed by the liver. Lower concentrations were found in mesenteric white adipose tissue (MWAT) and the brain. Moreover, flavanol metabolites followed a tissue-specific distribution pattern independent of dosage. In the kidney, glucuronidated metabolites were the most abundant; however, in the liver were mainly methyl-glucuronidated metabolites. In MWAT, free flavanols were dominant, and methylated metabolites were dominant in the brain. Concentration within a tissue was dependent on the administered dose. In conclusion, flavanol metabolites follow a tissue-specific distribution pattern and only the tissue concentration of flavanol metabolites is dependent on the administered dose.

Journal of Agricultural and Food Chemistry, 2014

This study describes the development and validation of a liquid chromatography−mass spectrometry ... more This study describes the development and validation of a liquid chromatography−mass spectrometry method for determination of a large number of flavanol colonic derivatives in biological samples. The method was validated with rat plasma after the intake of grape seed flavanols. The minimum plasma volume necessary to maintain good recovery values within the range of 83−110% for all of the standards was determined by micro solid-phase extraction (μ-SPE). In total, 16 commercial standards were used to measure 30 different phenolic compounds present at low concentration levels (micromolar). The chromatographic method enabled reliable quantification of plasma colonic flavanol derivatives with low limits of detection and quantification, achieving values of 0.03 nM and 0.10 nM, respectively. The developed method can be readily applied to determine all of the flavanol metabolites that are most likely responsible for the majority of biological effects of poorly absorbed flavanols.

Journal of Functional Foods

Abstract Involvement of nitric oxide (NO) in the antihypertensive effect of grape seed flavanols ... more Abstract Involvement of nitric oxide (NO) in the antihypertensive effect of grape seed flavanols was previously demonstrated. This study investigates the endothelial vasoprotective pattern and the role of Sirtuin-1 (Sirt-1) in this antihypertensive effect. Cafeteria diet-fed hypertensive rats (CHR) were administered water or 375 mg/kg of a low-molecular grape seed polyphenol extract (LM-GSPE) rich in flavanols and sacrificed 6 h post-administration. Plasma endothelin-1 (ET-1) and aortic expression of NO pathway genes, NADPH oxidase subunit 4 (NOX4) and ET-1 genes were measured. LM-GPSE decreased plasma ET-1, upregulating eNOS and Sirt-1 and downregulating aortic gene expression of ET-1 and NADPH, the endothelial major producer of free radicals, indicating the vasoprotective effect of grape seed flavanols. Additionally, twenty-four spontaneously hypertensive rats (SHR) were administered water or 375 mg/kg LM-GSPE and treated with 1 mg/kg sirtinol (inhibitor of Sirt-1). The antihypertensive effect of LM-GSPE was completely abolished by sirtinol, indicating that grape seed flavanols decrease BP in a Sirt-1-dependent manner.

Scientific Reports

Circulating microparticles (MPs) are procoagulant due to the surface containing phosphatidylserin... more Circulating microparticles (MPs) are procoagulant due to the surface containing phosphatidylserine (PS), which facilitates coagulation. We investigated if MPs improve hemostasis in HA plasma models. MPs isolated from pooled normal human plasma were added to severe, moderate and mild HA plasma models (0%, 2.5%, 20% FVIII). The MPs’ effect on hemostasis was evaluated by calibrated automated thrombogram (CAT) and overall hemostasis potential (OHP) assays, while fibrin structure was imaged by standard confocal, stimulated emission depletion (STED) microscopy and scanning electron microscopy (SEM). MPs partially restored thrombin generation and fibrin formation in all HA plasma models. The procoagulant effect of MPs requires PS exposure, to a less extent of contact pathway activation, but not tissue factor exposure or in vitro stimulation of MPs. MPs partially normalized the fibrin structure, and using super-resolution STED, MPs attached to fibrin were clearly resolved. In summary, our r...

Nutrients

The antihypertensive properties of different doses of a chicken foot hydrolysate, Hpp11 and the m... more The antihypertensive properties of different doses of a chicken foot hydrolysate, Hpp11 and the mechanisms involved in this effect were investigated. Spontaneously hypertensive rats (SHR) were administered water, Captopril (50 mg/kg) or Hpp11 at different doses (25, 55 and 85 mg/kg), and the systolic blood pressure (SBP) was recorded. The SBP of normotensive Wistar-Kyoto (WKY) rats administered water or Hpp11 was also recorded. Additionally, plasmatic angiotensin-converting enzyme (ACE) activity was determined in the SHR administered Hpp11. Moreover, the relaxation caused by Hpp11 in isolated aortic rings from Sprague-Dawley rats was evaluated. Hpp11 exhibited antihypertensive activity at doses of 55 and 85 mg/kg, with maximum activity 6 h post-administration. At this time, no differences were found between these doses and Captopril. Initial SBP values of 55 and 85 mg/kg were recovered 24 or 8 h post-administration, respectively, 55 mg/kg being the most effective dose. At this dose,...

British Journal of Nutrition

The effects of grape-seed polyphenols against the development of hypertension and other cardiomet... more The effects of grape-seed polyphenols against the development of hypertension and other cardiometabolic conditions associated with the metabolic syndrome (MetS) were studied in rats fed a high-fat, high-carbohydrate diet, known as the cafeteria (CAF) diet. Two groups of Wistar rats were fed standard (STD) or CAF diets for 12 weeks. The CAF diet-fed rats were administered different doses of a low-molecular-weight grape-seed polyphenol extract (LM-GSPE) (25, 100 and 200 mg/kg per d) or vehicle daily, and the STD diet-fed rats were administered LM-GSPE (100 mg/kg per d) or vehicle using ten animals per group. Body weight (BW), waist perimeter (WP) and systolic and diastolic blood pressures (BP) by the tail-cuff method were recorded weekly. The animals were housed in metabolic chambers every 2 weeks to estimate daily food and liquid intakes and to collect faeces and urine samples. The plasma lipid profile was analysed at time 0 and on the 4th, 7th, 10th and 12th weeks of the experiment....

Food Chemistry, 2017

Flavanols, which exert several health benefits, are metabolized after ingestion. Factors such as ... more Flavanols, which exert several health benefits, are metabolized after ingestion. Factors such as the host physiological condition could affect the metabolism and bioavailability of flavanols, influencing their bioactivities. This study aimed to qualitatively evaluate whether a pathological state influenced flavanol plasma bioavailability. Standard and cafeteria (CAF) diet fed rats, a robust model of metabolic syndrome (MeS), were administered 1000mg/kg of flavanol enriched grape seed polyphenol extract (GSPE). Flavanols and their metabolites were quantified by HPLC-MS/MS in plasma before and at 2, 4, 7, 24, and 48h after GSPE ingestion. Results showed that in CAF administered rats the maximum time of plasma flavanol concentration was delayed and these animals presented higher levels of plasma phase-II metabolites as well as altered microbial metabolites. In conclusion, this study demonstrated that MeS pathological state modified flavanol bioavailability, supporting the hypothesis that flavanol metabolism, and therefore flavanol functionality, depend on the organism's state of health.

Molecular Nutrition & Food Research, 2016

Studying the flavanol metabolism is essential to identify bioactive compounds, as beneficial effe... more Studying the flavanol metabolism is essential to identify bioactive compounds, as beneficial effects of flavanols have been attributed to their metabolic products. However, host-related factors, including pathological conditions, may affect flavanol metabolism and, thus, their bioactivity. This study aims to elucidate whether hypertension affects grape seed flavanol metabolism, influencing their bioactivity in relation to hypertension. Grape seed flavanols' effect on blood pressure (BP) was studied in spontaneously hypertensive rats (SHR) and healthy Wistar rats 6 h after grape seed extract administration (375 mg/kg). Animals were then sacrificed, and plasma bioavailability and aorta distribution of flavanol metabolites were studied by HPLC-MS/MS in both the groups. Grape seed flavanols were only able to decrease BP in SHR. Plasma total flavanol metabolites showed similar levels, being the difference noticed in specific metabolites' concentrations. Specifically, microbial metabolites showed quantitative and qualitative differences between both health states. Moreover, aorta total concentrations were found decreased in SHR. Interestingly, flavanol microbial metabolites were specifically increased SHR aortas, showing qualitative differences in small phenolic forms. This study demonstrates important differences in bioactivity and target tissue metabolite levels between healthy and diseased rats, indicating potential metabolites responsible of the anti-hypertensive effect.

diometabolic abnormality was significantly and negatively associated with physical activity level... more diometabolic abnormality was significantly and negatively associated with physical activity level, education level, alcohol intake and olive oil intake and positively associated with BMI. Conclusions: The prevalence of cardiometabolic abnormalities related to obesity is similar to that found in US population. Smoking, physical activity level and alcohol intake contribute to the explanation of the prevalence of metabolic non-healthy obee subjects in the Spanish population, as in the US population. However, in Spain olive oil intake, independently of education level, contributes significantly to the explanation of the variance in the prevalence of metabolic abnormalities.

The Journal of Nutritional Biochemistry, 2013

The regular consumption of flavonoids has been associated with reduced mortality and a decreased ... more The regular consumption of flavonoids has been associated with reduced mortality and a decreased risk of cardiovascular diseases. The proanthocyanidins found in plasma are very different from the original flavonoids in food sources. The use of physiologically appropriate conjugates of proanthocyanidins is essential for the in vitro analysis of flavonoid bioactivity. In this study, the effect of different proanthocyanidin-rich extracts, which were obtained from cocoa (CCX), French maritime pine bark (Pycnogenol extract, PYC) and grape seed (GSPE), on lipid homeostasis was evaluated. Hepatic human cells (HepG2 cells) were treated with 25 mg/L of CCX, PYC or GSPE. We also performed in vitro experiments to assess the effect on lipid synthesis that is induced by the bioactive GSPE proanthocyanidins using the physiological metabolites that are present in the serum of GSPE-administered rats. For this, Wistar rats were administered 1 g/kg of GSPE, and serum was collected after 2 h. The semipurified serum of GSPE-administered rats was fully characterized by liquid chromatography tandem triple quadrupole mass spectrometry (LC-QqQ/MS(2)). The lipids studied in the analyses were free cholesterol (FC), cholesterol ester (CE) and triglycerides (TG). All three proanthocyanidin-rich extracts induced a remarkable decrease in the de novo lipid synthesis in HepG2 cells. Moreover, GSPE rat serum metabolites reduced the total percentage of CE, FC and particularly TG; this reduction was significantly higher than that observed in the cells directly treated with GSPE. In conclusion, the bioactivity of the physiological metabolites that are present in the serum of rats after their ingestion of a proanthocyanidin-rich extract was demonstrated in Hep G2 cells.

Molecular Nutrition & Food Research, 2016

SCOPE Dietary flavanols produce beneficial health effects, and once absorbed, they are recognized... more SCOPE Dietary flavanols produce beneficial health effects, and once absorbed, they are recognized as xenobiotics and undergo phase-II enzymatic detoxification. Flavanols health-promoting properties are mainly attributed to their metabolic products. This work aimed to elucidate whether rats of the opposite sex exhibited differences in the metabolism and distribution of ingested flavanols. METHODS AND RESULTS Acute doses of grape seed polyphenols were administered to male and female rats. After 1, 2 and 4 h, plasma, liver, mesenteric white adipose tissue (MWAT), brain and hypothalamus flavanol metabolites were quantified by HPLC-MS/MS. Results indicated important sex-related quantitative differences in plasma and brain. Moreover, remarkable sex-related differences in the distributions and types of flavanol metabolites were also observed between liver and brain. CONCLUSIONS This study demonstrated that sex differentially influences the metabolism and distribution of flavanols throughout the bodies of rats, which may affect the physiological bioactivities of flavanols between males and females.