Olive oil polyphenols decrease blood pressure and improve endothelial function in young women with mild hypertension

Depósito de investigación de la Universidad de Sevilla https://idus.us.es/ “This is a pre-copyedited, author-produced version of an article accepted for publication in AMERICAN JOURNAL OF HYPERTENSION following peer review. The version of record Moreno-Luna R, Muñoz-Hernandez R, Miranda ML, Costa AF, Jimenez-Jimenez L, Vallejo-Vaz AJ, Muriana FJ, Villar J, Stiefel P. Olive oil polyphenols decrease blood pressure and improve endothelial function in young women with mild hypertension. Am J Hypertens. 2012; 25(12): 1299-304 is available online at: https://doi.org/10.1038/ajh.2012.128” ” 1 Olive Oil Polyphenols Decrease Blood Pressure and Improve Endothelial Function in Women with High-Normal Blood Pressure or Stage 1 Essential Hypertension Rafael Moreno-Luna1, Rocio Muñoz-Hernandez1, Maria L. Miranda1, Alzenira F. Costa1, Luis Jimenez-Jimenez2, Antonio J. Vallejo-Vaz1, Francisco J.G. Muriana3, Jose Villar1 and Pablo Stiefel1 1 Unidad Clinico Experimental de Riesgo Vascular (UCAMI-UCERV), Instituto de Biomedicina de Sevilla (IBIS) SAS, CEIC, Universidad de Sevilla, Hospital Virgen del Rocio, Seville, Spain 2 Servicio de Laboratorio Clinico, Hospital Virgen del Rocio, Seville, Spain 3 Laboratory of Cellular and Molecular Nutrition, Instituto de la Grasa, CSIC, Seville, Spain RUNNING TITLE: Olive oil polyphenols and hypertension WORD COUNT: 4016; NUMBER OF FIGURES: 2; NUMBER OF TABLES: 1 SUPPLEMENTAL ONLINE MATERIAL IS NOT SUBMITTED. AUTHORS LIST FOR INDEXING: Moreno-Luna, Muñoz-Jimenez, Miranda, Costa, Jimenez-Jimenez, Vallejo-Vaz, Muriana, Villar, Stiefel 1 Supported by CITOLIVA Foundation, Instituto de Salud Carlos III (RD06/0014/0035), and Junta de Andalucía (CVI-4352) grants. 2 2 Author disclosures: R. Moreno-Luna, R. Muñoz-Jimenez, M.L. Miranda, A.F. Costa, L. Jimenez-Jimenez, A.J. Vallejo-Vaz, F.J.G. Muriana, J. Villar, P. Stiefel have no conflicts of interest. 3Abbreviations used: asymmetric dimethylarginine, ADMA; blood pressure, BP; coronary heart disease, CHD; C-reactive protein, CRP; hyperemic area, HA; ischemic reactive hyperemia, IRH; oxidized LDL, ox-LDL; perfusion units, PU; total cholesterol, TC. Correspondence: Pablo Stiefel, Instituto de Biomedicina de Sevilla (IBIS) SAS, CEIC, Universidad de Sevilla, Hospital Virgen del Rocio, C/ Manuel Siurot s/n, 41013 Seville, Spain. Telephone number: + 34 955013095; Fax number: + 34 955013473; E-mail: [email protected] 3 ABSTRACT 1 This study aims to examine the influence of a polyphenol-rich olive oil on blood 2 pressure (BP) and endothelial function in twenty-four women with high-normal BP or 3 stage 1 essential hypertension. We conducted a double-blind, randomized, crossover 4 dietary-intervention study. After a homogenization period of 4 months (baseline 5 values), two diets were used, one with polyphenol-rich olive oil (Diet A), the other with 6 polyphenol-free olive oil (Diet B). Systolic and diastolic BP, serum or plasma 7 biomarkers of endothelial function, oxidative stress, and inflammation, and ischemia- 8 induced hyperemia in the forearm were measured at baseline and after 2 months on Diet 9 A or Diet B. When compared to baseline values, only Diet A led to a significant (P < 10 0.005) decrease in systolic (–7.91 ± 1.97 mm Hg) and diastolic (–6.65 ± 2.10 mm Hg) BP. 11 A similar trend was found for serum asymmetric dimethylarginine (–0.092 ± 0.001 12 mol/L, P < 0.005), oxidized LDL (–28.23 ± 8.51 g/L, P < 0.005), and plasma C- 13 reactive protein (–1.90 ± 0.52 mg/L, P < 0.005). Diet A also elicited an increase in 14 plasma nitrites/nitrates (+4.69 ± 1.36 mol/L, P < 0.001) and hyperemic area after 15 ischemia (+345.5 ± 78.8 perfusion units/sec, P < 0.001). We concluded that the regular 16 consumption of a diet containing polyphenol-rich olive oil decreases BP and improves 17 endothelial function in women with high-normal BP or stage 1 essential hypertension. 18 19 20 21 22 23 24 25 4 Introduction 26 Endothelial dysfunction is one of the phenomena associated with the development of 27 atherosclerosis and cardiovascular disease in patients with hypertension (1). It is mainly 28 characterized by decreased bioavailability of NO (2) and increased levels of ox-LDL 29 (3). One of the predominant mechanisms in NO inactivation is a perturbation of the L- 30 arginine–NO pathway by oxidative stress leading to elevations of plasma ADMA, 31 which in turn exacerbates oxidative stress (4). Both a pro-oxidant status and increased 32 ADMA are common features of disease states associated with atherosclerosis (4, 5) and 33 hypertension (6). 34 Food intake is an important factor affecting vascular reactivity. Short-term 35 feeding trials have shown the potential for certain foods to improve endothelial 36 function, either as isolated nutrients or as healthy food patterns (7). Several studies 37 suggest that some antioxidant compounds in food may limit oxidative damage and 38 restore endothelial function, thus slowing atherogenic development (8, 9). Polyphenol 39 intake has been associated with low CHD mortality rates (10). Prior studies have 40 reported that antioxidant and anti-inflammatory polyphenols in the diet improve 41 endothelial function and lipid profile (11). The minor components in virgin olive oil, 42 particularly the phenolic compounds, may contribute to the health benefits derived from 43 the Mediterranean Diet. In experimental studies, the phenolic compounds of virgin olive 44 oil showed strong antioxidant properties (12, 13). 45 Although there are a large number of studies linking the consumption of 46 phenolic compounds with a decrease in vascular events, recent evidence suggests that 47 the effects of food vary according to gender (14) and that women are underrepresented 48 in biomedical studies (15). Moreover, most of the studies on polyphenol healthy 49 5 properties are based on high-risk populations, thus underestimating the potential 50 benefits to low-risk populations (16). 51 In this study, we enrolled young women with high-normal BP or stage 1 52 essential hypertension and hypothesized that consumption of polyphenol-rich olive oil 53 would decrease BP and improve endothelial function. Additionally, we examined which 54 variables are most predictive of BP values and endothelial function biomarkers. 55 56 PATIENTS AND METHODS 57 Patients and design. Prior to the study, the Human Investigation Review Committee of 58 the Virgen del Rocio University Hospital approved all protocols, and all participants 59 provided written informed consent. The study was conducted according to the guidelines 60 of good clinical practice and principles expressed in the Helsinki Declaration by the 61 World Medical Association. 62 The study was conducted as double-blind, randomized, crossover. We 63 consecutively asked to enter the study to forty Caucasian women that were newly 64 diagnosed with high-normal BP (a systolic pressure of 120-139 mm Hg or diastolic 65 pressure of 80-89 mm Hg) or stage 1 essential hypertension (a systolic pressure of 140-159 66 mm Hg and diastolic pressure of 90-99 mm Hg). Six women refused to do so, and ten 67 more abandoned after the first dietary intervention. Therefore, twenty-four women 68 completed the study. Their average age was 26 years (range: 24-27 years) and BMI was 69 25.4 kg/m2 (range: 23.5-27.1 kg/m2). They had values of plasma TC between 3.88-4.14 70 mmol/L and TG between 0.51-0.56 mmol/L. All participants completed a comprehensive 71 health-related questionnaire that included lifestyle information (i.e., physical activity, 72 smoking habits, alcohol consumption, tea and coffee, and dietary habits), medical and 73 family history (especially those related to premature cardiovascular disease), and the use of 74 6 medications, nutritional supplements, and vitamins. The exclusion criteria included 75 previous history of cardiovascular disease, any severe chronic illness, treatment with 76 antihypertensive or lipid-lowering drugs, and current smoking habits. 77 Following a homogenization period of 4 months on a healthy diet 78 (homogenization diet), women who met inclusion criteria ate a diet with polyphenol- 79 rich olive oil (Diet A) or a diet with polyphenol-free olive oil (Diet B) for 2-months 80 period and then immediately started the second dietary 2-months period of either the 81 Diet B or the Diet A. A set menu plan, which contained the same basic foods referred 82 for the Mediterranean Diet (17) and calories to the habitual diets, was provided to the 83 participants. They were instructed to avoid the adherence to any special foods or 84 supplements affecting BP, lipid metabolism or highly rich in polyphenols. The only 85 difference between diets (homogenization diet, Diet A, and Diet B) was the type of 86 dietary fat. Sunflower or corn oil was permitted for the homogenization diet, whereas a 87 virgin olive oil rich in polyphenols and the same oil without polyphenols (refined olive 88 oil) were provided for the Diet A and Diet B, respectively. During each dietary 89 intervention period, participants consumed 60 mL of the corresponding dietary fat daily. 90 This study design ensured that each participant consumed comparable energy from dietary 91 fats and amounts of polyphenols from virgin olive oil on the Diet A. To increase 92 compliance, highly motivated women were selected for the study and were given clear 93 written and verbal instructions. The participants understood the importance of adhering to 94 the diet. At the end of the two dietary periods, participants completed a 4-days diet record, 95 which was used to analyze their nutrient intake and compliance. The aim was to minimize 96 metabolic perturbations between test diets focusing on comparison of polyphenol-rich 97 olive oil versus polyphenol-free olive oil. More than 20 different virgin olive oils were 98 analyzed for the polyphenol content, which was measured by reverse-phase HPLC-diode 99 7 array detection as previously described (18). We selected a virgin olive oil containing 564 100 mg/kg polyphenols. A portion of this virgin olive oil underwent oil refining at the Instituto 101 de la Grasa (CSIC, Seville) to produce a polyphenol-free olive oil. 102 Data on brachial BP and IRH, and blood samples were obtained after the 103 homogenization diet (baseline values) and after the Diet A or Diet B. 104 105 BP measurement. Brachial systolic and diastolic BP was measured using an automated 106 oscillometric device (Omron M6 Comfort; Omron Healthcare Co) in the right arm, with 107 participants lying in the supine position for 10 minutes by a trained observer. Three BP 108 readings were taken at 2-minutes interval, and the mean was used for data analysis. 109 110 Assessment of endothelial function, oxidative stress, and inflammation biomarkers. 111 Participants were required to fast and avoid heavy exercise 12 hours prior to the 112 morning of the examination. Plasma and serum were obtained from blood samples, 113 stored at –80 ºC, and analyzed at the Clinic Laboratory Service of University Hospital 114 Virgen del Rocio. Details on analytical methods for each biomarker are provided below. 115 NO via total plasma nitrites/nitrates was measured by a colorimetric assay (NO, 116 Colorimetric Assay; Roche Applied Science) using a modular analyzer Power Wave 117 XS® (Biotek®). Serum endogenous ADMA (ADMA-ELISA; DLD Diagnostika 118 GmbH), ox-LDL (Human Ox-LDL ELISA Kit; Biomedica Medizinprodukte GmbH & 119 Co KG), and plasma high-sensitivity CRP (CRP ELISA Kit; Immundiagnostik AG) 120 were measured with enzyme immunoassays. 121 122 IRH measurement. Laser-Doppler linear Periflux System 5000 (Perimed SA) was used 123 to measure IRH. The participant was taken into a quiet room at our Day Hospital with 124 8 only the researcher and a nurse present. The room temperature was maintained at 22 ºC. 125 The technique and possible symptoms were explained in detail. With the participant in 126 the supine position and after a 15-minutes resting, the BP cuff was placed on the 127 patient’s arm, and the receptor probe was attached on the forearm at 15 cm from the 128 wrist. The BP cuff was then inflated to 40 mm Hg above the systolic BP and maintained 129 at this pressure for 4 minutes. During this period, the monitoring system showed how 130 PU fell steadily to reach the biological zero. Afterwards, the BP cuff was rapidly 131 deflated and how quickly PU rose above the pre-ischemic PU values was monitored. 132 The data were recorded and stored using PeriSoft for Windows. The values of HA after 133 the ischemia were automatically calculated. The same researcher to avoid variability 134 performed all measurements. 135 136 Statistical analysis. We compared the changes after the Diet A or Diet B with respect to 137 the homogenization diet (baseline values) using Student’s t-test for paired samples. The 138 Kolmogorov–Smirnov test was used to assess normality of distributions. When samples 139 were not normally distributed, the Mann–Whitney U test was then used. 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J Am Coll Cardiol. 2005;46:1864-8. 19 TABLE 1 Endothelial function, oxidative stress, and inflammation biomarkers in women with high-normal BP or stage 1 essential hypertension after 4 months on homogenization diet and changes after 2 months on Diet A (polyphenol-rich olive oil) or Diet B (polyphenol-free olive oil)1 Changes from baseline Biomarker Baseline Diet A Diet B Nitrites/nitrates (µmol/L) 19.69 ± 1.01 +4.69 ± 1.36a +0.83 ± 0.85b ADMA (µmol/L) 0.82 ± 0.02 –0.09 ± 0.00a –0.04 ± 0.01b Ox-LDL (µg/L) 153.04 ± 7.12 –28.23 ± 8.51a –6.93 ± 6.82b CRP (mg/L) 1.56 ± 0.47 –1.90 ± 0.52a –0.65 ± 0.56b 1 Data are presented as mean ± SEM, n = 24. Means of change in a row with superscripts without a common letter differ, P < 0.01. Asymmetric dimethylarginine, ADMA; C-reactive protein, CRP; oxidized LDL, ox-LDL. 20 FIGURE LEGENDS FIGURE 1 Changes in systolic (A) and diastolic (B) BP relative to baseline values in women with high-normal BP or stage 1 essential hypertension after 2 months on Diet A (polyphenol-rich olive oil). The line is the best-fit line, n = 24. 21 FIGURE 2 Changes in plasma nitrites/nitrates (A), serum ADMA (B), serum ox-LDL (C), plasma CRP (D), and HA (E) relative to baseline values in women with highnormal BP or stage 1 essential hypertension after 2 months on Diet A (polyphenol-rich olive oil). The line is the best-fit line, n = 24. Asymmetric dimethylarginine, ADMA; C- reactive protein, CRP; hyperemic area, HA; oxidized LDL, ox-LDL. 22