Unsalted tomato juice intake improves blood pressure and serum low‐density lipoprotein cholesterol level in local Japanese residents at risk of cardiovascular disease

Abstract The aim of this study was to investigate the effects of unsalted tomato juice intake on cardiovascular risk markers in local Japanese residents. Four hundred and eighty‐one local residents in Kuriyama, Japan, were enrolled in this study. Throughout the year of the study, they were provided with as much unsalted tomato juice as they wanted. Participants were screened for cardiovascular risk markers, such as blood pressure (BP), serum lipid profile, and glucose tolerance, before and after the study period. Of the study participants, 260 participated in a detailed study of their lifestyle factors. The average ages of the 184 male and 297 female participants were 56.3 ± 13.3 (mean ± SD) and 58.4 ± 11.7 years, respectively. BP in 94 participants with untreated prehypertension or hypertension was significantly lowered (systolic BP, 141.2 ± 12.1–137.0 ± 16.3 mmHg, p = 0.003; diastolic BP, 83.3 ± 10.1–80.9 ± 11.1 mmHg, p = 0.012, paired t test). Further, the serum low‐density lipoprotein cholesterol (LDL‐C) level in 125 participants with untreated dyslipidemia significantly decreased (155.0 ± 23.2–149.9 ± 25.0 mg/dl, p = 0.005, paired t test). These beneficial effects were not different between sexes and among the different age groups. No significant difference in lifestyle was found before and after the study. Unsalted tomato juice intake improved systolic and diastolic BP and serum LDL‐C level in local Japanese residents at risk of cardiovascular conditions.

149.9 ± 25.0 mg/dl, p = 0.005, paired t test). These beneficial effects were not different between sexes and among the different age groups. No significant difference in lifestyle was found before and after the study. Unsalted tomato juice intake improved systolic and diastolic BP and serum LDL-C level in local Japanese residents at risk of cardiovascular conditions.
Furthermore, 13-oxo-9, 11-octadecadienoic acid (13-oxo-ODA), a conjugated linoleic acid newly identified in tomato juice, was shown to have antidyslipidemic effects (Kim et al., 2012). Recently, we reported that unsalted tomato juice intake for 8 weeks improved hypertriglyceridemia in middle-aged Japanese women (Hirose et al., 2015), which prompted us to investigate the effects of tomato juice on cardiovascular risk markers, such as BP, and lipid and glucose metabolism, in local Japanese male and female residents over a wider age range in this study.

| Study population
The participants of the present study were recruited from among the local residents in Kuriyama, Hokkaido, Japan. We sent flyers of this study to the local residents aged 20-74 years and held several briefing sessions to the candidates to recruit as many participants as possible. The participants received as much unsalted tomato juice (Nippon Del Monte) as they wanted throughout the year of the study. The nutritional composition of the tomato juice used in the present study, which was the same product as that was used in the previous study, is shown in Table 1 (Hirose et al., 2015). Participants kept records of daily tomato juice consumption, as well as any medical treatment and changes therein. We collected the participants' diaries every 3 months and calculated overall tomato juice consumption and total days of tomato juice intake throughout the study period based on the diaries, and then calculated the frequency of tomato juice intake and the average tomato juice consumption per day across the entire cohort. Five hundred and forty-one residents, who accounted for 4.3% of the population of Kuriyama, were enrolled, and 481 (88.9%) completed this study ( Figure 1).
The study protocol was reviewed and approved by the Tokyo Medical and Dental University Review Board, and written informed consent was obtained from all participants. The study was conducted in accordance with the Declaration of Helsinki.

| Measurement
The participants underwent an annual medical checkup, which was conducted according to the public policy of the Japanese government to prevent lifestyle-related diseases, before and after the study were diagnosed with type 2 diabetes, and those with an FPG level 110-125 mg/dl and/or HbA1c 6.0%-6.4% were considered to have impaired glucose tolerance.

| Lifestyle factors
Of the 481 study participants, only 260 (54.1%) who underwent the medical checkup performed by the local government of Kuriyama participated in a detailed study of their lifestyle factors. This involved completing a questionnaire defined by the Japanese Ministry of Health, Labor and Welfare, before and after the intervention (Table 2). The questionnaire was composed of 22 questions, including past medical history, concurrent therapeutic agents, and lifestyle factors, such as diet, exercise, and smoking. Ten of these questions, namely those about exercise, dietary habits, change in body weight, sleeping, and smoking, were assessed with regard to possible lifestyle alterations.

| Statistical analyses
Statistical analyses were performed with GraphPad Prism version 5.0 (GraphPad Software). Paired t test was used to compare the results of physical and blood examination before and after the intervention, and differences between men and women were examined by unpaired t test. Changes in CVD risk markers among different age groups were compared with one-way analysis of variance (ANOVA), and changes in lifestyle were evaluated by chi-square test. p values <0.05 were considered statistically significant.

| Participants and tomato juice consumption
Four hundred and eighty-one residents were enrolled, including 184 men and 297 women, aged 21-74 years. The average age (years) of men and women was 56.3 ± 13.3 (mean ± SD) and 58.4 ± 11.7, respectively. Among all participants, the frequency of daily unsalted tomato juice intake throughout the study period was 92.0 ± 12.2%, and the average daily tomato juice consumption was 215 ± 84 ml.

Most of the study participants consumed about 1 bottle (200 ml)
of unsalted tomato juice every day ( Figure 2). Nine participants who consumed <100 ml/day tomato juice were excluded from further analysis. Table 3 presents the overall results of physical and blood examination in 481 participants before and after the intervention. No significant differences were found in any factor before and after the study period.
F I G U R E 2 Average daily tomato juice consumption and cumulative frequency: The number of participants for each segment of daily tomato juice intake (x-axis) is represented in bars on the lefthand axis. The line and right-hand axis show the cumulative percentage
In 127 participants with untreated dyslipidemia, the mean serum TG and HDL-C level did not change significantly after a year of tomato

| Difference between sexes
Next, we compared the effects of unsalted tomato juice intake on SBP, DBP, and serum LDL-C level between men and women. The

| Difference among age groups
We also compared the changes in SBP, DBP, and the serum level of LDL-C before and after the intervention among different age groups. Have you ever had stroke or have you ever received treatment for stroke?

5
Have you ever had heart disease or have you ever received treatment for heart disease?

6
Have you ever had chronic renal failure or have you ever received treatment for chronic renal failure (dialysis)?

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Have you ever had anemia? 8 Are you a current regular smoker? 9 Have you gained 10 kg or more since you were 20 years old?

10
Have you been exercising at least 2 days per week, at 30 min each at an intensity that causes a slight sweat, for at least 1 year?
11 Do you walk for at least 1 hr every day or have equivalent physical activities in your daily life?

12
Do you walk faster than people of your age and sex?

13
Have you had a weight gain or loss of 3 kg or more over the last year?

14
How fast do you eat compared to others?

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Do you have an evening meal within 2 hr before bedtime 3 days or more per week?
16 Do you eat after the evening meal 3 days or more per week?
17 Do you skip breakfast 3 days or more per week?

18
How often do you drink alcohol?

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How much do you drink a day, in terms of glasses of refined sake?

20
Do you feel refreshed after a night's sleep?

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Are you going to start or have you started lifestyle modifications?

22
Are you willing to get health counseling about lifestyle modifications if the opportunity arises?
Note: Defined by the Japanese Ministry of Health, Labor and Welfare.

| Change in lifestyle factors
Finally, we investigated whether or not the improvements in cardiovascular markers in participants at risk could be attributable to changes in lifestyle during the study period. Among the 260 residents who participated in the detailed lifestyle study, 40 had untreated prehypertension or hypertension, and 69 had untreated dyslipidemia. The lifestyle factors did not differ significantly between the groups either before or after the study period (chi-square test) (Table 4).

| D ISCUSS I ON
The present study showed that unsalted tomato juice intake may have contribution to lower SBP and DBP in local Japanese residents who had untreated prehypertension or hypertension, and improve serum LDL-C level in those who had untreated dyslipidemia. These ameliorative effects were not different between sexes and different age groups and could not be attributable to the alteration in lifestyle. To the best of our knowledge, the current study is the first to investigate the effects of tomato or tomato product intake on CVD risk markers over the course of a year and over a wide age range.
Tomatoes contain a variety of bioactive components that make them and their products, including tomato juice, beneficial for health (Engelhard et al., 2006;Hsu et al., 2008;Oshima et al., 1996;Paran et al., 2009;Ried & Falker, 2011;Sesso et al., 2012). Above all, lycopene is well known for its strong antioxidant activity and the inhibition of LDL oxidation, which plays a key role in the initiation and development of atherosclerosis. Several epidemiological studies have suggested that lycopene could contribute to the prevention TA B L E 3 Results of physical and blood examination in 481 participants before and after the study period Note: Data are presented as the mean and SD. There were no significant differences in any factor before and after the intervention.
F I G U R E 3 Systolic and diastolic blood pressure, and the serum LDL-C level before and after the intervention: (a) systolic and (b) diastolic blood pressure in 94 participants with untreated prehypertension or hypertension; (c) the serum levels of LDL-C in 125 participants with untreated dyslipidemia. Data are presented as the standard error of the mean. *p < 0.05, ** < 0.01, versus before the intervention, paired t test of atherosclerosis and CVDs (Hak et al., 2004;Klipstein-Grobusch et al., 2000;Kohlmeier et al., 1997;Rissanen et al., 2003). shown (Palozza et al., 2010;. Studies suggested that lycopene could reduce the production of proinflammatory cytokines through activation of peroxisome proliferator-activated receptor γ and inhibition of nuclear factor-κB. Furthermore, the beneficial effects on BP (Engelhard et al., 2006;Palozza, Simone, Catalano, Parrone, et al., 2011;Paran et al., 2009)  Note: Data show the change of lifestyle factors in 40 participants with untreated prehypertension or hypertension and in 69 with untreated dyslipidemia before and after the study period. No significant differences in lifestyle factors were found between the groups (chi-square test).

TA B L E 4 Change in lifestyle factors
lycopene have been reported (Gajendragadkar et al., 2014;Kim, Paik, et al., 2011;Zhu, Wang, & Xu, 2011). Although the mechanism is yet to be elucidated, our study showed the beneficial effects of unsalted tomato juice intake on SBP, DBP, and LDL-C; thus, our findings may support these reports of the beneficial properties of lycopene.
Esculeoside A, a tomato saponin, has also been suggested to have protective effects on dyslipidemia and atherosclerosis development by inhibiting ACAT (Fujiwara et al., 2007;Nohara, Ono, Ikeda, Fujiwara, & El-Asar, 2010). protein E-deficient mice (Fujiwara et al., 2007). Esculeoside A could have played a role in improving serum LDL-C level in our study participants although its amount in the tomato juice used in the current study was unknown.
Peroxisome proliferator-activated receptors (PPARs), of which three subtypes have been identified (α, β, γ), regulate energy homeostasis, including lipid and glucose metabolism (Braissant, Foufelle, Scotto, Dauca, & Wahli, 1996;Varga, Czimmerer, & Nagy, 2011;Wahli, Braissant, & Desvergne, 1995). PPARα is strongly expressed in tissues with a high mitochondrial and peroxisomal β-oxidation activity, such as the heart, liver, kidney, and intestine (Braissant et al., 1996), and activation of PPARα results in enhancement of fatty acid oxidation and improvement of dyslipidemia (Wahli et al., 1995). Oxo-octadecadienoic acid (oxo-ODA) found in tomato was recently reported to act as a PPARα agonist Kim et al., 2012;Takahashi et al., 2011). Although four structural isomers of oxo-ODA were identified that activate PPARα equally (Braissant et al., 1996;Kim et al., 2012), 9-oxo-10(E),12(E)-ODA and 13-oxo-ODA were the only ones detected in tomato juice . Kim et al. (2012) showed that 9oxo-10(E),12(E)-ODA decreased cellular accumulation of TG in mice hepatocytes, and 13-oxo-ODA reduced plasma and hepatic TG level and plasma glucose level in obese diabetic mice. PPARα  (Deplanque et al., 2003). Furthermore, the atherosclerosis protective effects of PPARα activation are also thought to occur through enhancement of anti-inflammatory response (Cao, Wen, & Li, 2014). The regulation of energy metabolism, oxidative stress, and the inflammatory response by activation of PPARα is expected to lead to the prevention of atherosclerosis and CVDs. In our study, oxo-ODAs in tomato juice could have contributed to the ameliorative effects on the serum LDL-C level and blood pressure.
The present study has some limitations. Firstly, as most of the study participants consumed about one bottle of tomato juice (200 ml) every day, we could not evaluate whether the effects of tomato juice on CVD risk markers depended on the level of consumption. Secondly, the detailed information of the diet each participant followed during the study period is lacking. We also did not assess the intake of other juices and nutritional supplements. Finally, the study on lifestyle factors was conducted in only about half of the participants and the findings may not be applicable to the whole study population.
In conclusion, our study shows that unsalted tomato juice intake could have improved systolic and diastolic blood pressure in Japanese residents who had untreated prehypertension or hypertension, and also decreased the serum LDL-C level in those who had untreated dyslipidemia. As tomato juice is an affordable and readily product, it could be practical as applied a nutritional intervention to prevent CVDs in people at risk.

ACK N OWLED G M ENTS
We appreciate the contribution of Ms. Mayumi Sogawa of the local government of Kuriyama in recruiting the study participants and collecting the data. Authors are thankful to Kikkoman Corporation for financial support.

MT received an unrestricted research grant from Kikkoman
Corporation.

E TH I C A L R E V I E W
This study was conducted in accordance with the Declaration of Helsinki. This study protocol was reviewed and approved by the Tokyo Medical and Dental University Review Board.

I N FO R M E D CO N S E NT
Written informed consent was obtained from all study participants.