EXAMPLES OF OUR RESEARCH PROJECTS

THE KUOPIO ISCAEMIC HEART DISEASE RISK FACTOR (KIHD) STUDY

The Kuopio Ischaemic Heart Disease Risk Factor (KIHD) Study is an ongoing prospective population-based cohort study designed to investigate risk factors for CVD, atherosclerosis and related outcomes in middle-aged men from eastern Finland, the population with one of the highest recorded rates of CHD. The study protocol was approved by the Research Ethics Committee of the University of Kuopio. All subjects gave informed consent.

The study population is a random sample of men living in the Kuopio city and neighbouring rural communities, stratified and balanced into four strata: 42, 48, 54, or 60 years at the baseline examination. A total of 2682 participants (82.9 % those eligible), were enrolled in the study between 1984 and 1989.

The four-year examinations for the KIHD study were carried out during 1991 to 1993 for 1038 men. Eleven-year examinations were carried out in 1998 to 1999 for men and women. Eighteen-year examinations started in the year 2006.

KIHD Study timeline in PDF

Dietary intake of nutrients was assessed quantitatively with a four day food recording at the KIHD baseline and 11-year examinations. Dietary intake assessment.

Up-to-date there are about 200 published scientific articles from KIHD Study.

More information:

Virtanen JK, Voutilainen S, Rissanen TH, Happonen P, Mursu J, Laukkanen JA, Poulsen H, Lakka TA, Salonen JT. High dietary methionine intake increases the risk of acute coronary events in middle-aged men. Nutr Metab Cardiovasc Dis 2006:16:113-120. PDF

Jyrki K. Virtanen, Sari Voutilainen, Pertti Happonen, Georg Alfthan, Jari Kaikkonen, Jaakko Mursu, Tiina H. Rissanen, George A. Kaplan, Maarit J. Korhonen, Juhani Sivenius, Jukka T. Salonen. Serum homocysteine, folate and risk of stroke. European Journal of Cardiovascular Prevention & Rehabilitation 2005;12:369-375.

Rissanen TH, Voutilainen S, Virtanen JK, Venho B, Vanharanta M, Mursu J, Salonen JT. Intake of fruits, berries and vegetables and mortality: the Kuopio Ischaemic Heart Disease Risk Factor (KIHD) Study. Journal of Nutrition 2003;133:199-204. PDF

Vanharanta M, Voutilainen S, Rissanen TH, Adlercreutz H, Salonen JT. Risk of Cardiovascular and All-Cause Death According to Serum Concentrations of Enterolactone: Kuopio Ischaemic Heart Disease Risk Factor Study. Archives of Internal Medicine 2003;163:1099-1104.

Rissanen T, Voutilainen S, Nyyssönen K, Salonen R, Kaplan GA, Salonen JT. Serum lycopene level and carotid atherosclerosis: the Kuopio Ischaemic Heart Disease Risk Factor (KIHD) Study. American Journal of Clinical Nutrition 2003;77:133-138. PDF

Voutilainen S, Rissanen T, Virtanen J, Lakka TA, Salonen JTS. Low folate intakes are associated with an excess risk of acute coronary events: the Kuopio Ischaemic Heart Disease Risk Factor Study. Circulation 2001;103:2674-2680. PDF

Our aim was to investigate the bioavailability of coffee phenols and the effects of filtered coffee consumption on serum lipid peroxidation, the activity of antioxidant enzymes and plasma tHcy concentration.

Forty-five (45) non-smoking voluntary men aged mean 26 years were recruited from the Kuopio area in eastern Finland through advertisement through e-mail and intranet of the University of Kuopio. Potential participants were screened in an interview for the following inclusion criteria: no severe obesity (BMI <32 kg/m2), no regular use of any drugs or supplement with antioxidative or lipid lowering properties, no chronic diseases like diabetes, CHD, claudication, cerebrovascular disease, hypothyreoidism or other major illness, willingness to abstain from coffee drinking or consume 3 or 6 cups of coffee for three weeks. From these subjects 35 men participated also short-terms study where they consumed 0, 1 or 2 cups of coffee. Volunteers were given freedom to participate to one or both of the studies. All criteria were ascertained prior to entering the study. A written informed consent was obtained from all participants. The study protocol was approved by the joint ethical committee for human research at the University of Kuopio and the Kuopio University Hospital.

Long-term study
Two-week wash-out period preceded the long-term study and during this period the use of coffee, tea, red wine, cocoa and chocolate were forbidden. Subjects were given caffeine tablets to be used if necessary to deal with withdraw symptoms. The maximum daily amount of caffeine was the amount comparable which would be obtained from the daily study bolus in the long-term study (0, 300 or 600 mg). After wash-out period subjects were free to choose whether they wanted to consume 0, 3 or 6 cups of coffee during the 3 wk clinical supplementation period. The study was not randomized to ensure compliance. Blood samples were drawn with Venoject vacuum tubes (Terumo) after overnight fast (10 hours).

Subjects collected 24-h urine excretion before study visits and were also advised to avoid the use of alcohol and analgesics three days and vigorous physical activity one day before the study visits. A four-day food recording was required before and during the last week of the intervention period to control for possible confounding factors and to check the compliance to given instructions. Food records were checked by a nutritionist together with the subjects and then analyzed by using the Nutrica software (version 2.5).

The coffee used in this study was normal filtered coffee packed in 500 g package. Subjects were instructed to measure the daily amount of coffee (teaspoon per one 1.5 dl cup) and to consume the daily amount in three portions.

Short-term study
The short term study was conducted directly after end-point measurements of long-term study. Subjects consumed 1/3 of the study bolus consumed in the long-term study (0, 1 or 2 cups), but otherwise continued to fast and blood sample was taken 1.5 hours after coffee ingestion. From the blood samples the following markers of lipid peroxidation were analyzed; formation of baseline conjugated dienes, hydroxy fatty acids and plasma F2-isoprostanes.

More information:
jaakko.mursu@uku.fi, sari.voutilainen@uku.fi

Jaakko Mursu, Sari Voutilainen, Tarja Nurmi, Georg Alfthan, Jyrki K Virtanen, Tiina H Rissanen, , Pertti Happonen, Kristiina Nyyssönen, Jari Kaikkonen, Riitta Salonen, Jukka T. Salonen. The effect of coffee drinking on plasma total homocysteine concentration and oxidation of serum lipids, a clinical trial. Free Radicals in Biology and Medicine 2005;38:527-34. PDF

Joulukuu 2005, Pari kolme kahvikupillista päivässä tekee hyvää, Pertti kertoo kahvin terveysvaikutuksista Hyvä Terveys -lehdessä. sivu 1 sivu 2 sivu 3

To address the question of the antioxidant function of catechins on total serum and isolated lipoproteins, we conducted a long-term placebo-controlled supplementation study in which phloem was used as the source of catechins.

Seventy-five non-smoking male volunteers aged 31-70 years were recruited from the Kuopio area in eastern Finland through newspaper advertisements. Potential participants were screened in an initial telephone interview for the following inclusion criteria by a public health nurse: 1) no severe obesity (body mass index, BMI < 32 kg/m2), 2) elevated serum cholesterol concentration (total cholesterol 6-9 mmol/L), 3) no regular use of any drug or supplement with antioxidative (ß-carotene, vitamins C or E) or lipid lowering properties 4) no chronic diseases like diabetes, CHD or other major illness 5) willingness to consume 70 g of dried rye bread per day for four weeks. All criteria were ascertained prior to entering the study by a physician. A written informed consent was obtained from all participants. The study protocol was approved by the joint ethical committee for human research at the University of Kuopio and the Kuopio University Hospital.

The study was a 4 wk randomized double-blind supplementation study. Subjects were randomly assigned to consume daily 70 grams of normal dried rye bread (placebo group, n = 30), rye bread in which 7% of the rye flour was substituted with phloem powder (low catechin, LC group, n = 30) or bread in which 14% of the rye flour was substituted with phloem powder (high catechin, HC group, n = 15). The placebo group received 0.6 mg, LC group 17.7 mg and HC group 35.8 mg of catechins daily from the study bread. The subjects were advised to discontinue the use of tea, red wine, cacao and chocolate one week prior to the study and to avoid the use of alcohol and analgesics three days before and vigorous physical activity one day before the study visits. A four-day food recording was required before and during the last week of the intervention period to control for possible confounding factors and to check the compliance to given instructions. The compliance was also checked by a questionnaire designed to assess the amount of breads eaten. Blood samples were drawn after overnight fast (10 hours). All measurement were done at baseline and after the 4 wk supplementation period.

In the phloem study the lipid peroxidation was evaluated by measuring the oxidation susceptibility of serum and LDL+VLDL to oxidation. Briefly, the resistance of serum lipids to oxidation was measured after the serum was diluted to a concentration of 0.67% in 0.02 mol/L phosphate buffered saline (PBS), pH 7.4. Oxidation was initiated by addition of 100 l of 1 mmol/L CuCl2 into 2 mL of diluted, prewarmed (30 C) serum.

The formation of conjugated dienes was followed by monitoring the change in 234 nm absorbance at 30 C on a spectrophotometer equipped with a six position automatic sample changer. The time required from the start to the maximal rate of the reaction (lag time) was determined.

We also analyzed the lignan content of phloem powder enriched rye bread and studied the dose-response relationship of the effect of dietary plant lignans derived from phloem on enterolactone production by measuring enterolactone concentration in serum. We found a significant increase in serum enterolactone concentration in the LP and HP groups compared with the placebo group (P=0.009 and P=0.003, respectively). Considerable interindividual differences were observed in the response to dietary lignans within the study groups. Our results indicated that plant lignans attached to insoluble fiber layer in phloem can be further metabolized and converted to enterolactone presumably by the bacteria present in the colon.

More information:
jaakko.mursu@uku.fi

Mursu J, Vanharanta M, Voutilainen S, Rissanen TH, Nurmi T, Porkkala-Sarataho E, Nyyssönen K, Virtanen JK, Salonen R, Salonen JT. Polyphenol-rich phloem enhances the resistance of total serum lipids to oxidation in men. Journal of Agriculture and Food Chemistry 2005;53:3017-22. PDF

Mursu J, Nurmi T, Vanharanta M, Voutilainen S, Salonen JT. Developing phytochemical products: a case study. Phytochemical Functional Foods, Edited by Ian Johnson and Gary Williamson, Cambridge, England, 2003. ISBN 0-8493-1754-1. 384 p.

Vanharanta M, Mursu J, Nurmi T, Voutilainen S, Rissanen TH, Salonen R, Adlercreutz H, Salonen JT. Phloem fortification in rye bread elevates serum enterolactone level. European Journal of Clinical Nutrition 2002;56:952-957. PDF

Yle:n Kotimaan kasvot -ohjelma 19.1.2004 "Palaako Pettu ruokapöytään". Ohjelma katsottavissa Real Audiona aukeavan sivun oikeasta alareunasta.



CHOCOLATE STUDY (2001)

Cocoa powder is rich in polyphenols and, thus, may contribute to the reduction of lipid peroxidation. Our aim was to study the effects of long-term ingestion of chocolate, with differing amounts of polyphenols, on serum lipids and lipid peroxidation ex vivo and in vivo. We conducted a 3 week clinical supplementation trial of 45 nonsmoking, healthy volunteers. Participants consumed 75 g daily of either white chocolate (white chocolate, WC group), dark chocolate (dark chocolate, DC group), or dark chocolate enriched with cocoa polyphenols (high-polyphenol chocolate, HPC group). In the DC and HPC groups, an increase in serum HDL cholesterol was observed (11.4% and 13.7%, respectively), whereas in the WC group there was a small decrease (-2.9%, p < 0.001). The concentration of serum LDL diene conjugates, a marker of lipid peroxidation in vivo, decreased 11.9% in all three study groups. No changes were seen in the total antioxidant capacity of plasma, in the oxidation susceptibility of serum lipids or VLDL + LDL, or in the concentration of plasma F2-isoprostanes or hydroxy fatty acids. Cocoa polyphenols may increase the concentration of HDL cholesterol, whereas chocolate fatty acids may modify the fatty acid composition of LDL and make it more resistant to oxidative damage.

More information:
jaakko.mursu@uku.fi

Jaakko Mursu, Sari Voutilainen, Tiina H Rissanen, Jyrki K Virtanen, Tarja Nurmi, Jari Kaikkonen, Kristiina Nyyssönen, Jukka T. Salonen. The effect of white chocolate, dark chocolate and cocoa polyphenol enriched chocolate on serum lipids and lipid oxidation. Free Radicals in Biology and medicine 2004;37:1351-59. PDF and a summary in Finnish.

23.9.2004, juttu Suklaatutkimuksen tuloksista YLE:n terveyssivuilla.

a href="http://www.kepka.org/eurolive">The Eurolive Study is aimed at assessing the beneficial effects of olive oil on human health. It is focused on examining the importance of the phenolic compounds of olive oil on oxidative stress and oxidative damage in humans and aimed at providing information to the European Consumers, and to the Olive Oil Industry, about the cost/benefit ratio of the different olive oils on the market.

Olive oils with low, medium and high phenolic content were administered to 30 healthy volunteers in six European centers (Spain, Italy, Germany-Potsdam, Germany-Berlin, Denmark and Finland) in three week periods with two week washout periods between different olive oils.

Three main points were examined:

1. The role of olive oil and olive oil phenolic compounds in the protection of oxidative stress.

2. Oxidative stress occurs after meals with fat. This is called "postprandial oxidative stress". The beneficial effect of olive oil and that of its phenolic compounds on the postprandial oxidative stress were examined.

3. Whether the substitution of other raw fats by olive oil results in healthier fatty acid composition of the LDL.

One important point of the study is that the olive oil administered dose was close to that used daily in Mediterranean countries (25 mL/day).

Expected Results - Benefits
Conclusions concerning the long-term benefits of virgin olive oil and common olive oil consumption will be derived. Conclusions will be available for Consumers, as well as for the Scientific Community, and the Olive Oil Industry. Consumers will be aware of whether or not the content of the phenolic compounds in olive oil protects against oxidative stress. This will give options to consumers in making choices taking into account the nutritional value of each type of olive oil and its price. this study also examines whether or not the consumption of olive oil has the same effects on oxidative damage across Europe. If the beneficial effects of olive oil on oxidative damage differ from country to country, consumers with major benefits should be aware in order to make choices about their nutritional habits. This European study will provide information to the Olive Oil Industry about the health characteristics of each type of olive oil and its cost/benefit ratio.

More information:
jaakko.mursu@uku.fi, jyrki.virtanen@uku.fi

ASAP is a 6-year randomized trial to study the effect of supplementation with vitamin E plus slow-release vitamin C on carotid atherosclerotic progression. The main purpose of the ASAP (Antioxidant Supplementation in Atherosclerosis Prevention) study was to test the effect of reasonable supplemented doses of vitamin E and vitamin C and their combination on the progression of common carotid atherosclerosis in middle-aged high-risk men and women. Subjects were not entered into the trial if they had: regular intake of antioxidants, acetosalicylate or any other drug with antioxidative properties, severe obesity (BMI >32 kg/m2), type 1 diabetes, uncontrolled hypertension (sitting diastolic BP >105 mmHg), any condition limiting mobility, or severe disease shortening life expectancy. Premenopausal women and those taking oral estrogen therapy were also excluded.

The study consisted of 8-week dietary counseling and placebo lead-in phase, a 3-year double-masked treatment period and a 3-year open treatment period. After the lead-in the subjects were randomly allocated to receive twice daily with meal either (1) 91 mg of d-alfa-tocopherol (corresponding to 100 mg of d-alfa-tocopheryl acetate and 136 IU of vitamin E), (2) 250 mg slow-release ascorbic acid, (3) both d-alfa-tocopherol and slow-release ascorbic acid in a single tablet (CellaVie®), or (4) placebo only. All tablets were identical in appearance, size and color.

After the double-blind 3-year period, the study was continued for another three years as an open study, during which period all the supplemented subjects received the vitamin combination and the placebo subjects continued without supplementation. The doses were chosen to keep the plasma ratio of vitamin C and E concentrations similar as in unsupplemented persons. The pilot studies established that a reasonably constant plasma level of vitamin C was achieved. The subjects were randomized separately in four strata of approximately equal size: (1) smoking (>5 cigarettes/day) men, (2) nonsmoking men, (3) smoking postmenopausal women, and (4) nonsmoking postmenopausal women. All participants signed a written informed consent. The study protocol was approved by the Research Ethics Committee of the University of Kuopio.

The subjects came to baseline visits and were randomized during 1994-95. Follow-up visits were 6, 12, 18, 24, 30, 36 and 72 months later. Supplements were given, returned tablets were counted and ultrasonographic assessment of common carotid artery (CCA) intima-media thickness (IMT) was carried out at all these eight visits. Of the 520 hypercholesterolemic men and women aged 45-69 years, 440 (84.6%) completed the study.

Dietary intake of nutrients was assessed quantitatively with a four-day food recording at the ASAP study baseline and 3-year examinations.

More information:

Voutilainen S, Morrow J, Roberts J, Alfthan G, Nyyssönen K, Salonen J. Correlation between Plasma Total Homocysteine Concentration and Plasma F2-Isoprostane in 100 men in Eastern Finland. Arteriosclerosis, Thrombosis and Vascular Biology 1999;19:1263-1266. PDF

Rissanen T, Voutilainen S, Nyyssönen K, Salonen R, Salonen JT. Low plasma lycopene concentration is associated with increased intima-media thickness of the carotid artery wall. Arteriosclerosis, Thrombosis and Vascular Biology 2000;20:2677-2681. PDF

Vanharanta M, Voutilainen S, Nurmi T, Kaikkonen J, Roberts JL, Morrow JD, Adlercreutz H, Salonen JT. Association between low serum enterolactone and increased plasma F2-isoprostanes, a measure of lipid peroxidation. Atherosclerosis 2002;160:465-469. PDF

Salonen R, Nyyssönen K, Kaikkonen J, Porkkala-Sarataho E, Voutilainen S, Rissanen T, Tuomainen T-P, Valkonen V-P, Ristonmaa U, Lakka H, Vanharanta M, Salonen JT, Poulsen H. Six-year effect of vitamin E and vitamin C supplementation on atherosclerosis progression: the Antioxidant Supplementation in Atherosclerosis Prevention (ASAP) study. Circulation 2003;107:947-953. PDF

Salonen JT, Nyyssönen K, Salonen R, Kaikkonen J, Porkkala-Sarataho E, Voutilainen S, Lakka TA, Rissanen T, Lakka H, Leskinen L, Tuomainen T-P, Valkonen V-P, Poulsen H. Kolmevuotisen C- ja E-vitamiinilisän vaikutus kaulavaltimon ateroskleroosiin. Suomen Lääkärilehti 2001;26:395-400. PDF



FOLIC ACID SUPPLEMENTATION STUDY(1999)

The folic acid supplementation study was a double blind placebo-controlled randomized trial. Healthy nonsmoking male subjects were recruited among the staff and students at the University of Kuopio. Exclusion criteria included regular intake of drugs, vitamins or antioxidants, obesity (BMI > 30 kg/m2), and severe diseases. Forty men were randomized to receive either placebo (microcrystalline cellulose) or 300 g folic acid (Foliren®) three times a day for 12 wk. All the subjects provided a written informed consent. The Research Ethics Committee of the University of Kuopio approved the study protocol.

This study was done and samples analyzed in co-operation with Department of Clinical Chemistry, Laboratory of Atherosclerosis Genetics, Tampere University Hospital, and University of Tampere, Medical School, Tampere, Finland (Professor Terho Lehtimäki and researchers Riikka Rontu and Päivi Hämelahti). Oy Verman Ab (www.verman.fi) delivered supplements and partial funded this study.

More information:
sari.voutilainen@uku.fi

Voutilainen S, Rissanen T, Seppänen K, Porkkala-Sarataho E, Kaikkonen J, Tuomainen T-P, Virtanen J, Lehtimäki T, Malin R, Penttilä I, Kaplan GA, Salonen JT. Folic acid increases serum paraoxonase activity: evidence from a double blind oral supplementation trial in men. Current Topics in Nutraceutical Research 2003;1:175-182.

For further information, please contact Sari Voutilainen (sari.voutilainen@uku.fi)