HEALTH AND NUTRITION NEWSNo. 19March 15, 2007

Table of Contents

PREFATORY NOTE
Current issues on nutrition in the clinical field; e.g. lifestyle-related diseases…..…Tsuyoshi Watanabe

CURRENT RESEARCH PROJECTS
Project for Energy Metabolism…………..…..…………..………………………..Kazuko Ishikawa-Takata

THOUGHTS ON HEALTH AND NUTRITION RESEARCH
Traveling the universe of gene……………………………..……….………………….......Kouichi Yamada

RESEARCH FINDINGS
AMP-activated protein kinase regulates PEPCK gene expression by direct phosphorylation of AREBP……...…….…..Erina Inoue, Jun Yamauchi
Young Japanese adults with high BMI tend to underreport energy intake – from the dietary records ofthe general population aged 30’s-60’s……..………………………...…...Hitomi Okubo, Satoshi Sasaki
Effects of age on ventilatory threshold and peak oxygen uptake normalized for regional skeletal
muscle mass: a cross-sectional study on Japanese men and women aged 20-80 years……………..……............ Motohiko Miyachi© National Institute of Health and Nutrition

Prefatory Note

Current issues on nutrition in the clinical field;
e.g. lifestyle-related diseases
Tsuyoshi Watanabe
Professor, Department of Internal Medicine 3 (renal disease,
hypertension, diabetes, internal secretion, metabolic disorder)
School of Medicine, Fukushima Medical University

Now, lifestyle-related diseases are the major social
problems in Japan, which include conventionallycalled as “adult diseases (e,g, diabetes,
hypertension, hyperlipidemia etc.)” and recently
well-advocated metabolic syndrome. The above
diseases would accompany serious complications
such as atherosclerosis, diabetic nephropathy,
diabetic retinopathy, neurosis, nephrosclerosis(caused by hypertension), end-stage renal disease,blindness and intolerable neurologic symptom,
leading to the lowered QOL among the population.
In particular, atherosclerosis is the base of
cardiovascular event, which is one of the top
mortality causes with the overwhelmingly highest
morbidity rate in the world.
Besides, with low economic growth and a fear of
bankruptcy of the public finances for medical
insurance due to the aging society, the proportion of
the expense for lifestyle-related diseases and theircomplications in the national medical expenditures
has been increasing, and now exceed other types ofhealth problems. In this sense, the government
decided to set lifestyle-related diseases control as
the most prioritized national health strategy in the
21st century, where “adult disease” was renamed“lifestyle-related diseases”.
Basically, susceptibility to lifestyle-related diseases
is genetically determined. Studies on the genetic
background have elucidated that diabetes,
hypertension and hyperlipidemia are caused bysingle-gene abnormalities, though they are
exceptional. In fact, most of lifestyle-relateddiseases are polygenic, and the responsible genes
were not selected for eliminationin the process ofhuman evolution (history of hunger), but rather
worked favorably for preservation of species(so-called “thrifty genes”). Consequently, the genetic

background of many Japanese people would
exceed the threshold level for the occurrence oflifestyle-related diseases. In particular, Japanesediet has been composed mainly of plants since thebeginning of human history. It is therefore likely thatthe genetic background with low digestive andabsorptive capacity was selected, in which one
would develop lifestyle-related diseases by highenergy and fat intake. It is also pointed out thatJapanese people have relatively longer small
intestine than meat-eating western people. We
should therefore bear in mind that Japanese is theethnic group who fatefully carry lots of thrifty genes.
Needless to say, a dramatic increase oflifestyle-related diseases is not caused by thechange in genetic factors. After the World War II,
Japan experienced high economic growth andbecame the world's second-largest economy. As aresult, the general people could enjoy rich foodsupply which was once accessible by the royalty
and aristocracy only. In particular, westernization oflifestyle shifted food preference from staple food,vegetable and fish to meat, oil/lipid and milk, leading
to establishment of the westernized gourmet culture.
This trend can be observed also in the annual
National Nutrition Survey (renamed to National
Health and Nutrition Survey in 2003) undertaken by
the Ministry of Health, Labour and Welfare, whichshows that the nutrient intake has been dramaticallychanged with decreased carbohydrate, increased
fat and slightly increased protein intake. It is said
that high fat diet, especially excess intake of animal
fat that contains high saturated fatty acid, could
induce visceral adiposity with enhanced insulin
resistance, which eventually cause lifestyle-relateddiseases. No significant change has been observedin the average energy intake per capita during the

of nutrient food and advancement in medical
science and health system, infectious diseases andundernutrition were controlled and average life
expectancy was prolonged. Consequently, Japan
became the aging society with the world longest
average life expectancy after the WWII. Apart fromthe financial problem on medical insurance, at theindividual level, it is good to secure one’s longevity.
On the other hand, the negative aspect came up at
the individual level as well, that is, lifestyle-relateddiseases. Most of lifestyle-related diseases wereclassified as “adult diseases” in the past, as themorbidity of these diseases increased with age.
Thus, the aging society is one of the factors
responsible to the increase of lifestyle-relateddiseases. In other words, we can regard
lifestyle-related diseases as a “payback” of social
progress. However, we should not sort out thisproblem by simply stepping back in our history,
which is practically impossible. Now, there are many
issues for the medical professionals to work out
including; 1) treatment of lifestyle-related diseases
and their complications, 2) interventions for
improvement of lifestyle behaviors, which does notmake the patients uncomfortable, 3) identification ofrisk group by elucidation of the genetic background,and 4) development of safe and effective
therapeutic drugs, which could also contribute toimprovement of lifestyle behaviors.
I think that the clinical nutrition should work on thesolutions for lifestyle-related diseases under thecurrent comfortable aging society. As a clinician incharge of lifestyle-related diseases, I would like towork out these issues multilaterally, in cooperation
with the professionals in other fields.

postwar period. Yet, many of general people still
express “living” as “eating”, which implicates that thepast hunger experience would unconsciously makeus to seek for gourmet diet (high fat diet). Hence,
the average Body Mass Index (BMI) of Japanesepeople has been increasing, except for young
women. On the other hand, there has been adecreasing trend in the average BMI of young
women, possibly due to dieting with the desire ofbeing slim, and some of them even develop
anorexia nervosa. In addition, the decreased
physical activity due to development oftransportation infrastructure and household electric
appliances would increase the risk of developing
lifestyle-related diseases. Physical activity used to
be a measure (not an objective) to establish living
environment and to secure necessary food, while
the current low physical activity would reflect affluent
and affluent livings where one’s frustration can bereleased by sports, game and even by fighting only.
As a result, body fat is accumulated (obesity),
especially visceral fat, by excess energy intake
which does not match physical activity level. It istherefore obvious that the changes in our lifestyle,especially food culture, have contributed to
substantial increase of lifestyle-related diseases.
Now, how the problems on diet can be solved in theclinical field? Historically, human beings continued
to pursue the missions to secure living environmentthat protect us against bacteria, animal, severe
climate, as well as food that protect us against
hunger and undernutrition, seeking for good health.
However, the progress of scientific technology,
especially after the industrial revolution, has
dramatically improved food production anddistribution system, leading to the change of wholesituation. In the 20th century, the scientifictechnology was mostly utilized for various wars andcolonial rule began. As a negative asset of the 20th
century, many developing countries still suffer frominadequate living environment and hunger in the
21st century too. On the other hand, the developed
countries also face with the global environmental
problems (e.g. environmental prolusion, global
warming etc.), which can not be simply solved by
the advancement of scientific technology. Well, what
did the easy access to nutrient food bring to ourhealth, then? Thanks to improvement in availability* The original Japanese version was translated by Project forInternational Research and Development, NIHN

3

Current Research Projects
Project for Energy Metabolism
Kazuko Ishikawa-Takata
Health Promotion and Exercise Programphysical activity level per day and for each activity isrequired. Now, there are the equipments by which
we can see our own energy expenditure (e.g.
pedometer). Yet, its accuracy is still questioned and
it is necessary to develop more precise equipments.
Furthermore, a simple and precise method tomeasure energy intake is also urgently required.
DLW method could provide the basic data toinvestigate the above aspects as well.
Utilizing the data obtained in the above (1) and (2),the research (3) examines the necessary physicalactivity level and energy expenditure for preventionof disease development and of decrease in thecapacity to live independently, with respect to boththe contents of energy expenditure and nutrient
intake. In addition, we have a collaborative study
with National Center for Geriatrics and Gerontology,
in which we investigate the factors associated withthe decreased capacity to live independently, so as
to propose adequate physical activity and diet.
Since my researches are basically consisted ofhuman studies, I appreciate the understanding andcooperation of the study subjects as well as thecooperation by many staff in other projects/institutes.
Energy metabolism isbased on energy intake,utilization and expenditure,
which is associated withall of nutrition, physiology
and exercise. I would liketo pursue the researches
that would contribute tomaintaining one’s healthand the capacity to live
independently, in terms ofEquipment for pre-treatment
both diet and exercise.

Currently, I belong to Project for Energy Metabolism
in Health Promotion and Exercise Program. In thisproject, I have been working on the researches onenergy metabolism and physical activity. The main
research objectives are as below;(1) To measure energy expenditure and physicalactivity level of various activities, and to
identify the influencing factors.
(2) To develop a simple methodology to assess
energy expenditure and physical activity level.
(3) To investigate the association with developing
disease and maintaining the capacity to live
independently among the elderly, in terms ofboth physical activity and nutrition.
For the research (1), we measure energy
expenditure in daily life, using doubly-labeled water(DLW) method, in which a stable isotope ofhydrogen and oxygen is used. This isotope isincluded in drinking water too, though the amount isvery little. In DLW method, the subjects are asked totake DLW, which contains more stable isotopes than
drinking water, and also to collect their urine once a
day for 1-2 weeks. Using the urine samples, theamount of CO2 produced from our bodies can bemeasured, and then energy expenditure can becalculated. The main advantages of this method
include;1) No restriction with the activities for the
subjects, 2) The required tasks are not too much
(only urine collection and record of collection time),
3) precision of the data is secured. Therefore, themeasurement can be made regardless of one’s age,from young infants to the elderly. In this way, energy
expenditure of various types of subjects, even theathletes, can be measured using DLW method. In
addition, we investigate the factors by which thedaily energy expenditure could change; e.g. type ofactivity, body composition, diet and season.
The research (2) is carried out as a joint study withother projects of our institute, university staffs andprivate companies. Whilst it has been wellacknowledged that DLW method is precise, its costis very high and it is also difficult to measure many
subjects. Besides, we can obtain only the averagedata of energy expenditure for the 1-2 weeks.
Therefore, a simple and precise method to measureof samples in DLW method

Stable isotope ratio mass spectrometer4

Thoughts on Health and Nutrition Research
Traveling the universe of gene
Kouichi Yamada
Nutritional Education Program
When I was looking for the information on
lifestyle-related diseases on the internet, I noted a
website on dieting which says “Let’s check yourobesity gene!”. That is, if one sends the fingernails
or cotton swab that wiped inner mouth to theinspection institute by post, and one’s polymorphism
in a gene can be analyzed. Three obesity genes are
examined by this method; beta 2 adrenergic
receptor (β2AR), beta 3 adrenergic receptor (β3AR)and uncoupling protein 1 (UCP1). In this website, it
is also written as follows; if one has gene mutation(exactly, alteration) in UCP1, s/he is susceptible tosubcutaneous obesity (pear-shaped). Twenty-five
percent of Japanese have the mutation in UCP1,
and their energy metabolism is 100kcal lower thanthat of people without the mutation. Women are
more likely to develop this type of obesity, withhaving a lot of subcutaneous fat in hip and thigh. I
was actually surprised to see the advice written in
the website, which says that “you should take staple
food regularly and less fatty foods, and also take inthe order of vegetable, staple food and meat/fish”.
Indeed, we are still studying to verify the accuracy of
information as well as the evidences.
Locations of single-nucleotide polymorphism(SNP) to be analyzed are Arg16Gly for β2AR,
Trp64Arg for β3AR and Met229Leu for UCP1,
according to the above website. As for UCP1,-3826A>G has been well examined since before,
though most of which reported no association withweight gain. Practically, it should be questionable
UCP1 gene expression can be attenuated by SNP
located 4kb upstream. Nobody has measured thepromoter activity of this region. Mori et al. reported
the Met229-->Leu as well as -112A-->C minor allele
were frequently observed in the type II diabetes
patients, and examined influence of -112 SNP on
UCP1 promoter activity using the reporter assay(Diabetologia 2001, 44, 373-6). They also reported
the linkage disequilibrium between -112 and Met229,
which shows that -112 may be more likely than
Met229 to be the causative SNP. In this way, thecause-effect relationships between SNPs in UCP1
and predisposition to obesity have not been fully
elucidated yet.
The above mentioned “big-three” genes and aperoxisome proliferator-activated receptor gamma(PPARγ2) are known as “four kings” of obesity
genes. Yet, in practice, hereditary factors of obesity
cannot be simply explained by these four genes only,
as more than 50 genes are known to be related toobesity. Besides, there is a new gene that has been
recently drawn attention, that is, RAGE (advanced
glycosylation end product-specific receptor gene).
Well, I could not understand what it meant, when I
heard of it for the first time. Later, searching in the
databases of Entrez-Nucleotide and Entrez-Gene, I
learned that RAGE was the gene located in humanchromosome 6p21.3 and I could also view its
exon-intron structure. I am amazed to see the great
gap between the current advanced technology andthe past one where it took several days forsequencing even dozens or hundreds base by
Maxam-Gilbert method or dideoxy method.
RAGE is the gene that encodes 404 or 302 aminoacid residue proteins. It is not so large gene, but isdivided into 11 exons. This number is rather big for
its size. For example, β2AR and β3AR have only 1exon and 2 exons, respectively. Besides, RAGE is
surrounded by many genes like a cluster housing,whereas the genes around β2AR are scattered withdozens kb of distance to the nearest gene. Likewise,
β3AR also has only 2 genes within 100kb distance.
Um… our creator is really full of whims, isn’t it?
When I look at a large amount of base sequencesin the SNP analyses, I feel that they look like mantra
or secret codes. Exons account for only 2% of
human genome (300 millions bp), and the proteincoding region is only 1.1%. Intron accounts for 24%
and the remainder is meaningless intergenic region.
So, exon looks like a green oasis in desert or evenlike a water planet floating in the universe. In order
to search unknown metabolic code, I would continue
to travel the universe of gene hereafter too

5Latest Research
AMP-activated protein kinase regulates PEPCK gene expression by direct
phosphorylation of AREBP
Erina Inoue, Jun Yamauchi (Nutritional Epidemiology Program)
ATP-activated protein kinase (AMPK) acts as an intracellular sensor for maintaining the energy balance, andis activated by hypoglycemia and exercise in skeletal muscle. The activated AMPK maintains the energybalance by direct phosphorylation of major metabolic enzymes for glucose and lipid and by changing these
enzyme activities. In addition, it has been recently proposed that the activated AMPK regulates gene
expression, and thus AMPK may directly affect activation of transcriptional factors.
Blood glucose is important energy source in human bodies, which is maintained at a certain level by keeping
the balance between hepatic gluconeogenesis and uptake of glucose by peripheral tissue. Excessive hepatic
glucose output through abnormally high levels of glycogenolysis and gluconeogenesis as well as poor uptake
glucose uptake are characteristics of diabetes. Gluconeogenesis is regulated by the key enzymes such as
phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase. PEPCK catalysts the irreversible
conversion during the initial reaction in hepatic gluconeogenesis. Thus, PEPCK gene expression is especially
important, which is mediated by several circulating hormones such as glucagons and glucocorticoids,
whereas it is suppressed by insulin. In the patients with insulin resistant diabetes, PEPCK gene expression
cannot be suppressed by insulin, leading to the vicious cycle for elevated blood glucose. Therefore, anothercontrol system is required, apart from the insulin signaling system. There are several lines of evidence
suggesting that the AMPK signaling pathways can lead to the inhibition of PEPCK gene expression, though
its mechanism is not fully understood.
In order to elucidate the molecular mechanism of AMPK observed in PEPCK gene expression, we searched
for an unknown transcription factor, with the hypothesis that AMPK could mediate the transduction signal that
leads to the repression of PEPCK gene expression through phosphorylation of unknown transcription factor.
Firstly, in order to identify responsive element(s) for AMPK in the human PEPCK gene promoter, functional
analysis was performed using a series of promoter deletion mutants in a transient Luciferase reporter assay
using primary rat hepatocytes. We found the DNA sequence which functioned as an AICAR dependenttranscription repression. To identify the AICAR responsive element binding protein (referred to as AREBP), a
yeast one-hybrid assay was carried out. Out of 108 colonies, three identical cDNA clones were isolated. There
are several putative AMPK phosphorylation consensus sites in the AREBP molecule.
Next, we tested whether the isolated AREBP would actually act as an AICAR dependent regulator of the
PEPCK gene by a transient Luciferase reporter assay using rat hepatoma cells. Although obvious differenceswere not observed with AREBP alone compared with mock transfection, the AREBP robustly repressedtranscription in the presence of AICAR. In contrast, AREBPS470A binging was not influenced by AMPK
phosphorylation, even in the presence of AICAR. In addition, when the expression of endogenous AREBP
was knocked down using the RNA interference (RNAi) technique, the repression of PEPCK expression
following AICAR treatment was eliminated by AREBP-RNAi.
These results showed that AREBP would act as an AICAR dependent transcriptional repressor for PEPCK
gene expression. It must be noted, however, that these results were derived from the analyses in vitro or atmolecular level. In order to investigate the functions of AREBP in vivo, further researches are in progressusing mouse with excessive AREBP expression. By understanding the regulation system of gluconeogenesis,
we expect that our findings will be utilized for prevention and control of diabetes in the future.

AMP-activated protein kinase regulates PEPCK gene expression by direct
phosphorylation of a novel zinc finger transcription factor.
Biochem Biophys Res Commun. 2006;351(4):793-9.
Inoue E and Yamauchi J
Nutritional Epidemiology Program, Bio-Index Project, National Institute of Health and Nutrition, Tokyo, Japan

Abstract:
AMP-activated protein kinase (AMPK) acts as an intracellular sensor for maintaining theenergy balance. Activation of AMPK switches on ATP-generating process while switches off ATP-consuming
process. It achieves these effects by phosphorylation of downstream metabolic enzymes. It has been
proposed that AMPK also regulates gene expression through phosphorylation of certain transcription factors;
however its molecular mechanism is not fully understood. Here we show the cloning and characterization of anovel zinc finger transcription factor referred to as AREBP. AREBP is phosphorylated at Ser(470) by AMPK.
Phosphorylation reduces the DNA-binding activity of AREBP. Transient transfection experiments indicate thatwild-type AREBP, but not Ser(470) to Ala(470) substituted non-phosphorylating mutant, represses gene
expression of the phosphoenolpyruvate carboxykinase (PEPCK), a key enzyme of gluconeogenesis. RNAinterference-mediated reduction of endogenous AREBP expression attenuates AMPK-induced PEPCK
down-regulation. These results implicate AREBP as a novel key modulator of PEPCK gene expression
regulated by AMPK.
6

Latest Research
Young Japanese adults with high BMI tend to under-report energy intake – from the
dietary records of the population aged 30’s-60’s.
Hitomi Okubo✝, Satoshi Sasaki (Nutritional Epidemiology Program)✝

Okubo H is currently with Kagawa Nutrition University

Reporting bias, underreporting or overreporting, is often observed in any type of self-reported dietaryassessment methods. This is one of the big headaches for dietitians. So far, all the available studies onreporting bias were conducted in Western countries, and few studies has been undertaken in Japan. Diet
record is one of the self-reported dietary assessment methods, which is believed to be reliable as the study
subjects record what they actually took. The purpose of the present study was to examine the relative
accuracy of self-reported energy intake obtained by diet records among various age ranges in the Japanese
population.
We selected three areas which have different geographical conditions in Japan: Osaka (urban), Nagano(rural inland) and Tottori (rural coastal). Out of 96 married couples aged ( 30 years who were recruited, 183
subjects (91 women and 92 men) with complete 16-day diet records were included in the present analysis.
The subjects completed 4-day semi-weighed diet records four times at 3-month intervals (16 days in total)from November 2002 to August 2003. We calculated the ratio pf energy intake to basal metabolic rate
(EI/BMR) to evaluate the relative accuracy of the reported energy intake. Low ratios describe subjects
reporting comparatively low energy intake relative to their energy requirement, hence high likelihood of
underreporting.
Mean EI/BMR was lowest in the subjects aged 30-39 years and highest in those aged 60 years or over (1.37
vs. 1.74 in male, 1.43 vs. 1.65 in female). A stepwise multiple regression analysis was performed to examine
the prediction for relative accuracy of reporting. Regardless of sex, age correlated positively and BMI
correlated negatively. Figure 1 and 2 visually show the joint effect of age and BMI on EI/BMR values bycross-classifying subjects by both variables, with tertile of BMI on the x-axis, age group (30-49, ≥ 50 years) on
the z and mean value of EI/BMR on the y. Compared with subjects classified into the lowest BMI and oldest
age group, subjects in the highest BMI and youngest age group had significantly lower EI/BMR (Fig 1).
Similar trend was observed in female as well (Fig. 2). These findings indicate that the characteristics of the
population prone to underreport energy intake were young people with high BMI.
Since the participants of this study
Figure 1
Figure 2(male)(female)might be highly health-conscious,
we must be cautious to apply thesetrends to the whole general
Japanese population. Yet, ourfindings would suggest that
dietitians should bear in mind thepossibility of underreporting when ahighlownutritional
assessment
is
low
low
high
high
Age
high
low
undertaken for the subjects withAgehigh BMI or young ones.
EI/BMR were adjusted for physical activity level and living area. * p<0.05, **p<0.01, ***p<0.001

The influence of age and body mass index on relative accuracy of energy intake
among Japanese adults
Public Health Nutrition. 2006; 9(5): 651-657.
Okubo H1), Sasaki S1), Hirota N2), Notsu A3), Todoriki H4), Miura A5), Fukui M6) and Date C7)1) Scientific Evaluation of Dietary Reference Intakes Project, National Institute of Health and Nutrition, Tokyo, Japan
2) Department of Living Sciences, Nagano Prefectural College, Nagano, Japan: 3) Tottori College, Tottori, Japan:
4) Department of Environmental and Preventive Medicine Faculty of Medicine, School of Medicine, University of
Ryukyus, Okinawa, Japan. 5) Department of Nutritional Health, Kwassui Women’s College, Nagasaki, Japan:
6) Department of Statistics, Osaka City University Medical School, Osaka, Japan. 7) Department of Food Sciences andNutrition, School of Human Environmental Sciences, Mukogawa Women’s University, Hyogo, Japan

Abstract:
Objective: To examine relationships between the ratio of energy intake to basal metabolic rate (EI/BMR) andage and body mass index (BMI) among Japanese adults. Design: Energy intake was assessed by 4-day
semi-weighed diet records in each of four seasons (16 days in total). The EI/BMR ratio was calculated from
reported energy intake and estimated basal metabolic rate as an indicator of reporting accuracy. Setting:Residents in three areas in Japan, namely Osaka (urban), Nagano (rural inland) and Tottori (rural coastal).
Subjects: One hundred and eighty-three healthy Japanese men and women aged ≥ 30 years. Results: The
oldest age group (≥60 years) had higher EI/BMR values than the youngest age group (30–39 years) in both
sexes (1.74 vs. 1.37 for men; 1.65 vs. 1.43 for women). In multiple regression analyses, age correlated
positively (partial correlation coefficient, β=0.012, P < 0.001 for men; β=0.011, P < 0.001 for women) and BMI
correlated negatively (β=-0.031, P < 0.001 for men; β=-0.025, P < 0.01 for women) with EI/BMR.
Conclusion: Age and BMI may influence the relative accuracy of energy intake among Japanese adults.
7

Latest Research
Effects of age on ventilatory threshold and peak oxygen uptake normalized forregional skeletal muscle mass: a cross-sectional study on Japanese men and women
aged 20-80 years.
Motohiko Miyachi (Health Promotion and Exercise Program)【Background】Ventilatory threshold (VT) is a valuable index of cardiorespiratory fitness as well as physical exercise
intensity which is required for developing an exercise program for health promotion. However, few studies have examined
the age-associated decline of VT. This study therefore aimed to demonstrate normative VT values and age-associateddecline of VT normalized for skeletal muscle (SM) mass in 1,463 Japanese men and women aged 20-80 years.
【Methods】 Fourteen hundred and sixty-three healthy Japanese men and women, who are members of a fitness club inTokyo, participated in this study (807 men and 656 women aged 20-80 years, with the mean age of 49.3 ± 13.5 years). SM
mass was determined by B-mode ultrasound and estimated using the equations developed by Sanada. VT and peakoxygen uptake (VO2peak) were determined during treadmill walking, for which VO2 were measured using an
automated breath-by-breath mass spectrometry system. We developed the protocol in which the subjects warmed-up for 3
min, and then, the treadmill speed and grade were increased alternately for each successive minute of walking. VO2peak was
estimated using the heart rate (HR)-VO2 relationship during sub-maximal exercise and the predicted maximal heart rates(HR)(220 minus age). VT was estimated from ventilatory equivalents for oxygen (VE/VO2) and carbon oxide (VE/VCO2). VT
was determined as the point where VCO2 increased sharply compared to VO2, or the one where VE/VCO2 became unchanged
and VE/VO2 increased.
【Results and Discussion】Total and regional (trunk and thigh) SM mass were negatively correlated with age in both men
and women (Total SM mass decreased 0.13kg in men and 0.02kg in women per year, and the thigh SM mass decreased
0.052kg in men and 0.014kg in women per year). VT was significantly associated with thigh SM mass independent of age,body mass and fat-free mass. The most important finding in this study was that age-associated declines were observed in
VT normalized for body mass in both men and women, but not in VT normalized for SM mass (Table). On the other hand,age associated declines were observed for VO2peak normalized for body mass as well as normalized for SM mass. Thus, thisstudy showed that the age-associated declines in VT are markedly blunted if normalized for SM mass rather than body
Table. Absolute and normalized VT in various age groups
mass. These results suggest thatthe decrease in VT with age isprimarily due to an age-related
decline of SM mass.
【 Conclusion 】 This study
provided cardiorespiratory fitness
data, using VT, according togender and age group. Ourfindings would suggest that it isimportanttoprevent
theage-related decline in SM masssoastoprevent
theage-associated declines in VT,
namely cardiorespiratory fitness.

Effects of age on ventilatory threshold and peak oxygen uptake normalised for regionalskeletal muscle mass in Japanese men and women aged 20-80 years.
European Journal of Applied Physiology. 2007; 99:475-483
Sanada K1), Kuchiki T2), Miyachi M3), McGrath K4), Higuchi M5), Ebashi H6),

1) Consolidated Research Institute for Advanced Science and Medical Care, Waseda University, Tokyo, Japan
2) Division of Integrated Humanistic and Cultural Studies, Graduate School of Integrated Science and Art,
University of East Asia, Shimonoseki, Japan
3) National Institute of Health and Nutrition, Tokyo, Japan
4) Department of Physiological Sciences and Sports Performance, National Institute of Fitness and Sports, Kanoya, Japan
5)Faculty of Sport Sciences, Waseda University, Tokorozawa, Japan
6) Faculty of Integrated Cultures and Humanities, University of East Asia, Shimonoseki, Japan

Abstract: Ventilatory threshold (VT) is an important predictor of cardiorespiratory fitness, such as peakoxygen uptake (VO2peak) and is a valuable index of aerobic exercise intensity. However, little is known about therole of skeletal muscle (SM) mass in the age-associated decline of VT. Therefore, the present study was
performed to investigate the effects of age on cardiopulmonary fitness normalised for regional SM mass in 1,463
Japanese men and women, and to determine the relevance of VT normalised to SM mass based on age andgender. Total, trunk and thigh SM mass were measured using an ultrasound method, while VO2peak and VT were
determined during treadmill walking. VO2peak was estimated using the predicted maximum heart rate (HR) and the
HR- VO2 relationship for sub-maximal treadmill walking. There were significant negative correlations between VT
normalised for body mass and age in men and women (P < 0.001). Age-associated declines were also observedin VT normalised for body mass in both men and women; however, VT normalised for SM mass was not
significantly different with age. Significant correlations were also observed between thigh SM mass and VT inboth men and women. These results suggest that thigh SM mass is closely associated with VO2peak and/or VT inboth men and women, and the decrease in VT with age is predominantly due to an age-related decline of SM
mass. Moreover, this study provides normative cardiorespiratory fitness data regarding VT normalised SM massin healthy men and women aged 20-80 years.
8