nature publishing group
No Evidence of an Effect of Alterations in
Dietary Fatty Acids on Fasting Adiponectin
Over 3 Weeks
Fiona E. Lithander1,2,3, Geraldine F. Keogh1,2, Yu Wang2, Garth J.S. Cooper2,4, Tom B. Mulvey2,
Yih-Kai Chan1,2, Brian H. McArdle5 and Sally D. Poppitt1,2,4
objective: Little is known about the effects of alterations in fatty acid classes on adiponectin, a hormone secreted by
the adipocyte known to be important in the development of diabetes and cardiovascular disease (CVD). Any factor,
including diet, that may positively influence adiponectin gene expression or increase circulating levels might be useful
for improving such metabolic abnormalities. We investigated the effects of alterations in dietary fatty acid saturation
on fasting serum adiponectin and associated peptides.
Methods and Procedures: Double-blind, randomized, crossover, 2 × 3-week residential intervention trial where
18 mildly hyperlipidemic adult men were provided with a high saturated:unsaturated fat (SFA:USFA) and lower
SFA:USFA treatment separated by an uncontrolled 4-week washout. Only fatty acid profile was altered between
treatments. Fasting blood samples were collected on days 0, 1, 7, 14, 21, 22 of each intervention period for the
measurement of adiponectin, tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), high-sensitivity C–reactive protein
(hsC-RP), leptin, and ghrelin.
Results: Body weight was kept constant (±1 kg) throughout each treatment. There was no detectable difference
in fasting adiponectin at baseline (mean day 0 + day 1) between the treatment groups (mean ± s.d.; highSFA:USFA =
7.0 ± 1.7 vs. lowSFA:USFA = 6.7 ± 1.4 µg/ml, P > 0.05). There were neither significant between-treatment effects of fatty
acid saturation (diet × time, P > 0.05) on serum adiponectin nor any significant between-treatment effects on serum
TNF-α, IL-6, hsC-RP, leptin, or ghrelin (P > 0.05).
Discussion: Fasting serum adiponectin was not detectably affected by alterations in dietary fatty acid profile in mildly
hyperlipidemic men. There was no evidence that an increase in SFA content of the diet significantly worsened fasting
serum adiponectin over a 3-week intervention period.
Obesity (2008) doi:10.1038/oby.2007.97
Adipose tissue–derived adiponectin is an insulin-sensitizing
hormone that is associated with decreased risk of cardiovascular
disease (CVD) and type 2 diabetes (T2DM) (1,2). Circulating
levels of adiponectin have been shown to be inversely correlated
with total levels (3,4), and visceral adiposity (5) and adverse
metabolic states such as dyslipidemia (6), hyperglycemia, and
insulin resistance (2,7,8). A suppressed level of adiponectin
gene expression in the adipose tissue of obese subjects has also
been demonstrated (3,9). Importantly, adiponectin is linked
to fat metabolism per se whereby exogenous administration of
globular adiponectin can increase free fatty acid oxidation by
11–19% in muscle in mice, increase removal of free fatty acid
from the circulation, and lead to the loss of adipose mass (10).
In addition, it has been shown that chronic infusion of adi-
ponectin produced from Escherichia coli significantly amel-
iorated insulin resistance and improved lipid profiles in both
lipoatrophic diabetic mice and diet-induced obese mice (11).
Therefore, any factor that may positively influence adiponec-
tin gene expression or circulating levels of this hormone might
be a useful agent for improving insulin resistance, risk factors
for T2DM, and CVD, or potentially even decrease the adipose
mass in obese individuals.
Alterations in diet have been shown to affect the markers of
disease risk such as glucose, lipids, and insulin. The relation-
ship between dietary fat, especially saturated fatty acid (SFA),
1Human Nutrition and Metabolic Unit, University of Auckland, Auckland, New Zealand; 2School of Biological Sciences, University of Auckland, Auckland, New Zealand;
3School of Medical Sciences, University of Auckland, Auckland, New Zealand; 4Department of Medicine, University of Auckland, Auckland, New Zealand; 5Department
of Statistics, University of Auckland, Auckland, New Zealand. Correspondence: Fiona E. Lithander (email@example.com)
Received 12 June 2007; accepted 15 July 2007; advance online publication 17 January 2008. doi:10.1038/oby.2007.97
and CVD has long been understood (12,13), and it is known
that a diet rich in SFA can lead to increases in total cholesterol
and low-density lipoprotein-cholesterol (12), both of which
are independent risk factors for CVD. A diet rich in SFA is also
known to adversely affect the risk of T2DM (14). In light of
the inverse association between adiponectin (both circulating
and gene expression) and CVD and T2DM, and the positive
association between dietary fat intake and CVD and T2DM,
it seems pertinent to explore the link between fasting serum
adiponectin and fat intake, and in particular alterations in fatty
Long-term changes in dietary intake can influence adi-
ponectin indirectly through changes in adipose mass (3,4),
but whether diet per se has an independent effect on this
hormone has yet to be established. Intervention trials, which
make acute changes in diet, have shown mixed results with
the evidence of both an increase (15) and a decrease (16) in
circulating adiponectin up to 4 h following consumption of
meal. Alternately, other trials, including one from our own
laboratory (17), have shown that circulating adiponectin does
not alter postprandially in response to a meal (18,19). The
relationship between medium-term dietary manipulation and
adiponectin has been investigated in two trials. Circulating
levels have been shown to be unaffected by alteration in the
fat content of a eucaloric diet over 7 days involving 21 sub-
jects (20). Bonnet and colleagues have demonstrated higher
levels of the hormone in the individuals supplemented with
high ω-3 polyunsaturated fatty acids (PUFAs) fatty fish at least
three times per week in addition to 20 g high monounsatu-
rated fatty acids (MUFAs) rapeseed oil per day over 10 weeks
compared with baseline levels (21).
Adiponectin has been shown to be closely related to cytokines
such as tumor necrosis factor-α (TNF-α) and interleukin-6
(IL-6). Although both of these hormones play an important
role in carbohydrate and lipid metabolism (22,23), unlike
adiponectin they are positively correlated with T2DM, CVD,
and obesity. Dietary fatty acid composition has been shown to
affect circulating TNF-α, IL-6, and C–reactive protein (C-RP).
Intervention trials, which last from 4 to 12 weeks, have demon-
strated either an increase (24) or a decrease (25–28) in TNF-α
and IL-6 as a result of feeding diets high in PUFA. Similarly,
C-RP has been shown to increase (29), decrease (30,31), or
not change (32) in response to alterations in fatty acid profile.
Alterations in diet where body fat remains unchanged have
been shown not to affect fasting leptin concentrations (33),
and it has been proposed that circulating leptin may be driven
by total adipose mass (34) as opposed to being affected by
diet. There is little evidence to indicate that ghrelin responds
to medium/longer-term feeding, especially alterations in fatty
Hence, this trial was designed to investigate whether fast-
ing levels of serum adiponectin and the associated peptides
TNF-α, IL-6, C-RP, leptin, and ghrelin are altered by changes
in dietary fatty acid saturation made over a 3-week period in
a group of hyperlipidemic men known to be at risk of CVD
through elevation of blood cholesterol.
methods and procedures
Eighteen men with mild hyperlipidemia (recruited on the basis of low-
density lipoprotein-cholesterol 3.0–5.0 mmol/l) who were otherwise
healthy completed both arms of the trial. At the time of participation
in the trial, they were taking no medication for hyperlipidemia or any
other condition, nor had they any current or previous history of CVD,
T2DM, or any other metabolic disorder. Written consent was obtained
from each participant, and ethical approval obtained from the Auckland
Ethics Committee, Auckland, New Zealand.
protocol and diets
All subjects in this randomized, double-blind, crossover intervention
trial were required to reside at the University of Auckland Human
Nutrition and Metabolic Unit for both dietary periods, which com-
prised two treatment arms each of 3-week duration. Work from our
own (35) and other laboratories (36) have demonstrated that 3 weeks
is of sufficient duration to see a decrease in serum total cholesterol and
low-density lipoprotein-cholesterol when dietary SFA are replaced by
unsaturated fats. It is reasonable to hypothesize that if such traditional
risk factors change within a 3-week time frame, emerging risk factors
such as adiponectin may change within the same period.
Between treatments, subjects returned home for a washout period of
4 weeks, during which time they were asked to resume their habitual diet
and exercise patterns. The energy and macronutrient content of the diet
was initially calculated using the dietary program, Foodworks (version
2.05, New Zealand Edition, Xyris Software, Brisbane, Australia, 1999),
and then verified by direct chemical analyses of duplicate diet samples.
Eight duplicate 5-day diets were collected at intervals throughout the
trial, homogenized, and an aliquot frozen for later chemical analysis. This
enabled the composition of the diet to be verified, and also demonstrated
that there were no seasonal trends in composition.
Subjects were randomized using stratification to ensure that half
were given the high saturated fatty acid:unsaturated fatty acid treat-
ment (highSFA:USFA) and half the low SFA:USFA (lowSFA:USFA) on entry into
the study. All subjects then crossed over on to the other treatment arm.
Subjects were provided with breakfast, lunch, and dinner in addition
to between-meal snacks in the form of a 5-day rotation diet. Subjects
were maintained in energy balance throughout the trial based on an
estimate of basal metabolic rate (37) and energy expenditure, and food
intake was altered daily to maintain a constant body weight (±1 kg).
Subjects weighed themselves daily after an overnight fast and after void-
ing the bladder. Breakfast and dinner were taken under supervision at
the Human Nutrition and Metabolic Unit, while lunch was packed and
subjects could consume this meal at their place of work or study. Data
on 24-h urinary nitrogen balance collected on day 10 and 20 of each
intervention period were used to assess independent dietary compli-
ance. Urinary losses of nitrogen were directly compared with dietary
protein intake using para-amino benzoic acid to verify the completeness
of collection (38–40). Decaffeinated, energy-free beverages were freely
available throughout the intervention, and alcohol was prohibited at
all times. Subjects were required to eat only and all of the foods pro-
vided and no others. The dietary compositions of both treatments are
presented in Table 2.
Blood sampling and analyses
Blood and urine samples were routinely collected throughout both
intervention periods. Fasting venous blood samples were collected
on the morning of days 0 and 1 (pre-intervention baseline), 7, 14,
21, and 22. Once collected, blood samples were centrifuged and
serum stored at –80 °C until later batch analyses. Samples were ana-
lyzed for adiponectin, TNF-α, IL-6, high-sensitivity C-RP (hsC-RP),
leptin, and ghrelin. Adiponectin was analyzed by enzyme-linked
immunosor bent assay using an in-house assay. Serum samples were
diluted 1:5000 with phosphate-buffered saline and then added to
96-well microliter plates coated with monoclonal antibody against
human adiponectin (R & D System, Minneapolis, MN). After incu-
bation at room temperature for 120 min, wells were washed and
incubated for another 60 min with the biotinylated monoclonal anti-
body against adiponectin (R & D System, Minneapolis, MN). The
wells were again washed and incubated with streptavidin-conjugated
horseradish peroxidase for 60 min and then reacted with tetra-me-
thyl benzidine reagent for 15 min. To each well, 100 µl of 3 M HCl
was then added to stop the reaction and the absorbance at 450 nm
was measured. Intra- and inter-assay coefficients of variation were
6.2–7.9% and 3.8–6.3%, respectively. Lower limits of detection for
the assay were 0.5–2 ng/ml of adiponectin protein.
Serum IL-6 and TNF-α were analyzed by enzyme-linked immunosorb-
ent assay using a Duoset commercial kit (R & D System, Minneapolis,
MN). Then, 100 μl of sample was applied to 96-well microtitre plates
and incubated with the individual coating antibodies for 2 h at room
temperature. After washing with phosphate-buffered saline, the detec-
tion antibodies were applied for another 2 h at room temperature. The
bound immune-complexes were read at 450 nm. The standard curve was
generated for every set of samples assayed using the standards provided
in the kit. hsC-RP was analyzed using a Pointe Scientific (Lincoln Park,
MI) immuno-turbidometric commercial kit. A solution of latex particles
coated with the antibody specific to human C-RP aggregate was added to
form immune complexes. Increased light scattering, proportional to the
concentration of analyte, was measured on a COBAS Mira auto-analyzer
(Roche Diagnostics, Basel, Switzerland). C-RP concentration was cal-
culated using a calibration curve of C-RP standards and Prism software
(GraphPad, San Diego, CA) used to fit third-order polynomials to the
curve to calculate sample concentration. Assay range was 0.05–10 mg/l
with a sensitivity of 0.1 mg/l.
Total ghrelin concentrations were measured by radioimmunoassay
using a Linco Research commercial kit (Saint Charles, MI), using 125I-
labelled ghrelin as the tracer. Primary antibody was rabbit anti-ghrelin,
secondary antibody was goat anti-rabbit IgG, and precipitation was
achieved using polyethylene glycol (PEG) in a phosphate and Triton-
X100 buffer. Hundred microliters of sample and standards were incubated
with primary antibody at 4 °C overnight. After 24 h, 100 µl of tracer was
added and incubated overnight at 4 °C. After 48 h, 1.0 ml of precipitat-
ing reagent was added, tubes incubated at room temperature for 20 min,
and then centrifuged at 1,700g for 20 min at 4 °C. The supernatant was
decanted and the pellets were counted on a Wallac 1480 Wizard gamma
counter (Wallac Finland Oy, Turku, Finland). Linco Research reagents
were prepared and stored according to the manufacturer’s instructions.
Curve fitting and sample concentration were computed using the Mul-
tiCalc software supplied with the counter. Coefficient of variation for
this assay across the full standards range was 3.3%. The lowest level of
detection was 0.311 ng/ml.
Serum leptin was measured by enzyme-linked immunosorbent assay
using a Duoset commercial kit (R & D System, Minneapolis, MN).
Hundred microliters of sample was added to 96-well microtitre plates
and incubated with the individual coating antibodies for 2 h at room
temperature. After washing with phosphate-buffered saline, the detection
antibodies were applied for another 2 h at room temperature. The bound
immune-complexes were measured at 450 nm. The standard curve was
generated for every set of samples assayed using the standards provided
in the kit.
Paired t-test analyses (two tailed) were used to identify any differences
in dietary energy or macronutrient composition between the lowSFA:USFA
and highSFA:USFA diets as taken by the subjects. All anthropometric and
metabolic variables including body weight, adiponectin, TNF-α, IL-6,
hsC-RP, leptin, and ghrelin were analyzed using linear mixed model
ANOVA (SAS: PROC MIXED, SAS version 8.0, SAS Institute, Cary, NC,
2001). The dietary treatment, the arm of the trial (stratum), the inter-
vention period, and the study day within period factors were explicitly
modeled as fixed factors, as was the treatment/day interaction, which
addressed whether the trajectory over time during the intervention
period differed between treatments (diet × time). Subjects within strata
were treated as random, as were their interactions with day and inter-
vention period. Repeat baseline measures before intervention (d0, d1)
were combined into a single mean value to reduce variability at base-
line. Repeat data collected at the end of the intervention (d21, d22) were
not combined. Variable intervals between blood collections were also
included in the analyses such that the unequal number of days between
measurements was modeled as an autoregressive order 1 process with
constant day-to-day correlation. Biochemical assays were analyzed in
triplicate and presented as a mean ± s.e.m. Statistical significance was
based on 95% limits (P < 0.05).
Subject characteristics are shown in Table 1. One subject with-
drew from the trial on day 13 of the highSFA:USFA treatment and
was replaced in the randomization. All other subjects com-
pleted the trial. Body weight was kept constant and was strictly
maintained within the limits of ±1 kg during the two interven-
tion arms. There was neither any significant difference in body
weight at day 0 between the two treatments (P > 0.05) nor any
change during the intervention, which would have affected the
fasting parameters (P > 0.05), as shown in Figure 1.
The calculated macronutrient composition of the diet was
38%, 49%, and 13% of total energy for fat, CHO, and protein,
respectively. Dietary cholesterol was lower on the lowSFA:USFA
treatment, but there was no difference between total fat, CHO,
and protein between treatments (P > 0.05). An indepen-
dent measure of dietary compliance using the 24-h nitrogen
balance method was carried out on four occasions for each
table 1 Baseline characteristics of the 18 men who
completed both arms of the intervention. all measurements
made at screen visit
Baseline Mean s.d. Range
Age (years)39.713.9 23.0–67.0
Body weight (kg)81.913.162.4–114.6
Waist (cm)93.89.9 78.0–114.0
Hip (cm)101.4 7.8 90.0–118.0
Waist:hip ratio 0.9 0.00.8–1.0
Total cholesterol (mmol/l)5.8 0.7 4.8–6.6
LDL-cholesterol (mmol/l)3.9 0.53.1–4.6
HDL-cholesterol (mmol/l) 1.2 0.3 0.9–1.8
TAG (mmol/l)1.5 0.7 0.5–3.1
TC:HDL-C ratio4.9 1.1 3.3–6.9
SBP (mm Hg) 127 21104–182
DBP (mm Hg)817 68–92
Glucose (mmol/l) 4.8 0.44.0–5.3
BMI, body mass index; DBP, diastolic blood pressure; HDL, high-density lipoprotein;
LDL, low-density lipoprotein; SBP, systolic blood pressure; TAG, triacylglycerides;
TC:HDL-C ratio, Total cholesterol:high-density lipoprotein–cholesterol ratio.
subject. There was reasonable long-term dietary compliance.
The mean ± s.e.m. of urinary N as a percentage of dietary N
was 74 ± 4%. The composition of the diet on both treatments
as measured by direct chemical analysis is shown in Table 2.
SFAs, MUFAs, and PUFAs accounted for 53, 32, and 30 g/d
of the total diet in the highSFA:USFA treatment, respectively, and
38, 41, and 31 g/d of the total diet in the lowSFA:USFA. When cal-
culated as a percentage of total daily energy intake (%), total
saturates were 5% lower and total unsaturates 3% higher on
the lowSFA:USFA treatment. The most substantial differences were
in palmitic (C16:0, −10.7 g/d), myristic (C14:0, −3.4 g/d), oleic
(C18:1, +10.8 g/d) acids.
There was no significant difference at baseline (mean day 0 +
day 1) between the highSFA:USFA and the lowSFA:USFA treatments
in serum adiponectin (mean ± s.d.; 7.0 ± 1.7 vs. 6.7 ± 1.4 μg/
ml, P = 0.19), TNF-α (563.1 ± 372.2 vs. 606.4 ± 425.3 pg/ml,
P = 0.09), IL-6 (20.8 ± 28.9 vs. 18.9 ± 24.9 pg/ml, P = 0.19),
hsC-RP (1.4 ± 1.3 vs. 1.4 ± 0.9 mg/l, P = 0.72), or ghrelin
(783.2 ± 131.2 vs. 785.3 ± 211.0 pg/ml, P = 0.95). Leptin was
significantly higher at baseline (mean day 0 + day 1) on the
highSFA:USFA treatment (2186.4 ± 536.6 vs. 2002.7 ± 338.5, P =
0.04). Table 3 shows the effects of both treatments on fast-
ing levels of these hormones between day 0 and the end of
the intervention period (day 22). Figure 2 shows the change
relative to d0 in adiponectin, TNF-α, IL-6, and hsC-RP.
Figure 3 shows the change relative to d0 for leptin and ghre-
lin. Fasting adiponectin levels were not affected by treatment,
demonstrating that alterations in fatty acid saturation did
not affect the circulating adiponectin (diet × time, P = 0.10).
TNF-α (diet × time, P = 0.74), IL-6 (diet × time, P = 0.22),
and hsC-RP (diet × time, P = 0.39) were also not affected by
fatty acid saturation of the diet, and there was no significant
effect of fatty acid saturation on leptin (diet × time, P = 0.56)
or ghrelin (diet × time, P = 0.65).
Little is known about the effects of diet on serum adiponectin
in humans and the extent to which dietary intake may modu-
late this adipokine. In this trial we have shown that a diet rich
in USFA such as MUFA, compared with a diet high in SFA, has
no effect on the fasting levels of total adiponectin or associated
Body weight (kg)
0 17 14
Delta body weight
Body weight (kg)
Figure 1 No significant change in body weight during 3 weeks on
highSFA:USFA (open circles) and lowSFA:USFA (closed circles) treatments.
Mean ± s.e.m.
table 2 composition of the highsFa:usFa and the lowsFa:usFa
treatments as measured by direct chemical analysis;
mean ± s.d.a
Energy intake, EI (range, MJ/d)
EI (mean, MJ/d)b
13.3 ± 1.513.3 ± 1.60
CHO (% of energy)
53 ± 253 ± 30
Protein (% of energy)
13 ± 1 13 ± 10
Fat (% of energy)
34 ± 134 ± 30
232 ± 93
18 ± 0.4
236 ± 55
13 ± 0.4
Total SFA (calculated, en%)d
SFA profile (g/d)
Total52.9 ± 5.5
3.1 ± 1.1
8.3 ± 1.3
28.0 ± 2.7
11.8 ± 1.5
10 ± 0.7
37.9 ± 2.9
1.6 ± 0.2
4.9 ± 0.6
17.3 ± 1.3
13.3 ± 1.3
12 ± 0.7
Total MUFA (calculated, en%)d
MUFA profile (g/d)
Total 32.3 ± 5.6
1.8 ± 0.3
29.7 ± 4.9
7 ± 1
41.2 ± 4.4
0.8 ± 0.9
40.5 ± 3.8
8 ± 1
Total PUFA (calculated, en%)d
PUFA profile (g/d)
Total30.2 ± 4.3
27.1 ± 3.5
3.1 ± 1.1
31.4 ± 6.2
27.7 ± 5.0
3.7 ± 1.3
MUFA, monounsaturated fatty acid; PUFA, polyunsaturated fatty acid; SFA, satu-
rated fatty acid; USFA, unsaturated fatty acid.
aResults are the mean values for eight duplicate portions analyzed from both
highSFA:USFA and lowSFA:USFA treatments; bno significant difference between treat-
ments (P > 0.05). Minor fractions of fatty acids not shown; ccholesterol intake
shown for a 13.3 MJ diet; dcalculated from Foodworks dietary program.
peptides, TNF-α, IL-6, C-RP, leptin, or ghrelin, when given to a
group of mildly hyperlipidemic men over a 3-week period.
The adipocyte secretes adiponectin in the form of distinct
complexes (41–44). Trimers are the building blocks of low-
molecular weight adiponectin; two subunits of the adiponec-
tin trimer link to form a hexamer or middle-molecular weight
adiponectin (41,43), while hexamers are the raw material for
the formation of high-molecular weight adiponectin. Post-
translational modifications are required for folding and assem-
bly into the higher-order structures (41). Different multimers
of adiponectin trigger different signal transduction pathways
and exercise distinct functions on its target tissues (45–47).
Any agent or intervention, including dietary intake that causes
a change in the circulating level of full-length adiponectin,
would necessarily have to interfere with at least one of the steps
involved in the process of producing these multimers.
Epidemiological evidence that has investigated the relation-
ship between dietary intake and circulating full-length adi-
ponectin is inconclusive. In a cross-sectional analysis using
semiquantitative food frequency questionnaires, Pischon
and colleagues (48) demonstrated in a study of 532 men that
moderate alcohol intake was related to higher serum adi-
ponectin concentrations, and conversely a diet rich in carbo-
hydrate, which promotes glycemia, was associated with lower
table 3 effects of highsFa:usFa and lowsFa:usFa dietary treatments on serum markers measured at baseline (mean day 0 + day 1) and
on days 7, 14, 21, and 22
day 0 + day 1 Day 7Day 14 Day 21 Day 22
7.0 (0.3)7.2 (0.5) 6.9 (0.4)7.3 (0.4) 6.5 (0.4)
563.1 (62.0)566.3 (99.2) 546.6 (87.4)560.7 (99.8) 548.4 (99.2)
20.8 (4.8)26.2 (8.0) 20.4 (7.0) 19.3 (6.5)20.5 (6.9)
hsC-RP (mg/l)1.4 (0.2) 1.7 (0.6) 1.6 (0.4)1.9 (0.4) 1.5 (0.3)
Leptin (pg/ml)2186.4 (89.4)* 2101.6 (121.9)2031.7 (118.7) 2041.2 (96.9)2182.9 (122.8)
Ghrelin (pg/ml)783.2 (21.9) 841.8 (50.8)888.9 (53.7) 841.3 (37.4)847.0 (54.7)
6.7 (0.2) 6.9 (0.4)6.6 (0.4) 7.0 (0.7)7.8 (0.6)
606.4 (70.9)593.3 (101.7)563.4 (93.1) 548.9 (88.5)526.4 (84.6)
18.9 (4.2) 19.4 (6.2)18.5 (6.1) 21.3 (7.1) 20.8 (6.8)
hsC-RP (mg/l)1.4 (0.2)1.2 (0.2)1.9 (0.6) 1.2 (0.2)1.0 (0.2)
Leptin (pg/ml) 2002.3 (55.6)*2049.9 (86.0) 1971.5 (87.8) 1972.0 (64.7)2026.3 (80.1)
Ghrelin (pg/ml) 785.3 (35.2)806.8 (45.5)820.5 (51.7) 854.1 (65.9)855.7 (78.9)
Significant difference at baseline, paired t-test. Mean (s.e.m.).
hsC-RP, high-sensitivity C–reactive protein; IL-6, interleukin-6; SFA, saturated fatty acid; TNF-α, tumor necrosis factor-α; USFA, unsaturated fatty acid.
*P < 0.05; no significant effects of treatment over time, ANOVA, P > 0.05.
017 14 2122
Figure 2 Change from baseline in fasting adiponectin, tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and high-sensitivity C–reactive protein
(hsC-RP) during the 3-week highSFA:USFA (open circles) and lowSFA:USFA (closed circles) treatments. Mean ± s.e.m.
concentration. Yannakoulia and co-workers (49) used 3-day
food records and found no significant correlation between
total energy or any macronutrient and serum adiponectin in a
group of 114 individuals. However, the difficulties in obtaining
reliable data on food intake by dietary interviews, records, or
diaries are well known (50–52).
Intervention studies have been restricted to acute postpran-
dial trials rather than investigating the longer-term effects of
feeding on fasting levels, as in our current experiment. These
postprandial trials have shown mixed results, yet the differ-
ences in the effect of feeding on postprandial adiponectin
cannot readily be explained by the type of meal provided.
Adiponectin both increased (15) and decreased (16) up to 4 h
after test meal, while there was no effect of feeding on serum
levels up to 6 h after test-meal in other trials (17–19,53). Some
studies probed the effects of a mixed meal (15,19,53) while
others investigated the effect of high-fat feeding per se (16–18).
An experiment carried out in our laboratory tested the effects
of a high-SFA meal vs. a low-SFA meal, and no effect of fat
quality on circulating adiponectin was seen (17).
We are aware of two trials that have examined the longer-
term effects of dietary fat on adiponectin in humans. In a
crossover trial, Berk and colleagues (20) fed a group of lean
and obese subjects a eucaloric diet comprising either 30% or
50% energy from fat each for a period of 7 days. Although cir-
culating adiponectin did not change in response to alterations
in dietary fat content over that period, there was a significant
correlation between the changes in adiponectin levels for low
fat to high fat with baseline insulin sensitivity. Bonnet and
colleagues (21) investigated the impact of an increased intake
of dietary ω-3 fatty acid (PUFA) on glucose metabolism and
adipokine concentration in a group of 20 healthy subjects over
a period of 10 weeks. Diets were supplemented with fatty fish
at least three times per week in addition to 20 g high-MUFA
rapeseed oil per day, and the results showed an increase in
dietary ω-3 fatty acid consumption to be associated with a
17% increase in circulating adiponectin when compared to
Of particular importance is the fact that in the current trial
body weight was kept constant and was strictly maintained
within the limits of ±1 kg during the two intervention arms.
This is critical because the size of adipose tissue depots is
known to be one of the key determinants of the concentra-
tion of circulating adiponectin. It is known that hormone
levels are decreased in obese humans (54), restored to nor-
mal levels after weight loss (55), and negatively correlated to
body fat and to waist-to-hip ratio (54). Any change in adipose
tissue mass, whether an increase or decrease, can alter total
adiponectin concentration. Circulating adiponectin is also
affected by disease state. Although participants in the current
trial were hyperlipidemic, they were taking no medication
for their hyperlipidemia or any other condition, nor had they
any current or previous history of CVD, T2DM, or any other
Adipose-derived pro-inflammatory cytokines, TNF-α and
IL-6, have been shown to be closely related to adiponectin
(56,57). In the current trial, no effect of alterations in dietary
fatty acid saturation was seen on these markers. A number of
large dietary supplement trials have investigated medium-term
alterations in fatty acid profile, and have revealed changes in
these inflammatory markers. Over an intervention period last-
ing 4–6 weeks, the marine-derived ω-3 PUFA eicosapentenoic
acid (20:5n−3) was shown to inhibit the production of TNF-α
and IL-6 (25,26). Although over periods of up to 12 weeks
(26,27), α-linolenic acid (18:3n−3) an ω-3 PUFA, derived
from plant sources such as flaxseed oil, may decrease TNF-α
and IL-6 at high doses (28), it appears to be less effective in
doing so than its marine-derived counter-parts. ώ-6 PUFAs
have been shown to be predominantly pro-inflammatory (24),
while healthy men (29) and individuals with T2DM (58) derive
anti-inflammatory effects from fats rich in MUFA fed over a
Changes in fatty acid saturation had no effect on circulating
C-RP in the current trial. In a cross-sectional study, a high-fat
diet that was predominantly ω-3 PUFA was shown to be nega-
tively associated with C-RP (59). Although C-RP has previ-
ously been shown to both increase (29) and decrease (30) in
response to medium-term (4–5 weeks) feeding, an interven-
tion study involving healthy men showed that conversion to
the Mediterranean diet for 90 days did not lead to an improve-
ment in C-RP (32) whereas in subjects with the metabolic syn-
drome such a dietary change lead to a decrease in C-RP after
2 years (31). Although weight loss (60) and physical activity
(61–63) are known to modulate C-RP, the effects of diet on this
acute-phase protein are inconclusive.
Diurnal changes in the release of leptin with a decline early
morning followed by a nighttime peak, where concentrations
are up to two times higher than nadir levels (64–67), compli-
cate the interpretation of circulating leptin data. In the current
Figure 3 Change from baseline in fasting leptin and ghrelin during
the 3-week highSFA:USFA (open circles) and lowSFA:USFA (closed circles)
treatments. Mean ± s.e.m.
trial, leptin did not change in response to fatty acid profile.
However, it might have been more appropriate to measure lep-
tin levels more frequently than simply an early morning fasting
sample at seven time-points across the two 3-week interven-
tion periods. Long-term variations in diet, when body fat
remains unchanged, have previously been shown not to alter
fasting leptin concentrations (33), and it has been suggested
that circulating leptin may be driven not by diet but rather by
total adipose mass (34).
Ghrelin is an anorectic hormone most abundant in the stom-
ach, the most important role of which seems to be the stimu-
lation of appetite and regulation of energy homeostatsis (68).
A number of trials have indicated that ghrelin responds post-
prandially to CHO feeding (69–72), although data suggest that
it may not respond in the same way to fat feeding (17,71,73).
There is little evidence, however, to indicate that this hormone
responds to longer-term feeding, especially alterations in fatty
acid profile. The lack of response of ghrelin to alterations in
fatty acid profile in our trial supports this idea.
In conclusion, little is known as yet about the effects of die-
tary fat intake and specifically fatty acid saturation on adipo-
kines such as adiponectin. Evidence from postprandial trials is
emerging such that this adipokine is insensitive to acute intake
of fat, but less is known about the effect of medium-term
feeding. There was no evidence from the current trial that an
increase in the SFA content of the diet significantly worsened
fasting serum adiponectin, TNF-α, IL-6, C-RP, leptin, or ghre-
lin in this group over the 3-week intervention period.
We thank Glyn Muir who ran the metabolic kitchen and prepared all
food for this trial. We are also grateful to Cathelijne Reincke for her
considerable assistance during her studentship. Also thanks are due to
Veronica Howsman, Santuri Rungan, Chao-Yuan Chen, Nicola Mohan,
Anna Mackey, Natasha Smith, and Sarah Thornber for their technical
assistance. Cynthia Tse provided laboratory and administrative support
throughout. We thank Shelia Bingham, MRC Cambridge, UK, for provision
of PABA-chek tablets. Thanks must also go to all participants who were
resident at the Human Nutrition and Metabolic Unit for several months
during this crossover trial.
The authors declared no conflict of interest.
© 2008 The Obesity Society
1. Spranger J, Kroke A, Möhlig M et al. Adiponectin and protection against
type 2 diabetes mellitus. Lancet 2003;361:226–228.
2. Hotta K, Funahashi T, Arita Y et al. Plasma concentrations of a novel,
adipose-specific protein, adiponectin, in type 2 diabetic patients.
Arterioscler Thromb Vasc Biol 2000;20:1595–1599.
3. Hu E, Liang P, Spiegelman BM. AdipoQ is a novel adipose-specific gene
dysregulated in obesity. J Biol Chem 1996;271:10697–10703.
4. Arita Y, Kihara S, Ouchi N et al. Paradoxical decrease of an
adipose-specific protein, adiponectin, in obesity. Biochem Biophys
Res Commun 1999;257:79–83.
5. Asayama K, Hayashibe H, Dobashi K et al. Decrease in serum adiponectin
level due to obesity and visceral fat accumulation in children. Obes Res
6. Matsubara M, Maruoka S, Katayose S. Decreased plasma adiponectin
concentrations in women with dyslipidemia. J Clin Endocrinol Metab
7. Nadler ST, Stoehr JP, Schueler KL et al. The expression of adipogenic genes
is decreased in obesity and diabetes mellitus. Proc Natl Acad Sci USA
8. Pellmé F, Smith U, Funahashi T et al. Circulating adiponectin levels are
reduced in nonobese but insulin-resistant first-degree relatives of type 2
diabetic patients. Diabetes 2003;52:1182–1186.
9. Lihn AS, Bruun JM, He G et al. Lower expression of adiponectin mRNA
in visceral adipose tissue in lean and obese subjects. Mol Cell Endocrinol
10. Fruebis J, Tsao TS, Javorschi S et al. Proteolytic cleavage product of
30-kDa adipocyte complement-related protein increases fatty acid oxidation
in muscle and causes weight loss in mice. Proc Natl Acad Sci USA
11. Yamauchi T, Kamon J, Waki H et al. The fat-derived hormone adiponectin
reverses insulin resistance associated with both lipoatrophy and obesity.
Nat Med 2001;7:941–946.
12. Grundy SM, Vega GL. Plasma cholesterol responsiveness to saturated fatty
acids. Am J Clin Nutr 1988;47:822–824.
13. Grundy SM, Denke MA. Dietary influences on serum lipids and lipoproteins.
J Lipid Res 1990;31:1149–1172.
14. Lindstrom J, Eriksson JG, Valle TT et al. Prevention of diabetes mellitus
in subjects with impaired glucose tolerance in the Finnish Diabetes
Prevention Study: results from a randomized clinical trial. J Am Soc Nephrol
15. English PJ, Coughlin SR, Hayden K, Malik IA, Wilding JP.
Plasma adiponectin increases postprandially in obese, but not in lean,
subjects. Obes Res 2003;11:839–844.
16. Esposito K, Nappo F, Giugliano F et al. Meal modulation of circulating
interleukin 18 and adiponectin concentrations in healthy subjects
and in patients with type 2 diabetes mellitus. Am J Clin Nutr
17. Poppitt SD, Keogh GF, Leahy FE et al. Postprandial effects of lipid loading
and fatty acid saturation on the adipose-derived cardioprotective peptide
adiponectin and pro-inflammatory mediators IL-6, TNF-α and CRP.
Nutrition, in press.
18. Peake PW, Kriketos AD, Denyer GS, Campbell LV, Charlesworth JA.
The postprandial response of adiponectin to a high-fat meal in normal
and insulin-resistant subjects. Int J Obes Relat Metab Disord
19. Karlsson FA, Engstrom BE, Lind L, Ohrvall M. No postprandial increase of
plasma adiponectin in obese subjects. Obes Res 2004;12:1031–1032;
author reply 1032–1034.
20. Berk ES, Kovera AJ, Boozer CN et al. Adiponectin levels during
low- and high-fat eucaloric diets in lean and obese women. Obes Res
21. Bonnet F, Guebre-Egziabher F, Rabasa R et al. Impact of an enrichment
of the diet in omega-3 fatty acids on glucose metabolism and plasma
adiponectin [abstr]. Obes Rev 2005;6:75.
22. Hauner H, Petruschke T, Russ M, Röhrig K, Eckel J. Effects of tumour
necrosis factor alpha (TNF alpha) on glucose transport and lipid metabolism
of newly-differentiated human fat cells in cell culture. Diabetologia
23. Ritchie DG. Interleukin 6 stimulates hepatic glucose release from prelabeled
glycogen pools. Am J Physiol 1990;258:E57–E64.
24. Kelley DS. Modulation of human immune and inflammatory responses by
dietary fatty acids. Nutrition 2001;17:669–673.
25. Endres S, Ghorbani R, Kelley VE et al. The effect of dietary supplementation
with n-3 polyunsaturated fatty acids on the synthesis of interleukin-1
and tumor necrosis factor by mononuclear cells. N Engl J Med
26. Caughey GE, Mantzioris E, Gibson RA, Cleland LG, James MJ. The effect
on human tumor necrosis factor alpha and interleukin 1 beta production of
diets enriched in n-3 fatty acids from vegetable oil or fish oil. Am J Clin Nutr
27. Thies F, Nebe-von-Caron G, Powell JR et al. Dietary supplementation with
gamma-linolenic acid or fish oil decreases T lymphocyte proliferation in
healthy older humans. J Nutr 2001;131:1918–1927.
28. Rallidis LS, Paschos G, Liakos GK et al. Dietary alpha-linolenic acid
decreases C-reactive protein, serum amyloid A and interleukin-6 in
dyslipidaemic patients. Atherosclerosis 2003;167:237–242.
29. Baer DJ, Judd JT, Clevidence BA, Tracy RP. Dietary fatty acids affect plasma
markers of inflammation in healthy men fed controlled diets: a randomized
crossover study. Am J Clin Nutr 2004;79:969–973.
8 Download full-text
30. Jenkins DJ, Kendall CW, Marchie A et al. Effects of a dietary portfolio of
cholesterol-lowering foods vs lovastatin on serum lipids and C-reactive
protein. JAMA 2003;290:502–510.
31. Esposito K, Marfella R, Ciotola M et al. Effect of a mediterranean-style diet
on endothelial dysfunction and markers of vascular inflammation in the
metabolic syndrome: a randomized trial. JAMA 2004;
32. Mezzano D. Distinctive effects of red wine and diet on haemostatic
cardiovascular risk factors. Biol Res 2004;37:217–224.
33. Havel PJ, Kasim-Karakas S, Mueller W et al. Relationship of plasma leptin
to plasma insulin and adiposity in normal weight and overweight women:
effects of dietary fat content and sustained weight loss. J Clin Endocrinol
34. Weigle DS, Duell PB, Connor WE et al. Effect of fasting, refeeding, and
dietary fat restriction on plasma leptin levels. J Clin Endocrinol Metab
35. Poppitt SD, Keogh GF, Mulvey TB et al. Lipid-lowering effects of a modified
butter-fat: a controlled intervention trial in healthy men. Eur J Clin Nutr
36. McDonald BE, Gerrard JM, Bruce VM, Corner EJ. Comparison of the effect
of canola oil and sunflower oil on plasma lipids and lipoproteins and on in
vivo thromboxane A2 and prostacyclin production in healthy young men.
Am J Clin Nutr 1989;50:1382–1388.
37. Schofield WN. Predicting basal metabolic rate, new standards and review of
previous work. Hum Nutr Clin Nutr 1985;39(suppl 1):5–41.
38. Bingham SA, Cummings JH. Urine nitrogen as an independent validatory
measure of dietary intake: a study of nitrogen balance in individuals
consuming their normal diet. Am J Clin Nutr 1985;42:1276–1289.
39. Johansson G, Callmer E, Gustafsson JA. Changing from a mixed meal diet
to a Scandanavian vegetarian diet: effects on nutrient intake, food choice,
meal pattern and cooking methods. Eur J Clin Nutr 1992;46:707–716.
40. Bingham S, Cummings JH. The use of 4-aminobenzoic acid as a marker
to validate the completeness of 24 h urine collections in man. Clin Sci
41. Pajvani UB, Du X, Combs TP et al. Structure-function studies of the
adipocyte-secreted hormone Acrp30/adiponectin. Implications fpr metabolic
regulation and bioactivity. J Biol Chem 2003;278:9073–9085.
42. Patel SD, Rajala MW, Rossetti L, Scherer PE, Shapiro L. Disulfide-dependent
multimeric assembly of resistin family hormones. Science
43. Tsao TS, Murrey HE, Hug C, Lee DH, Lodish HF. Oligomerization
state-dependent activation of NF-kappa B signaling pathway by
adipocyte complement-related protein of 30 kDa (Acrp30). J Biol Chem
44. Waki H, Yamauchi T, Kamon J et al. Impaired multimerization of human
adiponectin mutants associated with diabetes. Molecular structure and
multimer formation of adiponectin. J Biol Chem 2003;278:40352–40363.
45. Tsao TS, Lodish HF, Fruebis J. ACRP30, a new hormone controlling fat and
glucose metabolism. Eur J Pharmacol 2002;440:213–221.
46. Pajvani UB, Hawkins M, Combs TP et al. Complex distribution, not
absolute amount of adiponectin, correlates with thiazolidinedione-mediated
improvement in insulin sensitivity. J Biol Chem 2004;279:12152–12162.
47. Kobayashi H, Ouchi N, Kihara S et al. Selective suppression of endothelial
cell apoptosis by the high molecular weight form of adiponectin. Circ Res
48. Pischon T, Girman CJ, Rifai N, Hotamisligil GS, Rimm EB.
Association between dietary factors and plasma adiponectin
concentrations in men. Am J Clin Nutr 2005;81:780–786.
49. Yannakoulia M, Yiannakouris N, Bluher S et al. Body fat mass and
macronutrient intake in relation to circulating soluble leptin receptor,
free leptin index, adiponectin, and resistin concentrations in healthy humans.
J Clin Endocrinol Metab 2003;88:1730–1736.
50. Livingstone MB, Prentice AM, Strain JJ et al. Accuracy of weighed dietary
records in studies of diet and health. Br Med J 1990;300:708–712.
51. Livingstone MBE, Davies PSW, Prentice AM et al. Comparison of
simultaneous measures of energy intake and expenditure in children and
adolescents. Proc Nutr Soc 1991;50:15a.
52. Black AE, Jebb SA, Bingham SA. Validation of energy and protein intake
assessed by diet history and weighed record against energy expenditure
and 24-hour urinary nitrogen excretion. Proc Nutr Soc 1991;50:108A.
53. Imbeault P, Pomerleau M, Harper ME, Doucet E. Unchanged fasting and
postprandial adiponectin levels following a 4-day caloric restriction in young
healthy men. Clin Endocrinol (Oxf) 2004;60:429–433.
54. Weyer C, Funahashi T, Tanaka S et al. Hypoadiponectinemia in obesity
and type 2 diabetes: close association with insulin resistance and
hyperinsulinemia. J Clin Endocrinol Metab 2001;86:1930–1935.
55. Yang WS, Lee WJ, Funahashi T et al. Weight reduction increases plasma
levels of an adipose-derived anti-inflammatory protein, adiponectin. J Clin
Endocrinol Metab 2001;86:3815–3819.
56. Kern PA, Di Gregorio GB, Lu T, Rassouli N, Ranganathan G. Adiponectin
expression from human adipose tissue: relation to obesity, insulin resistance,
and tumor necrosis factor-alpha expression. Diabetes 2003;52:1779–1785.
57. Fasshauer M, Kralisch S, Klier M et al. Adiponectin gene expression
and secretion is inhibited by interleukin-6 in 3T3-L1 adipocytes.
Biochem Biophys Res Commun 2003;301:1045–1050.
58. Ros E. Dietary cis-monounsaturated fatty acids and metabolic control in
type 2 diabetes. Am J Clin Nutr 2003;78:617S–625S.
59. Lopez-Garcia E, Schulze MB, Manson JE et al. Consumption of (n-3) fatty
acids is related to plasma biomarkers of inflammation and endothelial
activation in women. J Nutr 2004;134:1806–1811.
60. Heilbronn LK, Noakes M, Clifton PM. Energy restriction and weight loss on
very-low-fat diets reduce C-reactive protein concentrations in obese, healthy
women. Arterioscler Thromb Vasc Biol 2001;21:968–970.
61. Mattusch F, Dufaux B, Heine O, Mertens I, Rost R. Reduction of the plasma
concentration of C-reactive protein following nine months of endurance
training. Int J Sports Med 2000;21:21–24.
62. Ford ES. Does exercise reduce inflammation? Physical activity and
C-reactive protein among U.S. adults. Epidemiology 2002;13:561–568.
63. Smith JK, Dykes R, Douglas JE, Krishnaswamy G, Berk S. Long-term
exercise and atherogenic activity of blood mononuclear cells in persons at
risk of developing ischemic heart disease. JAMA 1999;281:1722–1727.
64. Havel PJ, Townsend R, Chaump L, Teff K. High-fat meals reduce 24-h
circulating leptin concentrations in women. Diabetes 1999;48:334–341.
65. Sinha MK, Ohannesian JP, Heiman ML et al. Nocturnal rise of leptin in lean,
obese, and non-insulin-dependent diabetes mellitus subjects. J Clin Invest
66. Laughlin GA, Yen SS. Hypoleptinemia in women athletes: absence of a
diurnal rhythm with amenorrhea. J Clin Endocrinol Metab 1997;82:318–321.
67. Saad MF, Riad-Gabriel MG, Khan A et al. Diurnal and ultradian rhythmicity
of plasma leptin: effects of gender and adiposity. J Clin Endocrinol Metab
68. Peeters TL. Ghrelin: a new player in the control of gastrointestinal functions.
69. Tschöp M, Wawarta R, Riepl RL et al. Post-prandial decrease of circulating
human ghrelin levels. J Endocrinol Invest 2001;24:RC19–21.
70. Shiiya T, Nakanzato M, Mizuta M et al. Plasma ghrelin levels in lean
and obese humans and the effect of glucose on ghrelin secretion.
J Clin Endocrinol Metab 2002;87:240–244.
71. Monteleone P, Bencivenga R, Longobardi N, Serritella C, Maj M.
Differential responses of circulating ghrelin to high-fat or high-carbohydrate
meal in healthy women. J Clin Endocrinol Metab 2003;88:5510–5514.
72. Greenman Y, Golani N, Gilad S et al. Ghrelin secretion is modulated
in a nutrient- and gender-specific manner. Clin Endocrinol (Oxf)
73. Erdmann J, Töpsch R, Lippl F, Gussmann P, Schusdziarra V.
Postprandial response of plasma ghrelin levels to various test meals in
relation to food intake, plasma insulin, and glucose. J Clin Endocrinol Metab