J. Biomedical Science and Engineering, 2013, 6, 144-151 JBiSE
http://dx.doi.org/10.4236/jbise.2013.62018 Published Online February 2013 (http://www.scirp.org/journal/jbise/)
Heart and skeletal muscle insulin resistance but not
myocardial blood flow reserve could be related to chronic
use of thiazolidione in patients with type-2 diabetes*
Ikuo Yokoyama1#, Toshiyuki Moritan2, Yusuke Inoue3
1Department of Cardiovascular Medicine, Clinical Research Center, Sanno Hospital and Sanno Medical Center, International Uni-
versity of Health and Welfare, Tokyo, Japan
2Department of Clinical Engineering, Faculty of Medical Engineering, Suzuka University of Medical Science, Suzuka, Japan
3Department of Radiology, Graduate School of Medicine, Kitasato University, Sagamihara, Japan
Received 3 November 2012; revised 2 December 2012; accepted 14 December 2012
ABSTRACT
Heart and skeletal muscle insulin resistance and ab-
normal myocardial flow reserve (MFR) occurs in pa-
tients with type-II diabetes. Improvement of heart
and skeletal muscle insulin resistance with rosiglita-
zone use over 16 weeks have been reported. However,
it is not clear whether chronic use of troglitazone can
improve heart and skeletal muscle insulin resistance
and MFR. Materials and Methods: To test the hypo-
thesis whether effects of troglitazone on heart and
skeletal muscle insulin resistance and MFR in pa-
tients with type-II diabetes, rest and dipyridamole
stress perfusion positron emission tomography (PET)
with 13N-ammonia and heart and skeletal muscle
18FDG PET scans under insulin clamping were un-de r-
taken before and 12 month after the initiation of tro-
glitazone therapy (400 mg/day) in 23 patients with
type-II diabetes. Twenty patients with type-II diabetes
without CAD and without medications were served as
controls. In controls, any medications were not added
from the first PET study and 12 months after the
second PET study. Results: Baseline myocardial bloo d
flow (MBF) was comparable before and after the tro-
glitazone group as was the controls. MBF during
dipyridamole administration (0.56 mg/min/kg) was
not significantly improved in troglitazone group and
controls. MFR was not improved in tro glitazone gro u p
and controls. In troglitazone group, whole body glu-
cose disposal rate (GDR; µmole/min/kg) significantly
improved (pre; 19.0 ± 9.55, post; 28.7 ± 15.3 , p < 0.05)
as did the skeletal muscle glucose utilization rate
(SMGU (µmole/min/kg); pre; 20.3 ± 12.0, post; 34.8 ±
10.6, p < 0.05) and the myocardial glucose utilization
rate (MGU (µmole/min/kg); pre; 339.7 ± 105.2 vs.
post; 410.0 ± 240.0, p < 0.05). GDR, SMGU and MGU
were unchanged in controls. Conclusions: Troglita-
zone can improve heart and skeletal muscle insulin
resistance in patients with type-II diabetes but not
MFR showing that co-existence of heart and skeletal
mus cle insulin resistance is implicated in patients with
type-II diabetes and impaired MFR is uncoupled with
insulin resistance in the whole body and heart and
skeletal muscle in patients with type-II diabetes.
Keywords: Insulin Resistance; Myocardial Insu lin
Resistance; Glucose; FDG; PET; Type-II Diabetes; Flow
Reserve
1. INTRODUCTION
Insulin resistance is defined as an impaired glucose
utilization response to the stimulatory effect of insulin
and has been recognized to have common essential role
in a variety of metabolic diseases including type-II dia-
betes mellitus [1]. Since insulin resistance is closely re-
lated to metabolic syndrome and an occurrence of coro-
nary artery disease (CAD) [2], investigation of the patho-
physiology of and effects of certain drugs on insulin re-
sistance is important for an early prevention of CAD.
Positron emission tomography (PET) allows in vivo
quantitative analysis of tissue glucose metabolism using
[18F]2-fluoro-2-deoxy-D-glucose (18FDG) and myocar-
dial blood flow (MBF) with 13N-ammonia or 15O-H2O.
PET can be used to evaluate insulin resistance and myo-
cardial perfusion abnormalities in subjects highly at risk
for CAD. Reduced skeletal muscle glucose utilization
(SMGU) under hyperinsulinemic euglycemic clamping,
*This study finished on March 2000, be caus e tr oglita zone was forbidden to
treat diabetes mellitus and was withd rawn from the market due to seri-
ous side effects about liver function. Patients who were treated with
troglitazone received no further treatment with troglitazone after March
2000. Therefore all data collection using troglitazone were finished
before Marc h 1.
#Corresponding author.

I. Yokoyama et al. / J. Biomedical Science and Engineering 6 (2013) 144-151 145
implying insulin resistance in the skeletal muscle, has
been reported in patients with type-II diabetes [3-5]. Re-
duced myocardial flow reserve (MFR) in patients with
type-II diabetes without evidence of CAD [6-8] have
also been reported suggesting an existence of microvas-
cular abnormality in the heart.
Effects of rosiglitazone on myocardial insulin resis-
tance in patients with type-II diabetes with CAD have
been reported [9]. Effects of pioglitazone onthe improve-
ment of whole body insulin resistance and myocardial
insulin resistance but not MFR have been reported in
patients with familial mixed combined hyperlipidemia
[10]. However, another investigation has shown that acu te
use of troglitazone within 12 weeks failed to improve
myocardial insulin resistance although skeletal muscle
and whole body insulin resistance were improved in the
same study patients with type-II diabetes without CAD
[11]. In addition, effects of thiazolidiones on both myo-
cardial insulin resistance and MFRin patients with type-
II diabetes with or witho ut CAD are not studied simulta-
neously. There has been no examination as to whether
thiazolidiones can improve heart and skeletal muscle
insulin resistance as well as microvascular dysfunction in
patients with type-II diabetes during the same study pe-
riods. Relatively long term (chronic) effects of thiazo lid-
iones on heart and skeletal muscle insulin resistance and
MFRmore than 16 weeks have not been done.
This study aimed to study chronic effects of troglita-
zone therapy over 12 months, on heart and skeletal mus-
cle insulin resistance and MFR and to certify MFR in
patients with type-II diabetes without CAD is uncoupled
with a recovery of insulin resistance in the whole body
and heart andskeletal muscle.
2. MATERIALS AND METHODS
2.1. Study Design
Study Populations
We recruited asymptomatic non-medicated patients with
type-II diabetes from the local clinics and local health
service centers. The diabetic patients satisfied the criteria
of having a fasting glucose level at admission (within
few week before the study) grater than 140 mg/dl and
hemoglobin A1c (HbA1c) greater than 7.0%. All study
subjects were highly negligible for CAD by using tread-
mill test, rest echocardiography and rest and dipyrida-
mole stress static myocardial perfusion PET.
Twenty-seven patients with type-II diabetes (21 males,
6 females) were recruited for troglitazone therapy group
(400 mg/day). Twenty patients with type-II diabetes (16
males, 4 females, HbA1c; 8.0 ± 1.4 percent, age; 52.4 ±
11.2) who had not been and would not be treated with
any kinds of medicines for diabetes during the study pe-
riods were served as controls from our independent data
base to this study. Treadmill test and rest echocardiogra-
phy were undertaken before the initiation of the study as
the initial screening test for CAD and heart diseases.
Heart and skeletal muscle 18FDG PET and 13N-ammonia
myocardial perfusion PET were undertaken just at the
time of initiation of this study and 12 months after the
initiation of this study in both groups. Static myocardial
perfusion PET images just at the time of the initiation of
this study was used to exclude CAD. In both groups,
conventional diet and exercise therapy (approximately
10,000 steps/day) wer e continued during the who le study
periods of 12 months between the first PET scan and the
second PET scan, therapies were not changed. In case of
any kinds of emergency and patient’s request to drop out
from this study, the study protocol was stopped. Physical
examination and blood check test was undertaken every
month during the study periods. Finally, 23 patients with
type-II diabetes (mean age 52.9 ± 12.1 yr; 18 males, 5
females, HbA1c; 8.18 ± 1.94 percent, reference value 4.0
- 5.8 percent) finished all of the study protocol using
troglitazone therapy and PET scan (3 were excluded due
to CAD and one was stopped the protocol due to non-
fetal side effect of liver damage). General characteristics
were shown in Table 1.
Before this study, we informed all subjects of the na-
ture of this study, after which they agreed to participate
in the protocol that was approved by the local Ethics
Committee.
2.2. Positron Emission Tomography (PET)
Myocardial 13N-ammonia images and heart and skeletal
muscle 18FDG images were obtained using a Headtome
IV PET scanner (Shimadzu Corp., Kyoto, Japan). This
Table 1. General characteristics of study patients with type-II
diabetes who were treated with troglitazone therapy.
Pre Post P-value
N (M/F) 23 (18/5) 23 (18/5) N/A
Body weight 61.5 ± 8.8 61.8 ± 8.4 NS
Height 162 ± 7.78 162 ± 7.69 NS
SBP 134 ± 12.32 129 ± 6.21 NS
DBP 73.2 ± 10.2 72.5 ± 6.55 NS
TC 203 ± 19.3 197 ± 20 .1 NS
HDL 53.7 ± 14.7 53.7 ± 13.3 NS
Triglycerid es 161 ± 74.0 153 ± 5 8 .0 NS
LDL 117 ± 26.1 112 ± 20.9 NS
HbA1c 8.19 ± 1.18 7.18 ± 0.90 <0.01
FBS 202 ± 47.3 165.2 ± 40.9 <0.01
FFA 0.72 ± 0.29 0.55 ± 0.24 <0.05
N: Number of patients; M: Male; F: Female; N/A: Not applicable; BW:
Body weight (kg); HT: Height (cm); BMI: Body mass index (kg/m2); BPS:
Systolic blood pressure (mmHg); BPD: Diastolic blood pressure (mmHg);
HbA1c: Hemoglobin A1c (percent (%)); FBS: Fasting plasma blood glucose
concentration (mg/dL); TC: Total cholesterol (mg/dL); HDL: High density
lipoprotein cholesterol; TG: Triglycerides (mg/dL); LDL: Low density lipo-
protein cho l esterol (mg/dL); NS: Not s ignificant.

I. Yokoyama et al. / J. Biomedical Science and Engineering 6 (2013) 144-151 146
PET scanner has seven imaging planes; in-plane resolu-
tion is 4.5 mm at full width at half maximum (FWHM)
and the z-axial resolu tion is 9.5 mm at FWHM. Effective
in-plane resolution was 7 mm after using a smoothing
filter. The sensitivities of the Headtome IV scanners are
14 and 24 kcps (kiro counts/seco nds) for direct and cross
planes, respectively. After acquiring transmission data for
a period of 8 min to correct for photon attenuation prior
to obtaining the PET emission images, 15 - 20 mCi of
13N-ammonia were injected and dynamic PET scanning
was performed over 2 min. Fifty-five min after the injec-
tion of 13N-ammonia to allow for decay of the radioactiv-
ity of the isotope, dipyridamole (0.56 mg/kg) was ad-
ministered intravenously over a 4-min period. Five min
after the end of the dipyridamole infusion, 15 - 20 mCi
of 13N-ammonia were injected again, and, exactly at the
same time, a second dynamic PET scan was performed
for 2 min. The dynamic PET scan was performed every
15 sec (8 times) during the 2-min period. Dynamic data
were obtained for seven slices. After the dynamic scans,
static 13N-ammonia images were obtained from the PET
study over 10 minutes and analyzed visually by three
independent specialists who had no other information on
the patients. Single channel ECG monitoring in limb
leads was performed during the PET study, because a full
ECG was not satisfactory due to a lack of a precordial
record as a result of a technical difficulty. There was thus
the possibility that the ECG data were unreliable and
they were therefore not used.
2.3. Determination of MBF
Regional MBF was calculated according to the two-
compartment 13N-ammonia tracer kinetic model [12].
The time activity curve of the left ventricular cavity was
used as an input function. The tracer spillover was cor-
rected by least square non-linear regression analysis to
calculate MBF with the assumption that both myocardial
and left ventricular radioactivity influenced one another.
Details can be found in our previously published paper
[6]. As reported in our previous paper, regions of interest
(ROIs) were placed at the septum, anterior wall, lateral
wall and infero-posterior wall on transaxial images. To
obtain input function, ROIs were placed on the left ven-
tricular cavity of each slice. We determined the MFR
value as follows:
MFRMBF during dipyridamoleMBFat rest
2.4. Preparation of 18FDG
18F was synthesized using the Cypris model 370 cyclo-
tron (Sumitomo JYUKI Industries, Ltd., Kyoto, Japan),
and 18FDG was synthesized with an automated system
based on the method reported by Ehrenkaufer et al. [13].
Radiochemical purity was more than 95%.
Acquisition of Myocardial Me tabolic Images
There was about 100 min interval after the acquisition of
13N-ammonia images during dipyridamole administration
to allow for decay of radioactivity of 13N-ammonia dur-
ing which time the blood glucose concentration was kept
constant at the level of about 90 - 105 mg/dl using hy-
perinsulinemic euglycemic clamp technique which was
initiated at the end of data acquisition of MBF during
dipyridamole administration. Then a second transmission
scan for a period of 10 min was undertaken and we in-
jected 18FDG (5 - 10 mCi) and collected dynamic PET
data for 1 hour and 45 seconds. During this interval, we
obtained 19 dynamic scans using the following protocol:
five 15-sec, three 30-sec, four 120-sec, four 300-sec and
three 600-sec scans.
2.5. Quantification of MGU and SMGU to
Estimate Heart and Skeletal Muscle Insulin
Resistance
The amount of glucose metabolized by various thoracic
organs (heart and skeletalmuscle of lumbar region) was
determined by calculating the tissue glucose utilization
rate. Following the method previously reported by Ohta ke
et al., [14] we obtained input function from the time ac-
tivity curve of the descending aorta corrected by seven
venous blood samplings. Using the input function, we
determined


13 2 3
kk kk by Patlak graphic analy-
sis [15]. And calculated both the MGU and SMGU by
substituting


13 2 3
kk kk in the following equa-
tion.
  
18
13 2 3123
TissueFDG uptake rate
BGBGBG3 LCkk kk  
k1 and k2 and k3 were rate constants of the following
chemical formula.
 

1
2
3
4
GluSGluskelt. M
Glu- 6-phosphateskelt. M
k
k
k
k




k4 is assumed to be zero in the myocardium and skeletal
muscle. BG1, BG2 and BG3 were serum glucose concen-
trations during the dynamic PET scan using 18FDG as
shown. S means serum and T means tissue. LC stands for
Lumped Constant, which was calculated to be 1.0 in
myocardium [16] and 1.2 in skeletal muscle cells, as re-
ported in human studies [17].
All PET data were corrected for dead time effects to
reduce error to less than 1%. To avoid the influence of
the partial volume effect associated with the object's size,
recovery coefficients (RC) obtained from experimental
phantom studies in our laboratory were used. The RC
was 0.8 when myocardial wall thickness was 10 mm. For
the correction of the partial volume effect, specialists in

I. Yokoyama et al. / J. Biomedical Science and Engineering 6 (2013) 144-151 147
our hospital measured wall thickness with 2-dimmen-
sional echocardiography. The RC was taken into consid-
eration in our program to measure the tissue glucose
utilization rate.
We obtained the MGU from the transaxial images. The
amount of MGU was determined by averaging the above
values. We also determined the SMGU from the lumbar
muscleaccording to a previously validated method [18].
To calculate the tissue glucose utilization rate and
MBF, we used the SUN Ultra-30 high-speed image
processing system (SUN Microsystems Japan Co., Ltd.,
Tokyo, Japan) with “Dir. View” software (Asahi Kasei
Information System Co., Ltd. Tokyo, Japan).
Whole body insulin resistance was determined by ob-
taining the whole body glucose disposal rate (GDR) dur-
ing hyperinsulinemic euglycemic clamping (mg/min/kg)
using a previously reported method [7]. Twelve months
after the initiation of troglitazone therapy, a second PET
scan, including 13N-ammonia resting and stress perfusion
imaging and heart and skeletal muscle 18FDG PET scan,
was undertaken to assess the effect of troglitazone on
insulin resistance in the whole body as well as in heart
and skeletal muscle and on MFR.
2.6. Power Calculation and Statistical Methods
From our reference data on PET flow measurement, the
estimated levels of myocardial blood flow during
dipyridamole stress are 250 ± 100 mL/min/100 g in con-
trol subjects and 180 ± 51.0 mL/min/100 g in the patients
with type-II diabetes mellitus. Assuming that troglita-
zonetherapy will increasemyocardial blood flow in the
patients with type-II diabetes mellitus to the similar leve l
in the control subjects, to detect statistically significant
differences with 80% power and with α = 0.05, a total of
23 patients is required as p = 0.033 with 10% dropout
patients. Values are expressed as the mean ± standard
deviation. Data before and after therapy were compared
by the paired t-test. Differences between two groups
were examined by the unpaired t-test or Mann-Whitney
test according to the distribution of measurement pa-
rameters. Differences between three groups were tested
by the multiple comparison method after analysis of
variance (ANOVA). A p value of less than 0.05 was con-
sidered significant.
3. RESULTS
3.1. GDR, SMGU and MGU
GDR (µmole/min/kg) significantly improved after the
troglitazone ther apy group (pr e; 19.0 ± 9.55, post; 28.7 ±
15.3, p < 0.05) as did the SMGU (pre; 20.3 ± 12.0
µmole/min/kg, post; 34.8 ± 10.6 µmole/min/kg, p < 0.05).
MGU also significantly improved after the troglitazone
therapy (339.7 ± 105.2 µmole/min/kg (pre) vs. 410.0 ±
240.0 µmole/min/kg (post), p < 0.05).
In control group, GDR (pre; 19.8 ± 9,28, post; 20.5 ±
12.9, p = NS), SMGU (pre; 21.0 ± 12.2 µmole/min/kg,
post; 23.4 ± 11.8 µmole/min/kg, p = NS), MGU were not
significantly changed 12 months after the first PET scan
(341.5 ± 116.3 µmole/min/kg (pre) vs. 358.2 ± 219.5
µmole/min/kg (p ost), p = NS).
3.2. Baseline MBF and MBF during
Dipyridamole Administration and MFR
In the patients with type-II diabetes the baseline MBF
(ml/min/100 g) was comparable before and after the tro-
glitazone ther apy (pre: 77.6 ± 11.6 vs. post: 74.5 ± 9.62).
However, MBF-DP (178 ± 50.9) was not significantly
improved (184 ± 61.2, p = NS) after the troglitazone
therapy. MFR was not improved by troglitazone (base-
line 2.23 ± 0.84 vs. p ost-therapy 2.27 ± 0. 9 0, p = NS).
In control group, the baseline MBF (ml/min/100 g)
was comparable between the first PET scan and the sec-
ond PET scan (first: 78.5 ± 12.4 vs. the second: 75.9 ±
10.5). MBF-DP in the patients with type-II diabetes (183
± 54.8) at the time of first PET scan was not significantly
changed 12 months after the first PET scan (180 ± 65.1,
p = NS). MFR at the time of first PET scan was not
changed 12 months after the second PET scan (first 2.29
± 0.81 vs. the secon d 2.37 ± 0.94, p = NS).
3.3. Serum Glucose Concentration
Plasma fasting glucose concentration before therapy (8 .68
± 1.87 µmole/liter) was significantly reduced after ther-
apy (7.41 ± 2.21 µmole/liter, p < 0.01). The average se-
rum glucose concentration in patients with type-II diabe-
tes during insulin clamping was (5.0 8 ± 0.68 µmole/liter).
In controls, plasma fasting glucose concentration (8.68 ±
1.87 µmole/liter) at the time of first PET scan was not
changed 12 months after the first PET scan (7.41 ± 2.21
µmole/liter, p < 0.01). The average serum glucose con-
centration in controls during insulin clamping was (5.08
± 0.68 µmole/lite r ).
3.4. Serum Insulin Concentration
Serum insulin co ncentration d uring in sulin clamping was
comparable between pre therapy (59.0 ± 17.2 µU/ml)
and post therap y (66.3 ± 38.0 µU/ml). There was no sig-
nificant difference between the serum insulin concentra-
tion at the beginning and at the end of the dynamic PET
scan. Plasma fasting insulin concentration in patients
with type-II diabetes (8.0 ± 3.1 µU/ml) before therapy
tended to be decreased compared with pre-therapy values
after therapy (7.5 ± 4.1 µU/ml), but remained signifi-
cantly higher than values for control subjects. There was

I. Yokoyama et al. / J. Biomedical Science and Engineering 6 (2013) 144-151 148
no significant relationship between MGU and plasma
insulin concentration in th e patients with type-II diabetes,
as well as between SMGU and plasma insulin concentra-
tion.
3.5. Serum Free Fatty Acid Concentration (FFA)
In the patients with type-II diabetes with troglitazone
therapy, serum FFA concentration during insulin clamp-
ing (0.72 ± 0.29 mEq/liter) was significantly reduced
after therapy (0.55 ± 0.24 mEq/liter, p < 0.05).
In control group, serum FFA concentration during in-
sulin clamping was not changed after the therapy (pre;
0.78 ± 0.29 mEq/liter post; 0.75 ± 0.35 mEq/liter, p =
NS).
4. DISCUSSION
4.1. General Discussion of Whole Results
This study investigated effects of chronic use of troglita-
zone on heart and skeletal muscle and whole body insu-
lin resistance and myocardial perfusion in patients with
type-II diabetes without CAD during the same study pe-
riods for the first time. Data were compared with those
who were non-medicated type-II diabetes without CAD.
SMGU and GDR were significantly improved 12 months
after the troglitazone therapy in patients with type-II
diabetes without CAD. These resultsare consistent with
those in vitro findings [19] and our previous study of
relatively short-term therapy over 3 months in vivo [20].
On the other hand, in control group,SMGU and GDR
were not changed 12 months after the first PET scan.
These results showed that chronic troglitazone therapy
could also preserve effect to improve the insulin resis-
tance in the skeletal muscle and whole body in patients
with type-II diabetes without CAD. In addition, MGU
was also improved significantly i n t he troglit azone therapy
group but such improvement was not seen in control
group. These results highly suggest that chronic use of
troglitazone can improve insulin resistance in both the
heart and skeletal muscle and whole body in patients
with type-II diabetes without CAD. Similar investigation
has been done in patients with type-II diabetes with CAD
using rosiglitazone [9]. In the report, effects of rosiglita-
zone on the heart and skeletal muscle insulin resistance
have been investigated over 16 weeks (relatively short
term periods of therapy comparing with our present
study) in patients with type-II diabetes with CAD [9].
Naoumova et al. reported that pioglitazone improved
heart and whole body insulin resistance but not MFR in
patients with familial combined hyperlipidemia but not
type-II diabetes [10]. Whereas Yokoyama et al. reported
that troglitazone therapy within 12 weeks improved ske-
letal muscle and whole body insulin resistance nut bot in
myocardial insulin resistance [11]. Therefore, it takes
long time to improve myocardial insulin resistance with
troglitazone in type-II diabetes.
4.2. Influence of a Co-Existence of CAD on
Myocardial Insulin Resistance in Type-II
Diabetes
No significant difference in MGU has been observed
between normal remote myocardial segments in patients
with CAD and type-II diabetes and normal remote myo-
cardial segments in patients with CAD without type-II
diabetes [21]. So it should be needed to negate the influ-
ence of a co-existence of CAD on the thiazolidione’s
response to myocardial insulin resistance. Whereas, it
has not been clear whether myocardial insulin resistance
could be altered by aco-existence of CAD in patients
with type-II diabetes. Therefore, effects of thiazolidione s
on heart and skeletal muscle insulin resistance in patien ts
with type II-diabetes without CAD has been remain un-
certain. In add ition, effects of any kinds of thiazo lidiones
over relatively long-term therapy periods (12 months) on
heart and skeletal muscle and whole body insulin resis-
tance in patients with type-II diabetes without CAD have
not been studied. As results of this study, relatively long
term therapy of troglitazone can improve both whole
body insulin resistance and myocardial insulin resistance
significantly in patients with type-II diabetes without
CAD. Since troglitazone is one of thiazolidionenes which
act as an insulin sensitizer, our results strongly support
that both heart and skeletal muscle and whole body insu-
lin resistance coexists in patients with type-II diabetes
without CAD. Furthermore, our results support such re-
ports that showed an existence of myocardial insulin
resistance in patients with type-II diabetes who also had
skeletal muscle and whole body insulin resistance [3-5].
The plasma FFA concentration was reported to be re-
lated to insulin resistance in patients with type-II diabe-
tes [22,23]. Enhancement of insulin stimulated suppres-
sive effect to decrease FFA via PPAR-gamma agonists
has been reported [24]. In our current study, the plasma
FFA concentration during insulin clamping was signifi-
cantly decreased 12 months after the troglitazone therapy
and both heart and skeletal muscle glucose utilization
and GDR weresignificantly increased in the troglitazone
therapy group however no changes were seen in the con-
trol group. These results indicate that the FFA con centra-
tion is a factor in the improvement of insulin resistance
in both the whole body and the heart and skeletal muscle
after the administration of troglitazone. An existence of
glucose-FFA cycle (so-called Randle’s cycle) has been
reported in human heart and skeletal muscle [25]. Our
current results suggest that troglitazone improves insulin
resistance in the heart and skeletal muscle and the whole

I. Yokoyama et al. / J. Biomedical Science and Engineering 6 (2013) 144-151 149
body inpatients with type-IIdiabetes withou t CAD via an
enhancement of the Randle’s cycle.
Another possible mechanisms to increase SMGU, MGU
and GDR by troglitazone use over 12 months could be
thought as follo ws. An activation of the pero xisome pro-
liferator-activated receptor (PPAR-γ) in adipocytes [26],
an increase in the number of small adipocytes and inac-
tivation of tumor necrotizing factor-a (TNF-α) via the
activation of PPAR-γ in adipocytes [26] can be factors.
Up-regulation of gene expression of PPAR-γ in skeletal
muscle cells by troglitazone may be another mechanism
for the improvement of SMGU in patients with type-II
diabetes. However, since adipocytes scarcely exist in the
heart, this discussion would not be applied to mecha-
nisms of improvement in MGU with troglitazone. Fur-
therinvestigation shou ld be required to improv e heart and
skeletal muscle insulin resistance in patien ts with type-II
diabetes without CAD after troglitazone therapy.
4.3. Troglitazone Did Not Improve MFR in
Patients with Type-II Diabetes without CAD
In this study, MFR was not improved in troglitazone
therapy group, contrasting to the results of an improve-
ment of insulin resistance in the whole body and heart
and skeletal muscle. In addition, MFR, MGU, SMGU
and GDR were not improved in the control group. Those
findings led to the expectation that the recovery of MFR
would not be related to the improvement of insulin resis-
tance not only in the whole body but also in heart and
skeletal muscle after troglitazone in patients with type-II
diabetes.On the other hand, glycemic control has been
implicated to be related to the reduced MFR in patients
with type II diabetes [6-8], but not to the whole body
insulin resistance [7,8]. Therefore, our current results that
troglitazone did not improve MFR in patients with
type-II diabetes but improved only insulin resistance in
the whole body highly support above studies. Moreover,
our present results have shown that the improvement of
heart or skeletal muscle insulin resistance does not lead
to recovery of MFR in patients with type-II diabetes. In
general, a reduced MFR is thought to be principally due
to a structural alteration in the microvascular bed surface
area due to micro embolism and/or atherosclerosis or
both. Contrary to anti-hyperlipidemic therapies which
have been reported to improve MFR in patients with hy-
perlipidemia without CAD [27-29] and those with CAD,
[30,31] anti-diabetic therap ies do not cure hyperglycemia
and/or insulin resistance completely and the deleterious
effects of diabetes on vessels might progress gradually
day by day. These factors can be a reason in our result as
to the no effect of troglitazone therapy on the r ecovery of
MFR in type-II diabetes. In patients with CAD and hy-
per-LDL cholesterolemia,delayed response of improve-
ment of MFRin response to lipid-lowering therapy with
fluvastatinhas been reported [31]. It has also been re-
ported that reduced MFR in patients with hypercholes-
terolemiaand CAD was improved by simvastatin but not
by pravastatin [29]. Therefore, kinds of drugs and dura-
tion of therapy can also be factors for the improvement
of MFR. Our current results support the knowledge that
abnormal coronary microvascular function is mainly
related to glycemic control in patients with type-II dia-
betes but not to insulin resistance [7,8]. These results
suggested that much stronger and stable effects on hy-
perglycemia and much longer therapeutic periods should
be required to improve MFR in patients with type-II dia-
betes rather than modest improvements in glycemic con-
trol and insulin resistance. It has been reported that ab-
normal microvascular function in patients with type-II
diabetes should be un-resolved important question to
answer [32]. Much stronger evidence should be required
how to treat abnormal microvascular dysfunction in pa-
tients with type-II diabetes.
4.4. Quantitative Analysis of Heart and Skeletal
Muscle Glucose Utilization Using PET and
18FDG
The significant improvement of both SMGU and GDR
indicates the validity of the PET quantitative method to
measure SMGU. However, previous animal experimental
studies suggested that the myocardial Lumped Constant,
which strands for the difference between glucose and
18FDG, is not constant [33,34]. Hariharan et al. only
showed that the bolus injection of insulin (but not insulin
clamping) did not increase myocardial 18FDG uptake,
whereas myocardial glucose uptake was increased spon-
taneously when a resected beating mouse heart was per-
fused by Krebs Henselite buffer [33]. However, clinical
myocardial 18FDG PET studies have shown that insulin
clamping effectively increased myocardial 18FDG uptake
in both normal contro ls and patients with type II diab etes
[3-5]. The method used in the animal studies [33,34]
cannot be applied to human studies. Furthermore, Ng et
al. have revealed that 18FDG uptake quite linearly corre-
lated with myocardial glucose uptake [34]. Therefore,
these animal experimental findings can only suggest that
MGU cannot be measured by PET and 18FDG after bolus
injection of insulin but they did not determine if MGU
can be measured byPET and 18FDG under hyperinsu-
linemic euglycemic clamping.
In this study, we determined SMGU from the lumber
muscle, because with this approach both MGU and
SMGU can be measured just at the same timewith nearly
the same accuracy as that of thigh muscle [18]. In the
manuscript of this method, a significant positive rela-
tionship between thigh muscle SMGU and lumbar mus-

I. Yokoyama et al. / J. Biomedical Science and Engineering 6 (2013) 144-151 150
cle SMGU and no significant difference in SMGU value
between thigh muscle and lumbar muscle have been
shown [18]. Therefore, determination of SMGU from the
lumbar muscle can be considered to be reliable. Orienta-
tion of lumber muscle can be determined with static
18FDG images after the dynamic study by changing the
color scale appropriately, so there is no problem as to the
placement of ROI. Even when orientation of lumbar
muscle is difficult to determine only by PET, X-ray CT is
very helpful to the ROI placement. Several reports cite
measurement of SMGU from the femoral muscle. [4,5,
11,25,35] However, this approach requires additional
data acquisition time over 22 - 30 min after the PET scan
of thoracic region, including frequent arterial or arterial-
ized or venous blood samplings to obtain input function
and dynamic data of femoral muscle 18FD G u pt ake.
4.5. Study Limitation
Since this study is open trial and not a placebo-control
double blind study, grade of evidences obtained from this
study would be under estimated. Because of serious liver
damage of troglitazone, we did not make such strict trial
and disappeared from the market. These are the l imit atio n
of this study.
5. CONCLUSION
Chronic troglitazone use can improve heart and skeletal
muscle insulin resistance similarly but not MFR in pa-
tients with type-II diabetes. These results lead an idea
that myocardial insulin resistance co-exists with skeletal
muscle and whole body insulin resistance in patients
with type-II diabetes without CAD. No change of MFR
after troglitazone therapy indicated that MFR was un-
coupled with the recovery of insulin resistance in the
whole body and heart and skeletal muscle in patients
with type-II diabetes without CAD.
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