pubs.acs.org/JAFC Published on Web 09/13/2010
©2010 American Chemical Society
J. Agric. Food Chem. 2010, 58, 10347–10355
Cardiac Contractile Dysfunction and Apoptosis
in Streptozotocin-Induced Diabetic Rats Are Ameliorated
by Garlic Oil Supplementation
HUI WEN LIU,2CHONG-KUEI LII,3DA-TIAN BAU,b,]PEI-MIN CHAO,3
AND WEI-WEN KUO*,2
†Department of Physical Therapy and Graduate Institute of Rehabilitation Science, China Medical
University, 91 Hsueh-ShihRoad, Taichung 404, Taiwan,‡Institute of Biochemistry and Biotechnology,
ChungShanMedicalUniversity, 110,Sec. 1,Chien-Kuo N. Road,Taichung402,Taiwan,§Departmentof
Biochemistry, School of Medicine, Chung Shan Medical University, 110, Sec. 1, Chien-Kuo N. Road,
Taichung 402, Taiwan,#Clinical Laboratory, Chung Shan Medical University Hospital, Taichung,
Taiwan,^Division of Cardiology, Armed Force Taichung General Hospital, No. 348, Sec. 2, Chungshan
Road, Taiping City, Taichung 411, Taiwan,XInstitute of Nutrition Science, Chun Shan Medical
University, 110, Sec. 1, Chien-Kuo N. Road, Taichung, Taiwan,2Department of Biological Science and
Technology, China Medical University, 91 Hsueh-Shih Road, Taichung, Taiwan,3Institute of Nutrition,
China Medical University, 91 Hsueh-Shih Road, Taichung 404, Taiwan,bGraduate Institute of Chinese
Medical Science, China Medical University, 91 Hsueh-Shih Road, Taichung, Taiwan, and]Terry Fox
Cancer Research Laboratory, China Medical University Hospital, 2 Yuh-Der Road, Taichung 404,
These authors contributed equally to this work.
Previous studies have suggested that garlic oil could protect the cardiovascular system. However, the
mechanism by which garlic oil protects diabetes-induced cardiomyopathy is unclear. In this study,
streptozotocin (STZ)-induced diabetic rats received garlic oil (0, 10, 50, or 100 mg/kg of body weight)
by gastric gavage every 2 days for 16 days. Normal rats without diabetes were used as control. Cardiac
contractile dysfunction examined by echocardiography and apoptosis evaluated by terminal deoxy-
nucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay were observed in diabetic rat
hearts. Additionally, a shift in cardiac myosin heavy chain (MHC) gene expression from R- to β-MHC
isoform, decreased levels of superoxide dismutase-1 (SOD-1) and cardiac R-actin, and elevated
cardiac thiobarbituric acid reactive substances (TBARS) and caspase- and p38-NFκB-leading apopto-
sis signaling activities were demonstrated in diabetic hearts. However, these diabetes-related cardiac
dysfunctions were almost dose-dependently ameliorated by garlic oil administration. In conclusion,
garlic oil possesses significant potential for protecting hearts from diabetes-induced cardiomyopathy.
KEYWORDS: Diabetes-induced cardiomyopathy; echocardiography; oxidative stress; superoxide
dismutase; garlic oil
Diabetes mellitus is one of the major risk factors for the
all diabetic mortality. The mortality of cardiac disease in
patients with diabetes is 2-4-fold higher than that in subjects
well documented in diabetes. Several pathologic processes
may initiate myocyte injury and dysfunction in patients with
Hyperglycemia, resulting from either insulin deficiency in type
1 diabetes or insulin resistance in type 2 diabetes, induces the
production of reactive oxygen species (ROS), which is the major
cause of diabetic myocardial injury. Due to low content of free
radical scavengers, the heart is susceptible to being damaged by
include abnormal gene expression, altered signal transduction,
and activation of pathways leading to programmed myocardial
cell death (2). Several antioxidant enzymes were identified to be
decreased in the diabetic heart, due to hyperglycemia, by an
oxidative mechanism found in both rats and humans. Interest-
ingly, the exogenous or insulin-mediated antioxidant ability can
inhibit thisrise in oxidative stress, indicatinga possible beneficial
Although apoptosis has long been considered a mechanism
for the elimination of redundant cells, it has only recently been
apoptosis is involved in mechanisms of many human diseases,
*Address correspondence to this author at the Department of
Biological Science and Technology, School of Life Science, China
Medical University, No. 91 Hsueh-Shih Road, Taichung 40402, Taiwan
(e-mail email@example.com; phone 886-4-2205-3366, ext. 2510;
J. Agric. Food Chem., Vol. 58, No. 19, 2010Ou et al.
including cardiovascular disorders, such as diabetic cardiomyo-
pathy (5). Apoptosis contributes to loss of cardiomyocytes in
and is recognized as a predictor of adverse outcomes in subjects
with failing hearts(6). Therefore,the assessmentof the apoptosis
process could be a good way to predict the development of heart
failure induced by diabetes, and the specificity of the related
signaling pathways involved in apoptosis must be evaluated.
as medicinal role for centuries. Due to its exhibition of inhibiting
preventing lipid peroxidation of oxidized erythrocytes and LDL,
ing enzyme (7), even today the medicinal use of garlic is wide-
spread and growing. The main property of garlic for therapeutic
effects is from the effective antioxidant activity against oxidative
damage in cardiovascular diseases (8). In addition, according to
Ryan’s paper (9), garlic is also the most commonly used alter-
native medication of diabetic patients, and it was also reported
garlic can improve hyperglycemia in diabetic patients (10, 11).
However, information about the effect of garlic ondiabetic heart
functions and related mechanisms is very limited.
Figure 1. Analysis profile of garlic oil by gas chromatography. Diallyl sulfide (peak 1), methyl allyl disulfide (peak 2), diallyl disulfide (peak 3), methyl allyl
trisulfide (peak 4), and diallyl trisulfide (peak 5) were quantified at 3.77, 2.75, 40.83, 7.17, and 38.93% in garlic oil, respectively.
Table 1. Physiological and Echocardiographic Parameters in Rat Heartsa
controlGO-0 GO-10GO-50 GO-100
At Basal Level
blood glucose, mg/dL
187.1 ( 14.4
74.6 ( 7.5
414.7 ( 10.7
5.72 ( 0.55
2.63 ( 0.27
51.0 ( 2.2
87.7 ( 1.0
0.20 ( 0.03
155.0 ( 14.1*
266.2 ( 59.8*
403.0 ( 22.8
5.27 ( 0.40
2.63 ( 0.05
49.6 ( 3.8
85.7 ( 2.8
0.19 ( 0.05
137.5 ( 17.5*
205.0 ( 42.2*
399.4 ( 30.0
6.12 ( 0.78
2.93 ( 0.54
49.9 ( 4.2
85.7 ( 3.6
0.20 ( 0.04
165.0 ( 15.0*
266.8 ( 47.5*
417.8 ( 39.4
5.25 ( 0.40
2.70 ( 0.30
49.0 ( 2.0
85.4 ( 1.7
0.19 ( 0.05
152.5 ( 12.5*
220.3 ( 49.7*
394.4 ( 21.2
6.01 ( 0.63
3.07 ( 0.31
49.8 ( 4.1
85.6 ( 3.4
0.19 ( 0.02
After 16 Days of Feeding of Different Doses of Garlic Oil
blood glucose, mg/dL
300.0 ( 13.1
68.9 ( 9.10
407.0 ( 13.3
6.58 ( 0.18
2.78 ( 0.14
58.1 ( 1.40
91.5 ( 0.90
0.24 ( 0.19
205.0 ( 15.0*
425.0 ( 35.4*
291.0 ( 14.2*
6.83 ( 0.38
3.83 ( 0.29*
44.0 ( 3.3*
80.2 ( 3.5*
0.10 ( 0.04*
162.5 ( 27.5*
367.0 ( 52.0*
288.0 ( 6.1*
5.47 ( 0.15*,†
2.60 ( 0.12†
43.7 ( 1.8*
80.1 ( 1.8*
0.10 ( 0.03*
200.0 ( 30.0*
406.0 ( 13.0*
320 ( 0.9*
6.87 ( 0.32
3.80 ( 0.06*
52.2 ( 1.3†
87.8 ( 1.0†
0.19 ( 0.03†
210.0 ( 20.0*
425.0 ( 35.0*
335.0 ( 3.1*,†
6.56 ( 0.15
3.05 ( 0.03†
53.4 ( 1.5†
88.5 ( 1.0†
0.20 ( 0.01†
aGO, garlic oil; HR, heart rate; BW, body weight; LVEDD, left ventricular end diastolic diameter; LVESD, left ventricular end systolic diameter; FS, fractional shortening; EF,
ejection fraction; CO, cardiac output; LVM, left ventricular mass; TL, tibia length. The percentage of LV fractional shortening (FS, %) was calculated as [(LVEDD - LVESD)/
LVESV)/LVEDV] ?100(%).Cardiac output(CO) was calculatedasstrokevolume? heartrate(L/min). Resultsare mean( SEof sixrats per group.GO-0,-10,-50,and-100
represent doses of 0, 10, 50, and 100 mg of garlic oil/kg of body weight, respectively. *, P < 0.05 compared with control rats; †, P < 0.05 compared with GO-0 group rats.
J. Agric. Food Chem., Vol. 58, No. 19, 2010
Therefore, in the present study, we hypothesize that cardiac
contractile dysfunction and apoptosis induced by diabetes can
the cardiac contractile function, oxidative stress-related pro-
teins, myofibrillar formation, apoptosis, and related signaling
MATERIALS AND METHODS
Materials. Fresh garlic (A. sativum) was purchased from the local
market, and garlic oil was prepared by steam distillation (12). The final
Hewlett-Packard, Palo Alto, CA), which was processed as described (13).
The constituent profile of the garlic oil is shown in Figure 1. The major
essential components include diallyl disulfide (DADS), diallyl trisulfide
(DATS), diallyl sulfide(DAS),andminoramountsofmany othervolatile
compounds. The monoclonal antibody against caspase 3 was purchased
from Cell Signaling Technology Inc. (Beverly, MA), and polyclonal
antibodies against cytochrome c, superoxide dismutase (SOD)-1, R-actin,
caspases 8 and 9, and truncated BH3 interacting domain death agonist
(tBid) were purchased from Santa Cruz Biotechnology Inc. (Santa Cruz,
Animal Model and Treatments. Male Wistarrats(4 weeks old) were
purchased from the National Animal Breeding and Research Center
(Taipei, Taiwan). The animals were kept under a 12 h light-dark cycle,
and ambient temperature was maintained at 25 ?C. Animals were given
free access to water and standard laboratory chow (Lab Diet 5001; PMI
Nutrition International Inc., Brentwood, MO). Housing conditions and
experimental procedures were performed according to the NIH Guide for
the Care and Use of Laboratory Animals, and all protocols were approved
by the Institutional Animal Care and Use Committee of China Medical
University, Taichung, Taiwan. After 1 week of acclimatization, diabetes
buffer, pH 4.5) into a lateral tail vein. At 3 days after injection, glycemia
was measured with the Accu-Check Compack kit (Roche Diagnostics
Gmbh, Mannheim, Germany). Only animals in which hyperglycemia had
been successfully induced were randomly separated into four groups
2 mL/kg of BW) every other day for 16 days. The other normoglycemic
of treatment, all animals were anesthetized and echocardiography was
performed. Then, they were sacrificed, and their hearts were removed for
In Vivo Cardiac Function. Transthoracic echocardiograms were
the garlic oil feeding by an echo machine (Vivid i, 10S transducer, GE
Medical Systems, Milwaukee, WI) using a 4-11 MHz phase-array
transducer. M-mode images were obtained in the parasternal long- and
short-axis views of the left ventricle.
TUNEL Assay. All of the procedure was described in our previous
Figure 2. (A)ExpressionofR-MHCandβ-MHC,analyzedbysemiquantitativeRT-PCR.GAPDHwasusedasaloadingcontrol.(B)ExpressionofR-actinof
Values are expressed as a percentage of the control group. The average result( SE of three independent experiments is shown. GO-0, -10, -50, and -100
J. Agric. Food Chem., Vol. 58, No. 19, 2010 Ou et al.
of TUNEL-positive cardiac myocytes was determined. All morphometric
a blinded manner.
Determination of the Thiobarbituric Acid Reactive Substance
of 10% trichloroacetic acid, centrifuged at 10000g for 10 min. One
milliliter of the supernatant was mixed with 1 mL of 0.4% thiobarbituric
acid (TBA) reagent in 0.2 N HC1 and 0.1 mL of 0.2% BHT in 95%
TBA-malondialdehyde (MDA) adduct was extracted with 3 mL of
isobutanol, and the fluorescence was measured with excitation at 515 nm
and emission at 550 nm. 1,1,3,3-Tetramethoxypropane (Sigma Chemical,
St. Louis, MO) was used as the standard for the determination.
Tissue Extraction. The left ventricle samples were homogenized for
protein extract in a PBS buffer (0.14 M NaCl, 3 mM KCl, 1.4 mM
KH2PO4, 14 mM K2HPO4) at a concentration of 1 mg of tissue/10 μL of
PBS for 5 min. The homogenates were centrifuged at 12000 rpm for
30 min. Then, supernatant was collected for further analysis.
Electrophoresis and Western Blot. The protein content of cardiac
tissue extract was analyzed using the Bradford protein assay. Extracted
to nitrocellulose membranes. All of the procedure was described in our
previous study (15). Nonspecific protein binding was stopped in blocking
buffer (5% milk, 20 mM Tris-HCl, pH 7.6, 150 mM NaCl, and 0.1%
Tween 20) and blotted with specific first antibodies in the blocking buffer
at 4 ?C overnight. For repeated blotting, nitrocellulose membranes were
Inc., Rockford, IL) at room temperature for 30 min. Signal intensity was
quantitated using a PhosphoImager. R-Tubulin was used as a loading
RT-PCR. Total RNA was extracted using the Ultraspec RNA
of the manufacturer. All of the procedure was described in our previous
study (15). The cDNA was amplified by PCR with R-MHC primers,
forward primer (50-GGCAG ATATG AAGGG AAGAT-30) and reverse
primer (50-CGAAC ATGTG GTGGT TGAAG-30); β-MHC primers,
forward primer (50-CTTCA ACCAC CACAT GTTCG-30) and reverse
primer (50-TATTG TAGTC CACGG TGCCA-30); and GAPDH pri-
mers, forward primer (50-TCCCT CAAGA TTGTC AGCAA-30) and
reverse primer (50-AGATC CACAA CGGAT ACATT-30).
Statistical Analysis. Statistical differences were examined by one
were expressed as the mean ( SE.
Improved Cardiac Contractile Dysfunction in Diabetic Rats As
Response to Garlic Oil Feeding. To assess cardiac function and
dimension in vivo, we performed echocardiography on rats. In
addition to significantly high blood glucose induced in diabetic
rats, all of the animals showed a normal appearance and had a
days after induction, diabetes decreased body weight and in-
creased blood glucose in all diabetic animal groups. However,
diabetes significantly decreased the heart rate (HR), which was
dose-dependently reversed by garlic oil (GO) feeding. The per-
centagesoffractional shortening(FS) and ejectionfraction(EF),
typically representing cardiac contractile function of rat hearts,
were significantly decreased in diabetic rat hearts and dose-
dependently reversed back to the control level at GO doses of
50and100mg/kgofBW. Thecardiacoutput(CO) showssimilar
results to HR, FS, and EF. Collectively, diabetic rats receiving
GO showed a better cardiac output and contractile function.
Shift in Myosin Heavy Chain Gene Expression from r to
β Isoforms and Decreased Levels of Cardiac r-Actin in Diabetic
Rat Hearts Were Reversed by Garlic Oil Feeding. To evaluate the
contractile function of cardiac muscle, we examined MHC gene
expressions in rat heart, using RT-PCR analysis. The result is
shown in Figure 2A. Compared with the control group, the
diabetic rats showed a significantly decreased level of R-MHC
and an increased level of β-MHC isoforms, which were dose-
dependently reversed by the administration of GO. Even the
sarcomeric contractile protein are shown in Figure 2B. This
myofibrillar protein in diabetic hearts was drastically reduced
and dose-dependently restored by treatment with GO. In the
Figure 3. (A) Expression of superoxide dismutase-1 (SOD-1) of cardiac muscle analyzed by Western blotting. Signal intensity was quantitated using a
garlic oil/kg of body weight, respectively. /, P < 0.05 compared with control rats; †, P < 0.05 compared with GO-0 group rats.
J. Agric. Food Chem., Vol. 58, No. 19, 2010
reached control level. On the contrary, the levels of nonsarco-
meric smooth muscle R-actin did not alter at all.
Decreased Antioxidant Enzyme Expression and Elevated Lipid
Peroxidation in Diabetic Rat Hearts Were Reversed by Garlic Oil
Supplementation. Intracellular ROS levels are regulated by anti-
of SOD-1 in diabetic hearts in response to GO administration.
resulting from the oxidative damage was assessed by measuring
the TBARS level in hearts. Interestingly, GO treatment dose-
suggesting that the reduced SOD-1 activity for ROS scavenger
and the increased lipid peroxidation were reversed by the GO.
Development of Apoptosis in Diabetic Rat Hearts Was Amelio-
rated by Garlic Oil Feeding. To clarify whether cardiac apoptosis
rats, DAPI staining and TUNEL assay were examined, and the
results are shown in Figure 4. Compared with control group, a
5-fold increase in apoptosis in the left ventricles of rats was
induced bydiabetes, and thisincreasewas significantly decreased
by GO in a dose-dependent manner. This indicates that GO
feeding can improve diabetes-induced cardiac damages.
Increased Activities of Caspase-Leading Apoptotic Signalings
and p38-NFKB Signaling in Diabetic Rat Hearts Were Decreased
by Garlic Oil Feeding. To further understand the signaling path-
ways involved in the cardiac apoptosis development induced by
protein levels in control and diabetic rat hearts. Compared with
the control group, a 2.2-fold increase of active caspase 3 was
observed in diabetic rat hearts. This induction was inhibited by
the administration of GO and reached significantly low levels at
doses of 50 and 100 mg/kg of BW (Figure 5A). These data are
consistent with the results of cardiac apoptosis in Figure 3.
Furthermore, compared with the control group, both caspase 8
and caspase 9 were elevated 2.5- and 3.4-fold, respectively, in
diabetic rat hearts. This elevation was decreased and reached
significantly decreased levels at a dose as low as 50 mg/kg of BW
(Figure 5B,C). The levels of released cytochrome c and tBid
showed similar results in Figure 5B,C. Collectively, both the
receptor-dependent and mitochondrion-dependent apoptotic
pathways are involved in diabetes-induced cardiac apoptosis,
which is inhibited by the administration of GO.
Protein p38 and its downstream nuclear factor κB (NFκB) are
known as regulators of pathophysiological gene expressions to
cause cell death, leading to left ventricle dysfunction (16). Hence,
are shown in Figure 5D. The significant increase of cardiac p-p38
level in diabetic hearts was significantly decreased at all three
doses of GO. The cardiac NFκB in diabetes was more activated
and as significantly attenuated after GO treatment at doses of 50
and 100 mg/kg of BW.
heart rate, cardiac contractile function, cardiac output, impaired
contractile velocity of cardiac muscle, and myofibril formation.
are improved by GO treatment. We also show that diabetes
develops cardiac apoptosis and oxidative stress and increases
suggest that these oxidative stress- and apoptosis-promoting
events might be associated with cardiac dysfunction in diabetic
cardiomyopathy. GO treatment, which reduces oxidative stress,
alleviates apoptosis development and counteracts activations of
up-regulated apoptotic signalings that might be considered to
ties of garlic. Interestingly, Anwar’s (10) study indicates that GO
effectively normalized the unbalanced antioxidant status by
decreasing lipid peroxide and increasing SOD activity and total
thiol in the blood and liver of rats with STZ-induced diabetes.
These results provide an explanation for our experimental data
demonstrating that GO treatment ameliorates cardiac dysfunc-
tion, myofibril malformation, and apoptosis with related sig-
naling activities in STZ diabetic rat hearts through decreasing
oxidativestress. Additionally, in agreement with our hypothesis
Figure 4. Apoptoticcelldeathinthehearts of controlanddiabeticratsfed
different doses of garlic oil. The degree of apoptosis in rat hearts was
measured by TdT-mediated deoxynucleotidyl UTP nick-end labeling
(TUNEL) assay. The percentage of TUNEL-positive cells was determined
control group.Data are means( SEofsix rats pergroup. GO-0, -10, -50,
and -100 represent doses of 0, 10, 50, and 100 mg of garlic oil/kg of body
weight, respectively. /, P < 0.05 compared with control rats; †, P < 0.05
compared with GO-0 group rats.
J. Agric. Food Chem., Vol. 58, No. 19, 2010Ou et al.
(Fgure 6), the study of Banerjee et al. (17) indicates that garlic
with powerful antioxidant ability can increase glutathione
in cardiac muscles. In our analysis of results by using gas
chromatography, more than 20 organosulfur compounds have
been found in GO. Among them, DADS and DATS are the
powerful antioxidant potential (18,19). They may play impor-
tant roles in improving diabetic cardiac disease with GO treat-
ment. In addition, although the treatment with GO failed to
improve the glucose intolerance caused by STZ, the oxidative
stress, downstream of hyperglycemia, was inhibited by GO
Contractile failure resulting in the decline in ventricular per-
formance is another characteristic of diabetic cardiomyopathy.
Several studies suggest that ventricular dysfunction associated
with diabetes mellitus is also linked to the production of
ROS (20), resulting from hyperglycemia. Although the model
of STZ-induced type I DM was used in the present study, we
believe that no matter which type of diabetes was examined,
hyperglycemia, as the common characteristic of both types of
diabetes, is the major cause of cardiac damage. The evidence of
Figure 5. Cardiacpro-apoptoticproteins(A)caspase3,(B)caspase8andtBid,(C)caspase9andreleasedcytochromec,and(D)reactiveoxygenspecies
(ROS) downstream proteins, phosphorylated p38, and NFκB in control and diabetic rats fed different doses of garlic oil. Protein levels in left ventricles of rat
average result ( SE of three independent experiments is shown. GO-0, -10, -50, and -100 represent doses of 0, 10, 50, and 100 mg of garlic oil/kg of body
weight, respectively. /, P < 0.05 compared with control rats; †, P < 0.05 compared with GO-0 group rats.
J. Agric. Food Chem., Vol. 58, No. 19, 2010
diabetic cardiomyopathy (DCM) detected clinically by echo-
cardiography is an early increase in diastolic myocardial stiffness
indicated as left ventricular diastolic dysfunction and a later
occurrence of left ventricular systolic dysfunction (21). Using
ananimal model,Schaffer and his colleagues observedthatSTZ-
induced diabetic rats demonstrated glucose intolerance and
contractility, and cardiac output (22), which are very consistent
with our results.
MHCs are the molecular motors of cardiac muscle contrac-
tion. TheRand βisoforms differ in contractile propertiesonthe
basis of their relative adenosine triphosphatase (ATPase) activ-
to β in ventricle may explain the systolic dysfunction in the left
ventricle during heart failure (23,24). We show here a reduced
R-MHC expression and an increased β-MHC expression in
by GO supplementation. A similar result in Aragno’s study (25)
demonstrates that reduced myocardial contractility caused by
oxidative stress in the diabetic heart evidenced by the shift of
MHC isoforms was reversed by the administration of dehydro-
epiandrosterone (DHAE), a physiological steroid with multi-
targeted antioxidant properties. Similarly, our observation also
diabetic heart was alleviated by GO treatment. In addition,
Dyntar’s study (26) demonstrates that normal rebuilding of
myofibrillar structures evaluated by the altered cardiac R-actin
level were disrupted in cultured adult cardiomyocyte exposed to
high glucose concentration. Interestingly, this impairment can
be fully improved by treatment of antioxidant NAC. These
contribute to the change of myosin chain gene expressions and
myofibrillar formation, leading to myofibril remodeling, and it
is likely that improved redox balance could be the underlying
mechanism of garlic’s beneficial effects on the diabetic heart.
apoptotic signalings, p38-NFkB signaling was also involved in
the development of cardiac apoptosis, which was prevented and
This finding is consistent with Riad’s results (16), indicating that
treatment of chronic p38 inhibition reduces the development of
cardiac and endothelial dysfunction in diabetic rats. We con-
p38 leads to the activation of transcription factor, nuclear factor
κB (NFκB) to induce many pathophysiologic developments such
by antioxidant supplementation. Another interesting study (19)
related the inhibition of lipopolysaccharide (LPS)-induced oxi-
GO component, supporting the therapeutic potential of GO on
oxidative stress-related diseases.
Recently, the endogenously generated hydrogen sulfide
(H2S) has been reported to demonstrate potent cardioprotec-
tive effects. Several mechanisms of cardiovascular actions of
H2S are investigated. The ATP-sensitive Kþ(KATP) channel is
the H2S-induced vasodilation (28) and infarct-limiting effect in
hearts (29). Furthermore, downstream of the KATPchannel
opening is the PKCε, which plays a role in cardioprotective
signaling following injury stimuli (30). Anti-apoptotic proper-
ties of H2S are also evidenced by the activation of several
survival kinases, including ERK and PI3K/Akt (31), which
may up-regulate endogenous antioxidant enzyme activities,
through the nuclearfactorE2-relatedfactor-2(Nrf-2)-dependent
pathway (32). Interestingly, a recent study by Benavides’s group
indicated that GO and its components DADS and DATS can
induce H2S production (33), and GO was identified to promote
Nrf-2 activation (34). Using the H2S inhibitor propargylglycine,
the main bioactive constituent of garlic, S-allylcysteine, was
identified to exert cardioprotective action through the H2S-
mediated pathway (35). On the basis of the observations of low
levels of H2S in the blood of STZ-treated rats (36), all of these
antidiabetic cardiomyopathy of garlic oil.
In conclusion, our results show that GO supplementation for
including cardiac contractile functions and structures, myosin
chain gene expressions, oxidative stress, and apoptosis and
related signaling activities. All of these phenomena might be
associated with the antioxidant potential of GO, which is attrib-
uted to the presence of organosulfur compounds that modulate
the cardiac antioxidant activity. A future study to investigate the
individual GO constituent compounds on improving diabetic
cardiac dysfunction is needed.
BW, body weight; CO, cardiac output; Cyt c, cytochrome c;
DATS, diallyl trisulfide; DAPI, 40,6-diamidino-2-phenylindole;
Figure 6. Proposedhypothesisthatgarlicoilamelioratescardiacdysfunc-
tion in rats with diabetes by down-regulating apoptotic signaling activities.
We speculate that garlic oil protects hearts by inhibiting oxidative stress in
diabetic rats. Each up arrow and down arrow represent increases and
decreases, respectively. These STZ-induced mitochondrion-, death re-
ceptor-, and p38-NFκB-dependent apoptotic signaling activities were
significantly improved after garlic oil supplementation. Through these
mechanisms, garlic oil treatment ameliorates cardiomyopathy in rats with
heart rate, and contractile function as well as myofibril formation.
J. Agric. Food Chem., Vol. 58, No. 19, 2010Ou et al.
DCM, diabetic cardiomyopathy; DHAE, dehydroepiandroster-
one; EF, ejection fraction; FS, fractional shortening; GAPDH,
glyceraldehyde-3-phosphate dehydrogenase; GO, garlic oil; GST,
glutathione S-transferase; H&E, hematoxylin and eosin; HR,
heart rate; IGF-I(R), insulin-like growth factor-I (receptor);
iNOS, inducible nitric oxide synthase; LPS, lipopolysaccharide;
LVEDD, left ventricular end diastolic diameter; LVESD, left
ventricular end systolic diameter; LVESV, left ventricular end
diastolic volume; LVESV, left ventricular end systolic volume;
LVM, left ventricular mass; MHC, myosin heavy-chain; NAC,
N-acetylcysteine; NFκB, nuclear factor-κB; OGTT, oral glucose
tolerance test; PI3K, phosphadidylinositol 30-kinase; PWT, pos-
terior wall thickness; ROS, reactive oxygen species; STZ, strepto-
zotocin; TBARS, thiobarbituric acid reactive substances; tBid,
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Received for review April 27, 2010. Revised manuscript received
August 18, 2010. Accepted August 18, 2010. This study is supported
by the National Science Council of Republic of China (Grant NSC96-
2320-B-039-035-MY3), by the Taiwan Department of Health Clinical
inpartbythe TaiwanDepartment ofHealth CancerResearchCenter of