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Up-regulation of PPARγ, heat shock protein-27 and -72 by naringin ... https://www.ncbi.nlm.nih.gov/pubmed/21736771 by AK Sharma - ‎2011 - ‎Cited by 78 - ‎Related articles

Article · June 2011with170 Reads
DOI: 10.1017/S000711451100225X. · Source: PubMed
Abstract
Naringin, a bioflavonoid isolated from grapefruit, is well known to possess lipid-lowering and insulin-like properties. Therefore, we assessed whether naringin treatment ameliorates insulin resistance (IR), β-cell dysfunction, hepatic steatosis and kidney damage in high-fat diet (HFD)-streptozotocin (STZ)-induced type 2 diabetic rats. Wistar albino male rats were fed a HFD (55 % energy from fat and 2 % cholesterol) to develop IR and on the 10th day injected with a low dose of streptozotocin (40 mg/kg, intraperitoneal (ip)) to induce type 2 diabetes. After confirmation of hyperglycaemia (>13·89 mmol/l) on the 14th day, different doses of naringin (25, 50 and 100 mg/kg per d) and rosiglitazone (5 mg/kg per d) were administered orally for the next 28 d while being maintained on the HFD. Naringin significantly decreased IR, hyperinsulinaemia, hyperglycaemia, dyslipidaemia, TNF-α, IL-6, C-reactive protein and concomitantly increased adiponectin and β-cell function in a dose-dependent manner. Increased thiobarbituric acid-reactive substances and decreased antioxidant enzyme activities in the serum and tissues of diabetic rats were also normalised. Moreover, naringin robustly increased PPARγ expression in liver and kidney; phosphorylated tyrosine insulin receptor substrate 1 in liver; and stress proteins heat shock protein (HSP)-27 and HSP-72 in pancreas, liver and kidney. In contrast, NF-κB expression in these tissues along with sterol regulatory element binding protein-1c and liver X receptor- expressions in liver were significantly diminished. In addition, microscopic observations validated that naringin effectively rescues β-cells, hepatocytes and kidney from HFD-STZ-mediated oxidative damage and pathological alterations. Thus, this seminal study provides cogent evidence that naringin ameliorates IR, dyslipidaemia, β-cell dysfunction, hepatic steatosis and kidney damage in type 2 diabetic rats by partly regulating oxidative stress, inflammation and dysregulated adipocytokines production through up-regulation of PPARγ, HSP-27 and HSP-72.
Up-regulation of PPARg, heat shock protein-27 and -72 by naringin
attenuates insulin resistance, b-cell dysfunction, hepatic steatosis and
kidney damage in a rat model of type 2 diabetes
Ashok Kumar Sharma
1
, Saurabh Bharti
1
, Shreesh Ojha
1
, Jagriti Bhatia
1
, Narender Kumar
2
, Ruma Ray
2
,
Santosh Kumari
3
and Dharamvir Singh Arya
1
*
1
Department of Pharmacology, Cardiovascular and Diabetes Research Laboratory, All India Institute of Medical Sciences,
New Delhi 110029, India
2
Department of Pathology, All India Institute of Medical Sciences, New Delhi 110029, India
3
Department of Plant Physiology, Indian Agricultural Research Institute, Pusa, New Delhi 110012, India
(Received 2 February 2011 – Revised 15 March 2011 – Accepted 15 March 2011 – First published online 21 June 2011)
Abstract
Naringin, a bioflavonoid isolated from grapefruit, is well known to possess lipid-lowering and insulin-like properties. Therefore, we
assessed whether naringin treatment ameliorates insulin resistance (IR), b-cell dysfunction, hepatic steatosis and kidney damage in
high-fat diet (HFD)streptozotocin (STZ)-induced type 2 diabetic rats. Wistar albino male rats were fed a HFD (55 % energy from fat
and 2 % cholesterol) to develop IR and on the 10th day injected with a low dose of streptozotocin (40 mg/kg, intraperitoneal (ip)) to
induce type 2 diabetes. After confirmation of hyperglycaemia (.13·89 mmol/l) on the 14th day, different doses of naringin (25, 50 and
100 mg/kg per d) and rosiglitazone (5 mg/kg per d) were administered orally for the next 28 d while being maintained on the HFD.
Naringin significantly decreased IR, hyperinsulinaemia, hyperglycaemia, dyslipidaemia, TNF-a, IL-6, C-reactive protein and concomitantly
increased adiponectin and b-cell function in a dose-dependent manner. Increased thiobarbituric acid-reactive substances and decreased
antioxidant enzyme activities in the serum and tissues of diabetic rats were also normalised. Moreover, naringin robustly increased
PPARgexpression in liver and kidney; phosphorylated tyrosine insulin receptor substrate 1 in liver; and stress proteins heat shock protein
(HSP)-27 and HSP-72 in pancreas, liver and kidney. In contrast, NF-kB expression in these tissues along with sterol regulatory element
binding protein-1c and liver X receptor- expressions in liver were significantly diminished. In addition, microscopic observations validated
that naringin effectively rescues b-cells, hepatocytes and kidney from HFD-STZ-mediated oxidative damage and pathological alterations.
Thus, this seminal study provides cogent evidence that naringin ameliorates IR, dyslipidaemia, b-cell dysfunction, hepatic steatosis and
kidney damage in type 2 diabetic rats by partly regulating oxidative stress, inflammation and dysregulated adipocytokines production
through up-regulation of PPARg, HSP-27 and HSP-72.
Key words: Insulin resistance: Type 2 diabetes: Naringin: Heat shock proteins: PPARg
Diabetes mellitus remains an important cause of morbidity
and mortality, has reached burgeoning epidemic proportions
affecting about 285 million individuals in 2010 and it is
estimated that the number of diabetic individuals will be
increased by 54 % till 2030
(1)
. More than 90 % of diabetic indi-
viduals have type 2 diabetes mellitus (T2DM), a consequence
of a high-fat diet, sedentary lifestyle and genetic predisposi-
tion, often diagnosed after metabolic dysfunction has taken
hold of multiple organ systems
(1,2)
. The myriad of disorders
linked with T2DM includes oxidative stress, inflammation,
insulin resistance (IR), insulin deficiency due to b-cell failure,
dyslipidaemia, hepatic steatosis, diabetic nephropathy and
retinopathy
(3 – 5)
. Emerging data indicate that chronic subacute
inflammation in T2DM manifested as dysregulated production
of C-reactive protein (CRP), TNF-a, IL-6 and adiponectin plays
a crucial role in IR
(2,6)
.
Tragically, the present therapy for diabetes including insulin
and various oral anti-diabetic agents is frequently constrained
*Corresponding author: D. S. Arya, fax þ91 11 26584121, email dsarya16@hotmail.com
Abbreviations: CRP, C-reactive protein; ER, endoplasmic reticulum; GSH þPx, glutathione peroxidase; HFD, high-fat diet; HOMA, homoeostasis model
assessment; HSP, heat shock protein; ip, intraperitoneal; IR, insulin resistance; IRS1, insulin receptor substrate 1; LXRa, liver X receptor-a; P-IRS1,
phosphorylated tyrosine 612 IRS1; SOD, superoxide dismutase; SREBP-1c, sterol regulatory element binding protein-1c; STZ, streptozotocin; T2DM,
type 2 diabetes mellitus; TBARS, thiobarbituric acid-reactive substances; TC, total cholesterol.
British Journal of Nutrition (2011), 106, 1713–1723 doi:10.1017/S000711451100225X
qThe Authors 2011
British Journal of Nutrition
by safety, tolerability, oedema, weight gain, lactic acidosis and
gastrointestinal intolerance
(7)
. Moreover, drugs from natural
resources are usually considered safe, and accessible interven-
tions are becoming more popular in the treatment of several
disorders including diabetes
(8)
. Human diets of plant origin
contain polyphenols, phytosterols, phyto-oestrogens, phytates
and PUFA, and so on. Among them, naringin, a polyphenol
and naturally occurring bioflavonoid in grapefruits and
oranges, has been reported to possess antioxidant, anti-
diabetic, lipid-lowering, anti-atherogenic and anti-inflammatory
activities
(9 – 12)
.
Interestingly, it is well known that activation of PPARg,a
nuclear receptor, can exert anti-inflammatory and antioxidant
effects in several cell types including pancreatic b-cells, hepato-
cytes and glomeruli
(13,14)
. Therefore, PPARgligands can offer
a therapeutic intervention in modifying diabetes and the
metabolic syndrome
(14)
. Besides, heat shock proteins (HSP)
are powerful antioxidant and anti-inflammatory proteins that
protect cells against many acute and chronic stressful
conditions
(15 – 17)
. Furthermore, activation of PPARgand HSP
prevents IR and complications of diabetes by inhibiting NF-kB
and c-jun amino terminal kinase activation
(13 – 17)
. Recently,
Jung et al.
(11)
reported that naringin up-regulated the PPARg
receptor; therefore, we hypothesise that it might ameliorate IR,
diabetes and its associated complications in high-fat diet
(HFD)streptozotocin (STZ)-induced type 2 diabetic rats.
With this background, for the first time, the present study
was undertaken to investigate the effects of naringin on
glucose homoeostasis, dyslipidaemia, hepatic steatosis and
kidney damage in HFD-STZ-induced type 2 diabetic rats. To
delineate the mechanisms whereby naringin exerts its insu-
lin-sensitising and anti-diabetic activity in diabetic rats, we
evaluated oxidative stress, CRP, adipocytokines, as well as
protein expression of PPARg, HSP-27, HSP-72, NF-kB, sterol
regulatory element binding protein-1c (SREBP-1c), liver X
receptor a(LXRa) and phosphorylated tyrosine and total
insulin receptor substrate. Morphological examinations of
pancreas, liver and kidneys were also performed to further
substantiate the beneficial effect of naringin on pathological
alteration induced by HFD-STZ treatment. In the present
study, the efficacy of naringin was compared with that of
the PPARgagonist rosiglitazone, which is an insulin-sensitising
anti-diabetic drug.
Materials and methods
Animals and diet
The experimental protocol was approved by the Institutional
Animal Ethics Committee and conformed to the Indian
National Science Academy guidelines for the use and care of
experimental animals in research. Normal rat chow (55 %
carbohydrate, 24 % protein, 6 % ash, 5·0 % moisture, 5 % fat,
4 % fibre, 0·6 % Ca and 0·4 % P, w/w) and a high-fat diet
(25 % coconut oil, 2 % cholesterol and 73 % normal rat chow;
provides 55 % of the animal’s energy as fat) were used and
animals were allowed access to food and water ad libitum.
Wistar albino male rats (170– 200 g, n72) were adapted to
the experimental conditions at 25 ^58C with a relative humid-
ity of 60 ^5 % on a light –dark cycle of 12 –12 h for 1 week.
Drug and chemicals
Naringin and STZ were purchased from Sigma Chemicals (St
Louis, MO, USA). Rosiglitazone was obtained as a gift
sample from Zydus Cadila (Ahmedabad, Gujarat, India). Rat
TNF-a(Diaclone Tepnel Company, France), IL-6, high-sensi-
tivity CRP (Bender MedSystems, Inc., Vienna, Austria), adipo-
nectin (Linco Research, Inc., St Charles, MO, USA) and insulin
(Mecrodia AB, Uppsala, Sweden) ELISA Kits were used. The
kits for blood glucose, NEFA and lipid profile were purchased
from Logotech Private Limited (New Delhi, India). All primary
and secondary antibodies were procured from Santa Cruz Bio-
technology, Inc. (Santa Cruz, CA, USA) except HSP-72 primary
antibody (Stressgen, Plymouth Meeting, PA, USA).
Induction of type 2 diabetes and dyslipidaemia
Animals were assigned to either regular rat chow (normal con-
trol) or the HFD. After 10 d of the HFD, overnight-fasted (12 h)
rats were injected a single injection of freshly prepared STZ
(40 mg/kg, intraperitoneally (ip), in 0·1 M-citrate phosphate
buffer, pH 4·5). The development of hyperglycaemia in rats
was confirmed by estimation of fasting serum glucose 72 h
after STZ injection. Rats with fasting serum glucose level
above 13·89 mmol/l were considered diabetic and recruited in
the study. The schematic diagram of the experimental protocol
is given in Fig. 1.
Experimental design
The rats were randomly divided into six groups comprising of
twelve animals in each and were treated with naringin and
rosiglitazone by oral administration for 28 d while being main-
tained on the HFD. Group I, normal control (normoglycaemic
rats fed with normal diet only), group II, diabetic control
(diabetic rats fed with HFD), groups IIIV, diabetes– HFD
rats, were administered naringin at 25, 50 and 100 mg/kg
per d, by mouth respectively
(12)
. Group VI, DiabetesHFD
rats, were administered rosiglitazone at 5 mg/kg per d.
At the end of the experiment, blood samples were withdrawn
from the tail vein of overnight-fasted rats. Subsequently, an glu-
cose tolerance test and insulin tolerance tests were performed.
The animals were then killed with an overdose of sodium pento-
barbitone (150 mg/kg, ip) and their pancreas, liver and kidney
were removed and processed for biochemical and microscopic
examination. Serum was separated by centrifugation (Biofuge;
1st day
14th day
11th day
STZ (40 mg/kg, i.p.) 1st day 28th day
N 25, 50 and 100 mg/kg and RSG 5 mg/kg, p.o.
42th day
High-fat diet (42 d)
(Confirmation of diabetes; FSG >13·89 mmol/l)
Fig. 1. Schematic diagram of experimental protocol. FSG, fasting serum
glucose; N, naringin; RSG, rosiglitazone; p.o., by mouth; i.p., intraperitoneally.
A. K. Sharma et al.1714
British Journal of Nutrition
Heraeus, Germany) at 3000 gfor 5 min and analysed for glucose,
total cholesterol (TC), TAG, LDL-C, HDL-C, insulin, adiponectin,
IL-6, CRP and TNF-a.
Glucose tolerance and insulin tolerance tests
Overnight-fasted rats were administered glucose (1 g/kg, ip)
and subsequently blood was withdrawn from the tail to esti-
mate blood glucose levels at 21, 30, 60, 120 and 180 min
using a glucometer (OneTouch; Johnson and Johnson, New
Brunswick, NJ, USA). Once blood glucose levels returned to
baseline, an insulin tolerance test was performed with insulin
(Humulin, 0·5 units/kg, ip; Eli Lilly, India) and blood glucose
levels were determined at 21, 15, 30, 45 and 60 min
(5)
.
Serum glucose, NEFA and lipid profile
Serum glucose, NEFA and lipid profiles (TC, TAG, LDL-C
and HDL-C) were estimated spectrophotometrically using
commercial kits.
Serum insulin, IL-6, C-reactive protein, TNF-
a
and adiponectin
Serum insulin, IL-6, CRP, TNF-aand adiponectin levels were
measured by ELISA kits following the manufacturers’
instructions.
Insulin resistance and
b
-cell function
Homoeostasis model assessment (HOMA) of IR (HOMA-IR)
and HOMA of b-cell function (HOMA-B) were calculated by
the HOMA method using the following equations
(18)
:
IR (HOMA-IR) ¼(fasting glucose (mmol/l) £fasting insulin
(mIU/ml))/22·5, and b-cell function (HOMA-B) ¼(20 £fasting
insulin (mIU/ml))/(fasting glucose (mmol/l) – 3·5).
Thiobarbituric acid-reactive substances, superoxide
dismutase and glutathione peroxidase activities
For measuring thiobarbituric acid-reactive substances (TBARS),
aliquots of 10 % homogenate of pancreas, liver and kidney were
prepared in ice-cold phosphate buffer (0·1 M, pH 7·4). To this
tissue homogenate, 8·1 % SDS (0·2 ml), 20 % acetic acid
(1·5 ml) and 0·8 % thiobarbituric acid (1·5 ml) were added and
heated for 60 min in a boiling water bath. After cooling, the
pink complex was extracted with 5 ml butanol – pyridine
(15:1) mixture. Finally, absorbance of the organic layer was
observed at 532 nm (Specord 200; Jena, Germany) and plotted
against a standard graph and expressed as mmol/g tissue
(19)
.
Tissue homogenate so obtained was centrifuged (Biofuge;
Heraeus, Germany) at 3000 gfor 15 min at 48C and the super-
natant was collected to assess superoxide dismutase (SOD),
glutathione peroxidase (GSH-Px) and protein. For SOD, 100 ml
supernatant, 2·95 ml phosphate buffer (0·1 M, pH 8·4) and
50 ml pyrogallol (7·5 mM) were added and the change in absor-
bance was recorded at an interval of 60 s for 2 min at 420 nm.
One unit of enzyme activity was defined as the amount of
Table 1. Biochemical and histological findings in different experimental groups at the end of the study
(Mean values with their standard deviations; n12)
Normal control Diabetic control Diabetic þN25 Diabetic þN50 Diabetic þN100 Diabetic þRSG5
Parameters Mean SD Mean SD Mean SD Mean SD Mean SD Mean SD
Body weight (g) 231·5 10·6 217·8 14·8 218·6 10·1 225·4 11·2 222·7 13·1 229·5 9·8
Serum glucose (mmol/l)
Initial (day 1) 4·6 0·5 15·7 1·7 15·8 1·3 16·5 1·1 16·1 2·2 16·3 2·7
Final (day 28) 5·0 0·3 16·7** 2·2 14·3 1·4 10·1††† 1·6 8·3††† 1·5 8·6††† 1·9
Insulin (pmol/l) 77·0 5·9 123·2** 7·1 110·9 5·4†† 94·7††† 2·4 88·2††† 4·0 89·6††† 4·9
HOMA-IR 2·46 0·14 12·54** 2·13 10·07 1·56† 6·12††† 1·19 4·64††† 0·63 4·92††† 1·10
HOMA-B 145·9 10·4 26·7** 3·5 34·9 5·0 44·3 10·1 58·7† 12·9 56·8† 10·9
TNF-a(pg/ml) 14·8 3·5 97·5** 4·1 86·1 6·4 52·5††† 7·5 41·3††† 5·1 44·6††† 2·7
IL-6 (pg/ml) 22·8 5·1 56·3** 8·7 50·5 7·8 41·8† 5·6 29·4††† 6·0 31·1††† 4·5
CRP (mg/l) 35 7 136** 13 131 15 102†† 17 73††† 14 75††† 1·6
Adiponectin (mg/ml) 32·4 1·5 18·6** 1·2 22·7 1·8 25·3†† 0·9 28·5††† 1·7 29·0††† 1·2
Islet area (mm
2
) 3237 582 1215** 296 1523 352 2389††† 638 2953††† 525 2820††† 602
Glomerular sclerosis (%) 3 1 9·45** 1·5 8·13 1·86 6·85 2·16 4·15††† 1·3 4·98†† 1·73
Glomerular area (mm
2
) 18 083 2045 16 104 2195 16 709 2345 17 801 2065 18 065 2278 18 049 2132
HOMA-IR, homoeostasis model assessment of IR; HOMA-B, homoeostasis model assessment of b-cell function; CRP, C-reactive protein.
Mean values were significantly different from those of the normal control: **P,0·001, **P,0·001 by one-way ANOVA followed by Bonferroni post hoc test
Mean values were significantly different from those of the diabetic control: †P,0·05, ††P,0·01, †††P,0·001 by one-way ANOVA followed by Bonferroni post hoc test.
Naringin ameliorates type 2 diabetes in rats 1715
British Journal of Nutrition
enzyme required to produce 50 % inhibition of pyrogallol auto-
oxidation under the assay conditions and expressed as U/mg
protein
(20)
.
For GSH-Px, 100 ml supernatant were added to the reaction
mixture containing 1 mM-glutathione reductase in a 0·1 M-Tris
HCL (pH 7·2). The reaction was started by adding 2·5 mM-
hydrogen peroxide and the absorbance was measured for
1 min at 340 nm. A molar extinction coefficient of 6·22 per
mM cm was used to determine the activity and was expressed
as U/mg protein
(21)
.
Further, protein estimation was carried out by adding 10ml
tissue supernatant to 100 mlof1
M-NaOH and 1 ml Bradford
reagent (Sigma Chemicals). The solution so obtained was vor-
texed and the absorbance was recorded at 595 nm
(22)
. Protein
content was determined from a standard curve using known
concentrations of bovine serum albumin (Sigma Chemicals).
Similarly, following the above-mentioned methods, we
assessed TBARS, SOD, GSH-Px and total protein in the serum.
Western blot analysis
Protein samples (50 mg) were separated by 12 % SDS –PAGE,
transferred to a nitrocellulose (MDI, Ambala, Haryana, India)
membrane that was blocked for 1 h with 5 % (w/v) dry milk
in Tris-buffered saline, and incubated overnight at 48C with
the mouse monoclonal primary antibody. The primary anti-
bodies used were as follows: PPARg(1:2000), insulin receptor
substrate 1 (IRS1; 1:1000), phosphorylated tyrosine 612 IRS1
(P-IRS1; Tyr162, 1:1000), HSP-27 (1:1500), HSP-72 (1:1500)
and b-actin (1:2000). The primary antibody was detected
with horseradish peroxidase-conjugated goat anti-mouse sec-
ondary antibody (1:10 000) and Bio-Rad Protein Assay
Reagent, and visualised by Bio-Rad Quantity One software
(release 4.4.0; Bio-Rad, Hercules, CA, USA).
Histopathological examination
Formaldehyde (10 %)-fixed pancreas, liver and kidney tissues
were embedded in paraffin, and serial sections (3 mm) were
cut using a microtome (Leica RM 2125; Leica,Germany). Each
section was stained with haematoxylin and eosin and at least
ten fields per slide were observed under a light microscope
(Nikon, Tokyo, Japan). Endocrine pancreatic damage was
assessed by evaluating changes in islet shape, area, cell
number and presence of inflammatory infiltrate. Liver histo-
pathological changes were graded as described by Jang
30
(a)
(c)
(b)
25
20
15
Glucose (mmol/l)
10
5
0
20
15
Glucose (mmol/l)
10
5
0
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0 min 30 min 60 min 0 min 15 min 30min 45 min 60 min120 min 180 min
Time Time
12
10
8
6
2
0
Concentration (mmol/l)
4
Normal control Diabetic control Diabetic + N25 Diabetic + N50 Diabetic + N100 Diabetic + RSG5
Fig. 2. (a) Glucose tolerance test (GTT); (b) insulin tolerance test (ITT); (c) serum lipid profile at the end of study in different experimental groups. Values
are means, with their standard deviations represented by vertical bars (n12). * Mean values were significantly different from those of the normal control
(P,0·001, one-way ANOVA followed by Bonferroni post hoc test). Mean values were significantly different from those of the diabetic control: **P,0·01,
***
P,0·001, one-way ANOVA followed by Bonferroni post hoc test. TC, total cholesterol. , Normal control; , diabetic control; , diabetic þN25;
, diabetic þN50; , diabetic þN100; , diabetic þRSG5; , TC; , TAG; , LDL-C; , HDL-C; , NEFA.
A. K. Sharma et al.1716
British Journal of Nutrition
et al.
(23)
. The severity of kidney damage was evaluated in terms
of morphological changes, such as hydropic changes in proxi-
mal convoluted tubules, glomerular sclerosis and widening
of matrix. Sclerotic-glomeruli were ascertained by counting
obsolescent glomeruli by a semi-quantitative scoring technique
and reported as a percentage of approximately 100 glomeruli
examined in each slide. Image-Pro Plus 4.0 software (Bethesda,
MD, USA) was used to quantify change in pancreatic islets
area and glomerular area. The pathologist performing histo-
pathological evaluation was masked to the treatment protocol.
Transmission electron microscopy examination
Karnovsky’s fixed tissues of pancreas and liver were washed
in phosphate buffer (0·1 M, pH 7·4, 68C) and post-fixed for
2 h in 1 % osmium tetroxide in the same buffer at 48C. The
specimens were then dehydrated with graded acetone and
embedded in araldite CY212 to make tissue blocks. Sections
(7080 nm) were cut by an ultra microtome (UltraCut E;
Reichert, Austria) and stained with uranyl acetate and lead
acetate and examined under a transmission electron micro-
scope (Morgagni 268 D; Fei Co., the Netherlands) by a
morphologist masked to the treatment protocol.
Statistical analysis
All results were expressed as the means and standard devi-
ations
(12)
. Statistical analysis was performed using SPSS software
package version 11.5.9 (San Francisco, CA, USA). The values
were analysed by one-way ANOVA followed by Bonferroni
post hoc test. P,0·05 was considered statistically significant.
Results
Effect on body weight
Animals’ body weight did not differ significantly between
groups at the end of experiment (Table 1).
Effect on hyperglycaemia, hyperinsulinaemia, insulin
resistance and
b
-cell function
Serum glucose, insulin and HOMA-IR levels were significantly
elevated by 3·3-, 1·6- and 5·2-fold, respectively, in diabetic con-
trol rats compared with the normal controls (Table 1). Naringin
dose dependently attenuated hyperglycaemia, hyperinsulinae-
mia and IR, so that the naringin 100 mg/kg group was not differ-
ent from the standard drug rosiglitazone group (Table 1).
(a1) (b1) (c1)
(a2) (b2) (c2)
(d1) (e1) (f1)
(d2) (e2) (f2)
SG
SG
SG
SG
SG
SG
MC
MC
MC
N
N
N
N
MC
N
N
Fig. 3. Light microscopic study of pancreatic islet (a1 f1, 20 £, scale bar 50 mm) and electron microscopic study of b-cells (a2– f2, 4000 £, scale bar 1mm) in
different experimental groups. (a1 & a2) Normal control; (b1 & b2) diabetic control; (c1–e1 and c2 e2) Naringin 25, 50 and 100 mg/kg per d respectively; (f1 & f2)
rosiglitazone treated. ,b-cells; , inflammatory cells. N, nucleus; MC, mitochondria; SG, secretory granules.
Naringin ameliorates type 2 diabetes in rats 1717
British Journal of Nutrition
Furthermore, as reflected in the results of the glucose tolerance
test followed by the insulin tolerance test, diabetic control
rats developed glucose intolerance and IR (Fig. 2a and b).
In contrast, naringin at 50 and 100 mg/kg per d corrected
impaired glucose utilisation and insulin insensitivity in diabetic
rats (Fig. 2a and b). In addition, diabetic rats showed reduced
(5·4-fold) b-cell function (HOMA-B), which was significantly
(P,0·05) improved by naringin at 100 mg/kg per d and rosi-
glitazone (Table 1).
Effect on dyslipidaemia and NEFA
Compared with normal controls, we found a 3·2-fold increase
in serum TC, a 6·7-fold increase in TAG, a 3·2-fold increase in
LDL-C and a 4·2-fold increased in NEFA in HFD-STZ-treated
diabetic control rats (Fig. 2c). Naringin at 50 and 100 mg/kg
significantly (P,0·001) decreased TC (5·8 (SD 0·6); 5·0 (SD
0·7) v. 9·4 (SD 1·3) mmol/l), TAG (2·6 (SD 0·5); 2·3 (SD 0·3) v.
4·6 (SD 1·1) mmol/l), LDL-C (3·0 (SD 0·4); 2·6 (SD 0·3) v.4·5
(SD 0·6) mmol/l) and NEFA (1·78 (SD 0·34) mmol/l; 1·02 (SD
0·15) v. 3·13 (SD 0·53) mmol/l) compared with diabetic control
rats and the effect was more pronounced than with rosiglita-
zone. In addition, naringin at 100 mg/kg showed a significant
(P,0·01) increase in HDL-C (1·07 (SD 0·06) v. 0·86 (SD 0·09)
mmol/l) compared with diabetic rats (Fig. 2c).
Effect on islets cell damage, islet atrophy and
b
-cell failure
Pancreatic histology of the normal control rats showed normal
islet cell mass (Fig. 3a1); whereas diabetic control rats had
severe damage on endocrine pancreas, including significant
reduction of pancreatic islet area and atrophy of cells
(Fig. 3b1; Table 1). Middle and high doses of naringin repaired
the injury of pancreatic islet cells and naringin at 100 mg/kg
and rosiglitazone increased islet area to near-normal control
levels (Fig. 3d1, 3e1; Table 1). Furthermore, on electron micro-
scopic analysis compared with normal control (Fig. 3a2),
b-cells from HFD-STZ-treated rats displayed a significantly dis-
tended endoplasmic reticulum (ER) and reduced insulin
secretory granule numbers (Fig. 3b2). Treatment with naringin
100 mg/kg per d and rosiglitazone attenuated ER distension
and preserved granule content, suggesting that naringin pre-
serves b-cell function by maintaining an adequate pool of
secretory granules (Fig. 3e2 and 3f2).
Effect on hydropic change of renal proximal convoluted
tubules and glomerular sclerosis
In our study, diabetic control rats showed substantially hydro-
pic change in proximal convoluted tubules and more widen-
ing of matrix (Fig. 4b) and administration of naringin at
100 mg/kg reversed these changes to near-nor mal control
(Fig. 4e). Furthermore, naringin at 100 mg/kg significantly atte-
nuated glomerular sclerosis, though there was no substantial
difference in glomerular area between groups (Table 1).
Effect on protein expression of PPAR
g
in liver and kidney,
heat shock protein-27, -72 and NF-
k
B in pancreas, liver and
kidney, and insulin receptor substrate 1, phosphorylated
tyrosine 612 IRS1 (Tyr162), liver X receptor and sterol
regulatory element binding protein-1c in liver
HFD-STZ treatment decreased PPARgexpression in liver and
kidney, while naringin at 50, 100 mg/kg and rosiglitazone
significantly (P,0·01) increased PPARgexpression (Fig. 5a).
(a) (b) (c)
(d) (e) (f)
Fig. 4. Light microscopic study of kidney tissue in different experimental groups (a– f, scale bar 50 mm). (a) Normal control (10 £); (b) diabetic control (20 £);
(c– e) Naringin 25, 50 and 100mg/kg per d (10 £); (f) rosiglitazone treated (10 £). , widening of matrix; , hydropic changes.
A. K. Sharma et al.1718
British Journal of Nutrition
Further, the decreased IR in naringin (50 and 100 mg/kg)-treated
rats was also associated with significantly increased P-IRS1
(Tyr162) expression without any change in total IRS1 expression
in liver (Fig. 5b). Interestingly, in our study naringin treatment
significantly increased the expression of HSP-72 (Fig. 5c) and
HSP-27 (Fig. 5d), and the effect of naringin at 100 mg/kg was
more pronounced than with rosiglitazone. Thus, it is very
likely that one of the mechanisms by which naringin improves
type 2 diabetes is by induction of HSP-72 and HSP-27.
To better understand the molecular mechanism underlying
anti-inflammatory activity of naringin, we assessed NF-kBexpre-
ssion. Naringin treatment decreased the NF-kB protein expression
in pancreas, liver and kidney dose dependently, which was
found to be overexpressed in HFD-STZ-induced diabetic rats
(Fig. 5e). Moreover, in our study observed hepatic steatosis in dia-
betic control rats was linked with increased expression of LXRa
and SREBP-1c protein, which were significantly (P,0·001) atten-
uated by naringin administered at 50 and 100mg/kg (Fig. 5f).
Effect on serum TNF-
a
, IL-6, C-reactive protein
and adiponectin
Serum TNF-a, IL-6 and CRP concentrations were significantly
elevated by 6·6-, 2·5- and 3·8-fold, respectively, in diabetic
control rats compared with normal controls(Table 1). In contrast,
serum adiponectin levels were significantly (P,0·001) de-
creased in diabetic control rats. Naringin reduced TNF-a, IL-6,
CRP levels and increased adiponectin levels dose dependently,
though the effect was significant at 50 and 100 mg/kg (Table 1).
Effect on thiobarbituric acid-reactive substances,
superoxide dismutase and glutathione peroxidase
Table 2 represents the effect of naringin on the activities of
enzymatic antioxidants and TBARS contents in serum, pan-
creas, liver and kidney. Diabetic control rats showed signifi-
cant (P,0·001) reductions in the activities of SOD, GSH-Px
Liver
(a) (b)
(c) (d)
(e) (f)
Liver
Liver
Kidney
***
*
**
†††† ††
†† ††
††
*
*
†††
††† †††
†††
††† †††
†††
†††
†††
†††
†††
†††
†††
†††
†††
†††
†††
†††
†††
†††
†††
†††
†††
†††
†††
††
††
***
***
*** ***
***
†††
††† ††† †††
†††
†††
††
††
350
300
250
200
150
100
50
0
300
250
200
150
100
50
0
350
300
450
Pancreas
Liver
Kidney
Pancreas
Liver
Kidney
Pancreas
Liver
Liver
Liver
Kidney
400
250
200
150
100
50
0
350
300
400
250
200
150
100
50
0
250
200
150
100
50
0
250
200
150
100
50
0
PPARγ β-Actin (%)HSP-72/β-Actin (%)
HSP-27/β-Actin (%)
NF-κB
p
65/β-Actin (%)
NF-κB
p
65
NF-κB
p
65
NF-κB
p
65
SREBP-1c and LXRα/
β-Actin (%)
β-Actin
β-Actin
β-Actin
SREBP-1c
LXRα
β-Actin
HSP-72
HSP-72
HSP-72 β-Actin
HSP-27
HSP-27
HSP-27
P-IRS (tyr612)/
β-actin (%)
P-IRS1 (Tyr612)
Normal control
Diabetic control
Diabetic + N25
Diabetic + N50
Diabetic + N100
Diabetic + RSG5
Normal control
Diabetic control
Diabetic + N25
Diabetic + N50
Diabetic + N100
Diabetic + RSG5
Normal control
Diabetic control
Diabetic + N25
Diabetic + N50
Diabetic + N100
Diabetic + RSG5
Normal control
Diabetic control
Diabetic + N25
Diabetic + N50
Diabetic + N100
Diabetic + RSG5
Normal control
Diabetic control
Diabetic + N25
Diabetic + N50
Diabetic + N100
Diabetic + RSG5
Normal control
Diabetic control
Diabetic + N25
Diabetic + N50
Diabetic + N100
Diabetic + RSG5
PPARγ
PPARγ
β-Actin
IRS1
Fig. 5. Various protein expressions in different experimental groups. (a) PPARgexpression in liver ( ) and kidney ( ); (b) total insulin receptor substrate 1 (IRS1,
) and phosphorylated tyrosine 612 IRS1 (P-IRS1; Tyr612, ) expression in liver; (c) heat shock protein (HSP)-72 expression in pancreas ( ), liver ( ) and kidney
(); (d) HSP-27 expression in pancreas ( ), liver ( ) and kidney ( ); (e) NF-kB expression in pancreas ( ), liver ( ) and kidney ( ); (f) sterol regulatory element
binding protein-1c (SREBP-1c, ) and liver X receptor-a(LXRa, ) protein expression in liver. Data are expressed as a ratio of normal control value (set to
100 %). Values are means, with their standard deviations represented by vertical bars (n12). Mean values were significantly different from those of the normal
control: *P,0·05, **P¼0·01 by one-way ANOVA followed by Bonferroni post hoc test. Mean values were significantly different from those of the diet control:
P,0·05, ††P,0·01, †††P,0·001 by one-way ANOVA followed by Bonferroni post hoc test.
Naringin ameliorates type 2 diabetes in rats 1719
British Journal of Nutrition
and an increase in the level of TBARS. When these diabetic
rats were treated with naringin, we observed a significant
(P,0·05) decrease in TBARS and increase in SOD and GSH-Px
activities in serum, pancreas, liver and kidney compared with
diabetic control rats in a dose-dependent manner. However,
naringin (100 mg/kg per d)-induced an increase in enzymatic
antioxidant activity and a decrease in TBARS which were
comparable with normal control and rosiglitazone groups.
Effect on hepatic steatosis
To assess the impact of naringin on changes in liver mor-
phology in HFD-STZ-induced diabetic rats, we performed
histopathological and ultrastructural analysis of liver tissue.
Diabetic control rats showed increased inflammatory cells
and a microvesicular hepatic steatosis on histopathological
examination (Table 3). Furthermore, the electron microscopic
analysis of hepatocyte revealed rarefied matrix, swelling of
rough ER cisternae and mitochondria, fusion or loss of mito-
chondrial cristae, degranulation of rough ER and lipid accumu-
lation in diabetic rats (data not shown). Naringin at 100 mg/kg
per d reversed these pathological changes, and the effect was
comparable with the normal control group (Table 3).
Discussion
In the present study, we have demonstrated that naringin
ameliorates IR, b-cell dysfunction, dyslipidaemia, hepatic
steatosis and pathological changes in kidney in HFD-STZ-
induced type 2 diabetic rats in a dose-dependent manner. Fur-
thermore, we established that this beneficial effect of naringin
is because of attenuation of oxidative stress, CRP, ‘offensive’
adipocytokines TNF-a, IL-6 and correction of ‘defensive’
adipocytokine adiponectin. Importantly, improved glycaemic
control with naringin is associated with increased PPARg,
HSP-27, HSP-72 and decreased NF-kB protein expression.
Herein, we also demonstrated that attenuation of hepatic stea-
tosis by naringin is linked with decreased hepatic LXRaand
SREBP-1c protein expression. Our findings as well as those
shown by other authors
(24 – 26)
have demonstrated that ani-
mals, when fed with a HFD and injected with a low dose of
STZ, display many characteristics of IR including hyperglycae-
mia, hyperinsulinaemia, impaired glucose tolerance test and
insulin tolerance test, decreased b-cell function (HOMA-B),
hyperlipidaemia, hepatic steatosis and kidney damage.
The primary basis of T2DM pathogenesis is unknown; how-
ever, there is a general agreement that IR followed by b-cell
failure is a major event in the development of the disease
(4)
.
Interestingly, in our study naringin treatment not only
decreased hyperglycaemia, hyperinsulinaemia and IR, but
also improved b-cell function and b-cell mass. Microscopic
examination of endocrine pancreas revealed decreased
b-cell mass, swelling of ER and diminished secretory granules
in b-cells of HFD-STZ-treated rats indicating b-cell failure,
which were mitigated by naringin. Our results suggest that
reversal of IR and b-cell failure by naringin in HFD-STZ-
induced diabetic rats is tightly associated with overexpression
of PPARg, HSP-27 and HSP-72. The accumulated evidence
Table 2. Lipid peroxidation and antioxidant parameters in different experimental groups
(Mean values with their standard deviations; n12)
Normal control Diabetic control Diabetic þN25 Diabetic þN50 Diabetic þN100 Diabetic þRSG5
Parameters Mean SD Mean SD Mean SD Mean SD Mean SD Mean SD
TBARS (mmol/g)
Serum 3·57 0·63 6·86* 1·03 5·44 1·49 4·36†† 0·86 3·98††† 0·32 4·13†† 0·54
Pancreas 2·13 0·35 9·63* 0·74 8·21† 0·60 4·51††† 0·41 3·07††† 0·69 3·73††† 0·30
Liver 8·39 1·80 24·53* 3·08 20·92 5·63 16·37††† 2·06 12·04††† 1·42 12·87††† 2·04
Kidney 6·45 2·04 18·78* 2·74 17·26 3·07 12·89††† 1·12 9·07††† 1·07 12·17††† 1·23
SOD (U/mg protein)
Serum 134·5 10·8 102·7* 12·5 113·2 8·6 118·9†† 7·2 126·9††† 8·4 119·5†† 8·0
Pancreas 63·2 5·4 34·9* 7·4 41·0 5·7 48·5†† 4·6 54·1††† 7·3 50·3††† 6·9
Liver 142·5 9·0 91·0* 10·3 102·4 11·5 12·4†† 13·5 132·0††† 8·4 129·1††† 13·5
Kidney 125·6 7·1 87·2* 9·5 105·8† 12·6 114·8††† 10·7 118·2††† 9·1 117·4††† 15·4
GSH-Px (U/mg protein)
Serum 14·02 2·45 10·45* 0·34 10·33 0·78 11·47† 0·46 12·71†† 0·67 12·80†† 5·9
Pancreas 1·64 0·22 0·76* 0·17 0·83 0·13 1·32††† 0·10 1·45††† 0·29 1·37††† 0·41
Liver 20·71 3·12 11·26* 2·43 13·02 4·05 15·14†† 2·85 16·07†† 2·83 14·69† 2·11
Kidney 15·65 2·46 7·09* 3·04 8·35 2·06 10·77†† 3·10 12·89††† 1·83 12·08††† 2·23
TBARS, thiobarbituric acid-reactive substances; SOD, superoxide dismutase; GSH-Px, glutathione peroxidase.
Mean values were significantly different from those of the normal control (*P,0·001 by one-way ANOVA followed by Bonferroni post hoc test)
Mean values were significantly different from those of the diabetic control: †P,0·05, ††P,0·01, †††P,0·001 by one-way ANOVA followed by Bonferroni post hoc test.
A. K. Sharma et al.1720
British Journal of Nutrition
suggests that PPARgactivation improves b-cell function and IR
by decreasing NEFA and pro-inflammatory cytokines, conse-
quently sparing skeletal muscle, liver and b-cells from detri-
mental metabolic effects of their high concentration
(4,13,14,27)
.
Notably, our findings support the hypothesis that stress pro-
teins play a significant role in the pathobiology of IR and
T2DM, as expression of HSP-72 and HSP-27 decreased in dia-
betic rats, and their expression was increased along with the
amelioration of diabetes in naringin- and rosiglitazone-treated
rats. HSP-27 and HSP-72 are ubiquitous molecular chaperones
induced in response to stressful conditions, such as inflam-
mation, hypoxia, hyperglycaemia, oxidative stress, and pre-
clude cell damage
(15 – 17,28)
. Recently, it has been proposed
that HSP-72 protects against obesity-induced hyperglycaemia,
hyperinsulinaemia and IR by inhibition of c-jun amino term-
inal kinase and inhibitor of kB kinase
(15)
, which are critical
inflammatory kinases in the development of IR and T2DM
(2)
.
Similarly, various studies
(16,17)
have reported that overexpres-
sion of HSP-72 and HSP-27 prevents the phosphorylation of
inhibitor kB, activation of NF-
k
Band TNF-
a
gene transcrip-
tion. Consistent with this, an interesting finding in our study
was that naringin showed enhanced expression of HSP-27
and HSP-72 along with waning of NF-kB expression, which
might be a plausible mechanism involved in its better or simi-
lar response to a rosiglitazone even though comparatively
lesser expression of PPARg.
In addition, abnormal production of adipocytokines, reac-
tive oxygen species and advanced glycated end products in
diabetes leads to NF-kB activation and increased production
of inflammatory mediators such as TNF-aand IL-6
(29)
. Many
aspects of biological processes, such as immune and inflam-
matory responses, as well as cell growth and apoptosis, are
in part regulated by the NF-kB system
(29,30)
. So we postulated
that amelioration of b-cell damage and IR by naringin might
be directly or indirectly associated with modulation of
NF-kB expression. Interestingly, in our study naringin admin-
istration dose-dependently attenuated NF-kB expression and
inflammatory mediators in diabetic rats, which shows its
potential importance in metabolic diseases and T2DM.
However, this inhibition of NF-kB activity is probably a
consequence of increased PPARg, HSP-72 and HSP-27
expression by naringin treatment
(4,13 – 17)
. Furthermore, IRS
protein isoform, IRS1, is a decisive link in hepatic insulin
signalling and increased serine/threonine phosphorylation in
response to TNF-a, and various stresses in diabetes may be a
key molecular lesion in hepatic IR
(31)
. In line with this assump-
tion, our results indicate that the molecular mechanisms
underlying IR in diabetes may involve TNF-a-mediated
activation of pro-inflammatory kinases (NF-kB) and phospha-
tases that inhibit tyrosine phosphorylation of IRS1 and insulin
action. Conversely, an enhanced level of P-IRS1 (Tyr162) and
improved action of insulin by naringin seem to be a result of
antagonism of NF-kB activation, and this might be a conse-
quence of increased PPARg, HSP-72 and HSP-27 expression
with naringin
(4,13 – 17)
.
To elucidate the role of oxidative stress in HFD-STZ-treated
diabetic rats and naringin-treated rats, we assessed oxidative/
antioxidant status by measuring reactive oxygen species-
induced products of lipid peroxidation (TBARS) and activity
of SOD and GSH-Px in serum, pancreas, liver and kidney. Con-
sistent with previous studies
(24,26)
, we observed reductions in
the activities of SOD and GSH-Px as well as increased TBARS
in serum and tissues of diabetic rats. Similar to Kim et al.
(10)
,
our findings suggest the antioxidant activity of naringin
increased the antioxidant enzymes and decreased TBARS
levels in serum and tissues. Since increased reactive oxygen
species production by mitochondria and ER leads to the acti-
vation of stress kinases, such as c-jun amino terminal kinase
and inhibitor of kB kinase, the two principal inflammatory
pathways disrupt insulin action by decreasing P-IRS1 (Tyr162)
and developing metabolic diseases
(2,3,32)
. Thus, drugs with anti-
oxidant potential may attenuate T2DM. Furthermore, during
the past decade many researchers have implicated increased
oxidative stress as the major culprit in diabetes pathogenesis
leading to dysregulated production of adipocytokines and
acute-phase reactants
(2,3,31)
. Naringin treatment reduced circu-
lating markers of chronic subacute inflammation, such as CRP,
TNF-aand IL-6, that promote IR and at the same time increased
adiponectin, which has insulin-sensitising activity. We speculate
that naringin inhibits dysregulated production of adipocyto-
kines and CRP in HFD-STZ-induced diabetic rats by decreasing
oxidative stress and by modulation of PPARg, HSP-27, HSP-72
and NF-kB protein expression
(13 – 17,30)
.
In our study diabetic rats showed markedly increased
NEFA, TAG, TC, LDL-C and diminution in HDL-C. IR leads to
decreased apoB100 intracellular degradation, resulting in
increased VLDLapoB production and secretion
(5)
, which sub-
sequently causes increased LDL-C and decreased HDL-C
(33)
.
Further, this increased dyslipidaemia in diabetic rats was also
associated with microvesicular hepatic steatosis, swelling of
ER and mitochondria in hepatocytes, which resembles fatty
hepatic lesions observed in human patients with T2DM.
Hepatic steatosis that often accompanies diabetes may be a
consequence of hyperinsulinaemia and IR causing increased
Table 3. Histological observations of hepatic tissue in experimental groups (n12)
Groups Swelling of hepatic cells Fat accumulation Displacement of nucleus Loss of nucleus Inflammatory cells
Normal control ––– –
Diabetic control 1111 1111 1111 1111 111
Diabetic þN25 1111 1111 1111 1111 111
Diabetic þN50 111 111 11 11 11
Diabetic 1N100 þþþ þþ
Diabetic þRSG5 þþ þþ þ þ þ
N25, naringin at 25 mg/kg per d; N50, naringin at 50 mg/kg per d; N100, naringin at 100 mg/kg per d; RSGS, rosiglitazone at 5 mg/kg per d; þþþþþþ, high
degree; þþþþ þ , severe; þþþþ, moderate; þþþ, fair; þþ, mild; þ, very mild; 2, no changes in histology.
Naringin ameliorates type 2 diabetes in rats 1721
British Journal of Nutrition
expression of LXRaand SREBP-1c, thereby decreased
NEFA oxidation and augmented lipogenesis in hepatocytes
(5)
.
Even increased oxidative stress, serum NEFA and TNF-ain
diabetic rats have also been reported to be crucially linked
with hepatic steatosis
(23,26,31)
. As such, the present results
raise the possibility that the beneficial effect of naringin on
dyslipidaemia and hepatic steatosis may be partly mediated
through the modulation of PPARg, HSP-27, HSP-72 and
NF-kB expression, which in turn leads to decreased LXRa
and SREBP-1c activation, and consequently inhibition of
lipogenic enzymes synthesis, lipogenesis and fat accumulation
in hepatocytes
(5,13 – 15)
. Recently, many studies have suggested
that naringin lowers the plasma and hepatic lipid levels
by suppression of hepatic fatty acid synthase, glucose-6-
phosphate dehydrogenase, HMG-CoA reductase activity, acyl
CoA/cholesterol acyltransferase activities and by increasing
faecal fat
(10,11)
.
Data presented in this study also add to the evidence that
HFD-STZ treatment produces pronounced kidney lesions in
rats because of associated hyperinsulinaemia, hyperlipidae-
mia, hyperglycaemia and increased oxidative stress
(24)
.
However, naringin prevented these pathological alterations
due to its insulin-sensitising, anti-inflammatory, anti-dyslipi-
daemic and antioxidant activity. Herein, we also demonstrated
that the mechanism for such an outcome is modulation of
PPARg, HSP-27, HSP-72 and NF-kB protein expression by
naringin in kidney tissue
(13,16,17)
.
Thus in our study, naringin exerts various pharmacological
effects in HFD-STZ-induced type 2 diabetic rats. Concerning
its toxic effects, it is reported that naringin is safe and
produced no lethality at a very high dose (5000 mg/kg, by
mouth) in mice
(34)
. Moreover, Lambev et al.
(35)
demonstrated
that LD50 of naringin by the ip route in the rat and guinea
pig is 2000 mg/kg. One major drawback associated with
this flavanone glycoside is inhibition as well as the down-
regulation of drug-metabolising cytochrome P450 enzymes
such as CYP3A4 and CYP1A2, which may result in drug
drug interactions in diabetic individuals
(34)
.
In summary, the present study finds naringin as effective as
rosiglitazone and unravels the mechanism whereby naringin
ameliorates IR, b-cell failure, hepatic steatosis and kidney
damage in a rat model of T2DM. Up-regulation of PPARg,
HSP-72, HSP-27 and suppression of NF-kB by naringin prevent
type 2 diabetes and its deleterious effects by attenuating
oxidative stress, inflammation and dysregulated adipocytokine
production in HFD-STZ-induced type 2 diabetic rats. Alto-
gether, these findings indicate that naringin may be an effective
therapeutic strategy for the treatment of diabetes and its
associated complications; further experimental and clinical
studies are required to explore the additional mechanisms and
establish its clinical utility.
Acknowledgements
This study was supported by grant from an institutional
funding agency, AIIMS, New Delhi, India. Special thanks
to Deepak Sharma for his valuable technical assistance. A. K. S.,
S. B., S. O., J. B., S. K. and D. S. A. designed the research.
A. K. S. and S. B. performed the research. N. K. and R. R.
analysed histopathological and electron microscopical
changes. A. K. S., S. B., S. O., J. B., S. K. and D. S. A. analysed
data and wrote the paper. All the authors read and approved
the final submitted manuscript. None of the authors has any
conflict of interest.
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Project
Dear all of you, thanks a lot for your supports and cooperation, given to us for the joint project, mentioned above! The Identifying, Preparation and Drafting a Project Proposal as well its Impleme…" [more]
Project
Objective: Functional genomics for identifications of promoters, genes and epialleles associated with abiotic stress tolerance and nutrient use efficiency of rice and wheat Activities: Molecular an…" [more]
Article
February 2008 · Nippon Jinzo Gakkai shi
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