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See discussions, stats, and author profiles for this publication at: https://www.researchgate.net/publication/221785011

[6]-Gingerol isolated from ginger attenuatessodium arsenite induced oxidative stress and

play s a corrective role in improving insulin

signaling in mice

ARTICLE in TOXICOLOG Y LETTERS · JANUARY 2012

Impact Factor: 3.26 · DOI: 10.1016/j.toxlet.2012.01.002 · Source: PubMed

CITATIONS

34

READS

75

6 AUTHORS, INCLUDING:

Debrup Chakraborty

Michigan State University

12 PUBLICATIONS 191 CITATIONS

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Avinaba Mukherjee

Jadavpur University


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Samrat Ghosh

University of Kalyani


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Anisur Rahman Khuda-Bukhsh

University of Kalyani

315 PUBLICATIONS 2,636 CITATIONS

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Available from: Debrup Chakraborty

Retrieved on: 24 March 2016

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Toxicology Letters 210 (2012) 34–43

Contents lists available at SciVerse ScienceDirect

Toxicology Letters

journal homepage: www.elsevier .com/ locate / toxlet

[6]-Gingerol isolated from ginger attenuates sodium arsenite induced oxidativestress and plays a corrective role in improving insulin signaling in mice

Debrup Chakraborty, Avinaba Mukherjee, Sourav Sikdar, Avijit Paul, Samrat Ghosh,Anisur Rahman Khuda-Bukhsh∗

Cytogenetics andMolecular Biology Laboratory, Department of Zoology, University of Kalyani, Kalyani 741235, West Bengal, India

a r t i c l e i n f o

Article history:

Received 28 November 2011

Received in revised form

30 December 2011

Accepted 2 January 2012

Available online xxx

Keywords:

Sodium arsenite

[6]-Gingerol

Oxidative stress

Hyperglycemia

GLUT4

Insulin signaling

a b s t r a c t

Arsenic toxicity induces type 2 diabetes via stress mediated pathway. In this study, we attempt to reveal

how sodium arsenite (iAs) could induce stress mediated impaired insulin signaling in mice and if an

isolated active fraction of ginger, [6]-gingerol could attenuate the iAs intoxicated hyperglycemic condi-

tion of mice and bring about improvement in their impaired insulin signaling. [6]-Gingerol treatment

reduced elevated blood glucose level and oxidative stress by enhancing activity of super oxide dismutase

(SOD), catalase, glutathione peroxidase (GPx) and GSH. [6]-Gingerol also helped in increasing plasma

insulin level, brought down after iAs exposure. iAs treatment to primary cell culture of -cells andhepatocytes in vitro produced cyto-degenerative effect and accumulated reactive oxygen species (ROS)

in pancreatic -cells and hepatocytes of mice. [6]-Gingerol appeared to inhibit/intervene iAs inducedcyto-degeneration of pancreatic -cells and hepatocytes, helped in scavenging the free radicals. Theover-expression of TNF and IL6 in iAs intoxicated mice was down-regulated by [6]-gingerol treatment.iAs intoxication reduced expression levels of GLUT4, IRS-1, IRS-2, PI3K, AKT, PPAR signaling molecules;[6]-gingerol mediated its action through enhancing the expressions of these signaling molecules, both

at protein and mRNA levels. Thus, our results suggest that [6]-gingerol possesses an anti-hyperglycemicproperty and can improve impaired insulin signaling in arsenic intoxicated mice.

© 2012 Elsevier Ireland Ltd. All rights reserved.

1. Introduction

Arsenic is a naturally occurring heavy metal that is present in

food, soil and water. It is released in the environment from both

natural and man-made sources (Tchounwou et al., 1999). Inorganic

arsenic and theirmetabolites (both As+3 andAs+5 forms) are known

to exert their toxic effects by a variety of mechanisms which may

lead to some serious health problems. Epidemiological data have

shown that chronic exposure of inorganic arsenical compounds to

humans are associatedwith liverinjury,peripheral neuropathy andanincreasedincidence of cancer of thelung, skin,and liver (Leonard

andLauwerys, 1980). In EastAsia alone, including Bangladesh, West

Bengal, India, Vietnam, Thailand and China, more than 30 million

people are chronically exposed to arsenic (Tseng et al., 1968). This

arsenic induced toxicity arises and sustains by generating stress

response through reactive oxygen species formation and antioxi-

dant depletion ( Jomova et al., 2011). According to a recent study,

sodium arsenite (iAs)is found to be associatedwith increasedblood

glucose level in experimental rats (Yousef et al., 2008).

∗ Corresponding author. Tel.: +91 33 25828750x315.

E-mail addresses:khudabukhsh [email protected], prof [email protected]

(A.R. Khuda-Bukhsh).

Hydroarsenicism is a major public health problem since mil-

lions of people worldwide are exposed to arsenic by drinking of

contaminated water ( Jones, 2007). Studies on mouse bone marrow

cells have predicted an increased level of chromosomal abnor-

mality and micronucleus formation after treatment with arsenic

(Banerjee et al., 2007) andtherebyhaveconfirmed itscytotoxic and

cytodegenerative effects. One of the plausible modes of action of

arsenic toxicity is by oxidative stress since it can stimulate produc-

tion of reactive oxygen species (ROS), resulting from an imbalance

between antioxidants and oxidants during arsenic metabolism(Goering et al., 1999; Sun et al., 2006).

On the other hand arsenic has been recently proposed as

an additional risk factor for diabetes (Silbergeld et al., 2008;

Longnecker and Daniels, 2001). According to recent surveys it is

found that the occurrence of diabetes is significantly higher in

arsenic-endemic villages in Taiwan and India than in the general

population (Zimmet, 1982; Wang et al., 1997; Belon et al., 2006).

The prevalence of diabetesmellituswas 2-fold higher in these areas

than in Taipei City and the Taiwan area in general.

From in vitro studies, the impairment of insulin secretion

(Diaz-Villaseñor et al., 2006) and the induction of oxidative stress

(Izquierdo-Vega et al., 2006) have been postulated for arsenic-

induced type 2 diabetes. Induction of stress via generation of free

oxygen radicals and antioxidant depletionled into this process and

0378-4274/$ – seefront matter © 2012 Elsevier Ireland Ltd. All rights reserved.

doi:10.1016/j.toxlet.2012.01.002

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36 D. Chakraborty et al./ Toxicology Letters 210 (2012) 34–43

Fig. 1. Mass spectra and structure of [6]-gingerol.

2.8. Animal treatment

After selectionof theoptimum dose of iAs(3 mg/kg), animals were randomized

and 36 mice were divided into six groups, consisting of six mice in each group and

they were treated for 15 weeks as follows:

Group 1: Normal control: animals received only water as vehicle.

Group 2: Arsenic control (iAs): animals received iAs at 3 mg/kg body weight oncedaily for12 weeks, orally.

Group 3: [6]-Gingeroltreated group (iAs+ [6]-gingerol): received iAs for12 weeks

followed by [6]-gingeroladministrationat a dose of 50mg/kgbody weightin alco-

hol once daily for next 3 weeks.

Group 4: [6]-Gingeroltreated group (iAs+ [6]-gingerol): received iAs for12 weeks

followed by [6]-gingeroladministrationat a dose of 75mg/kgbody weightin alco-

hol once daily for next 3 weeks.

Group 5: Alcohol treated group (iAs+ alcohol): received iAs for12 weeks followed

by alcohol administration (equal quantity as administeredin groups 3 and4) once

dailyfor next 3 weeks.

Group 6: [6]-Gingerol alone treated group: normal mice were treated with [6]-

gingerol (orally, 75mg/kg body weight, once daily)for 3 weeks to see whether the

drug alone has anyadverse effect on mice.

The animals were humanely sacrificed under light ether anesthesia and livers

were collected.

We fedthe drug orallyto all themice of differentexperimental groups throughgavage. No significant changes in ALT, AST activity were found between iAs control

and alcohol (drug vehicle) treated groups. There were also no significant differ-

ences of ALT and AST activities between normal control and [6]-gingerol alone

treated groups. Therefore, we excluded groups 5 and 6 from more in-depth stud-

ies. Mice showing blood glucose levels of more than 180mg/dl were considered as

hyperglycemic animals and included in our experiments.

2.9. Collection of blood and tissue samples

At the end of the experimental period, we kept t he animals on f as t f or 12h

prior tobeingsacrificed fordetermining glucoselevel intheirfasting blood.We also

collected the pancreas and liver tissues from each experimental animal and stored

them at −80 ◦C for further analysis.

2.10. Preparation of liver tissue homogenates

We collected the liver tissues from experimental mice, homogenized them in

lysisbuffer usingglass homogenizer and centrifugedat 12,000× g for30 min at 4 ◦C.

We collectedthe supernatant aftercentrifugation and usedit forfurtherexperiment

andestimated thetotal proteinaccordingto themethodof Bradfordusing crystalline

bovineserum albumin(BSA) as standard (Bradford, 1976).

2.11. Determination of pancreatic and hepatic arsenic contents

The arsenic contents of liver tissues of all experimental animals were analyzed

according to themethod described by Khuda-Bukhshet al. (2005), using an atomic

absorption spectrophotometer (AAS).

2.12. Determination of in vivo antioxidant capacity

We determined antioxidant capacity of [6]-gingerol on hepatic tissues of all

experimental animals by FRAP (ferric reducing antioxidantpotential) assay( Benzie

and Strain, 1999). We took the absorbance of the sample against reagent blank

(1.5ml FRAP reagent +50 l distilled water)at 593nm (Manna et al., 2009).

2.13. Biochemical analysis of blood and activity of antioxidant markers in liver

We estimated thebloodglucose level using a glucose estimation kit(Accu-chek

active), Rochediagnostics, Mannheim,Germany.We used livertissue homogenates

for various enzymatic assays. We undertookspectrophotometricanalysisof activity

of catalase (CAT) (Aebi, 1984), super oxide dismutase (SOD) (Fridovich, 1989), glu-

tathione peroxidase(GPx),level of totalglutathione (GSH) (Ellman, 1959) according

to the standard protocols.

2.14. Oral glucose tolerance test (OGTT)

Anoral glucose tolerancetest (OGTT) wasperformed onthe lastday of treatment

after overnight fasting. Blood wascollected from the tail vein of mice at time 0, 60,

90 and120 min after anoral glucose load of 3.0g/kgof body weight. Only water was

provided inside the cages during thecourse of experiment.

2.15. ELISA for activity measurement of different antibodies

We assayed the activity of plasma insulin and hepatic GLUT4 according to

the manufacturer’s protocol (Santa Cruz Biotechnology, Inc., USA) and quantified

t he m using an ELIS A r eader ( The rm o Scie nt ifi c, USA). We used pNPP (para-

nitrophenylphosphate) asa colordevelopingagent andmeasuredthe colorintensity

in 405nm wavelength.

2.16. Immunoblot analysis

We usedthe livertissue homogenates for immunoblotanalysis. Forthis we took

50mg oftissuesamplesin2mllysisbufferforproteinextraction.We undertookSDS-

PAGE (12.5%) electrophoresis of equal amounts of lysate protein and transferred

them to polyvinyl difluoride (PVDF) membrane. After blocking with 3% BSA, we


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D. Chakraborty et al./ Toxicology Letters 210 (2012) 34–43 37

Fig. 2. Effect of [6]-gingerol on the viability of iAs intoxicated hepatocytes and

pancreatic -cells. While the-cells were exposed to 10M iAs and differentcon-

centrations of [6]-gingerol for 72h, the hepatocytes were exposed for 8 h. The cell

viability was thendetected by MTT assay. Eachpoint expressedas mean±SD (N =6).

Significance, * p< 0.05 vs.normal control group. Significance,# p


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Fig. 3. Dose dependent inhibitory effect of [6]-gingerol on iAs induced free radical accumulation. 1 h after incubation with iAs (10M) cells were incubated with 50 and

75g/ml doses of [6]-gingerol. -Cells and hepatocytes incubated for next 71h and 7h, respectively with drug. A–D represents fluorescence microscopic observations

and E–H represents flowcytometric analysis of ROS accumulation in -cells. On the other hand, I–L represents fluorescence microscopic observations and M–P represents

flowcytometric analysis of ROSaccumulation in hepatocytes.A, E, I, M – normalcontrol cells,B, F, J, N – 10M iAsintoxicatedcells, C, G, K, O – iAsintoxicatedcells, incubated

with 50g/ml [6]-gingerol, D, H, L, P – iAs intoxicated cells, incubated with 75g/ml [6]-gingerol. The intra-cellular ROS was detected by DCFHDA method. Q–T represents

flowcytometric analysis of intracellular GLUT4 content in hepatocytes. Q – normal control cells, R – 10M iAs intoxicated cells, S – iAs intoxicated cells, incubated with

50g/ml [6]-gingerol, T – iAs intoxicated cells, incubated with 75g/ml [6]-gingerol.

Table 1

Effect of different concentrations of iAs on mice.

Cont 2 mg/kg 3 mg/kg 4 mg/kg 5 mg/kg

ALT (m/mg protein) 11.06 ± 3.10 16.6 ± 3.109 23.24 ± 0.560* 25.04 ± 0.66* 27.93 ± 1.17*

AST (m/mg protein) 5.33 ± 0.203 6.55 ± 1.66 14.007 ± 0.509* 13.86 ± 0.441* 17.32 ± 2.68*

Blood glucose (mg/dl) 92.5 ± 2.12 132 ± 5.65* 189.5 ± 7.77* 186 ± 8.48* 186 ± 9.89*

Data are expressedas mean±SD (N =6).* p< 0.05 vs. normal control group.


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Table 2

Dose dependent Influence of [6]-gingerol on iAs intoxicated hyperglycemic mice.

Cont iAs 25 mg/kg 50 mg/kg 75 mg/kg

ALT (m/mg protein) 11.06 ± 3.109 23.24 ± 0.56* 21.87 ± 1.37 15.55 ± 1.35# 13.51 ± 0.45#

AST (m/mg protein) 5.33 ± 0.203 14.007 ± 0.51* 12.78 ± 1.08 8.68 ± 0.71# 8.29 ± 2.2#

Blood g lucose (mg/dl) 92.5 ± 2.12 189.5 ± 7.77* 182 ± 8.48 122.5 ± 3.53# 110 ± 2.82#

Data are expressed as mean±SD (N =6).* p< 0.05 vs. normal control group.

# Significance, p< 0.05 vs. iAs intoxicated group.

Table 3

Role of [6]-gingerolon arsenic deposition in pancreas and liver tissues of experimental animals expressed in nmol/g unit.

Cont iAs 50 mg/kg 75 mg/kg

Pancreas 13.87 ± 4.2 43.015 ± 6.86* 26.435 ± 3.41 23.669 ± 3.74#

Liver 14.92 ± 6.71 49.27 ± 7.73* 27.51 ± 3.11# 25.13 ± 4.14#


# Significance, p< 0.05 vs. iAs intoxicated diabetic group.

Table 4

Role of [6]-gingerolon different anti-oxidant biomarkers of iAs intoxicated mice liver.

Cont iAs 50 mg/kg 75 mg/kg

GHb (%) 4.02 ± 0.28 10 ± 0.19* 7.05 ± 0.27# 5.32 ± 0.97#

SOD (m/mg protein) 38.68 ± 1.26 28.32 ± 1.73* 32.19 ± 1.29 35.35 ± 0.35#

Catalase (m/mg protein) 72.22 ± 4.08 37.2032 ± 2.97* 52.16 ± 3.54# 58.547 ± 0.95#

GSH (m/mg protein) 37.62 ± 2.65 21.92 ± 1.21* 28.53 ± 1.4 32.29 ± 3.65#

GPx (m/mg protein) 64.5 ± 5.82 48.36 ± 1.44* 54.16 ± 1.59 58.18 ± 2.36#

FRAP (%) 95.83 ± 5.89 54.16 ± 5.89* 67.91 ± 1.76 79.16 ± 5.89#


# Significance, p< 0.05 vs. iAs intoxicated diabetic group.

3.9. Dose dependent study of [6]-gingerol by FRAP assay

Dose dependent effect of [6]-gingerol against iAs toxicity has

been represented in Table 4. The intracellular ferric reducing

antioxidant potential decliningafter iAsintoxicationin animalswas

found to increase significantly with[6]-gingerol treatment at doses

of 50mg/kg and 75mg/kg bw, respectively.

3.10. Effect on plasma insulin and hepatic GLUT4 content

Under iAs intoxicated hyperglycemic condition, decreased lev-

els of plasma insulin andhepaticGLUT4were found. Administration

Fig.4. Impactof [6]-gingerol(50 and 75 mg/kg)treatment on oralglucose tolerance

in iAs mice. Data were expressed as mean±SD (N =6), Ap< 0.05 cont vs. iAs intox-

icated group and ap< 0.05 iAs vs. drug groups (A/a used to denote comperisons in

0 min interval groups, B/b used to denote comperisons in 60min interval groups,

C/c used to denotecomperisonsin 90min interval groups, D/dused to denotecom-

perisons in 120min interval groups).

of [6]-gingerol at both the doses increased the plasma insulin and

hepatic GLUT4 concentrations significantly (Fig. 5).

3.11. Immunoblot analysis

Administration of the drug down regulated the expression of

TNFwhichwas earlier found to increase in iAsintoxicated liver.Inaddition,we observedthat iAsintoxicationreducedthe expressions

of proteins related to insulin signaling, namely, Insulin, IRS1, IRS2,

AKT, PI3K, GLUT4 and PPAR (Figs. 6–7). However, expressions of these proteins were significantly up regulated after treatment with

[6]-gingerol.

Fig.5. Effectof [6]-gingerolon plasmainsulinand hepatic GLUT4 levelin iAsintoxi-

cateddiabeticmice.Data wereexpressed in histogramas mean±SD (N =6).* p


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Fig. 6. Immunoblot analysis of insulin, IRS1, TNF, IRS2, GLUT4, PI3K, AKT, PPAR

andGAPDH. GAPDH is used as an equal loading housekeepinggene.Lane 1. Normal

mice; Lane 2. iAs-intoxicated mice, Lane 3. iAs-intoxicated+ [6]-gingerol 50mg/kg

bw, Lane 4. iAs-intoxicated+ [6]-gingerol75 mg/kg bw. Signs indicate that a partic-

ular treatmenthas not(−) been or has been (+)given,respectively.

3.12. RT-PCR analysis

Results of RT-PCR confirmed that there was a significant dif-

ference in the expressions of mRNA between the control and

the iAs-intoxicated groups and between the iAs and drug-treated

groups (Figs. 8–9). Results of RT-PCR analysis also supportedthe results obtained from the western blot analysis. The primer

sequences of amplified genes are given in Table 5.

4. Discussion

Our experimental findings showed that iAs intoxication can

significantly reduce the pancreatic -cells and hepatocyte cell via-bility. But iAs intoxicated cells incubated with [6]-gingerol had the

Table 5

Primer sequences.

Primer name Primer s equences

TNF Fwd 5-GCACAGAAAGCATGATCCGC-3

Rev 5-CTTGGTGGTTTGCTACGACG-3

IL6 Fwd 5-AACGATGATGCACTTGCAGA-3

Rev 5-GAGCATTGGAAATTGGGGTA-3

IRS1 Fwd 5-AGAGTGGTGGAGTTGAGTTG-3

Rev 5-GGTGTAACAGAAGCAGAAGC-3

IRS2 Fwd 5-GGATAATGGTGACTATACCGAGA-3

Rev 5-CTCACATCGATGGCGATATAGTT-3

PI3K Fwd 5-TTAAACGCGAAGGCAACGA-3

Rev 5-CAGTCTCCTCCTGCTGCTGAT-3

AKT Fwd 5-CCTGGACTACCTGCACTCTCGGAA-3

Rev 5-TTGCTTTCAGGGCTGCTCAAGAAGG-3

GLUT4 Fwd 5-AAGATGGCCACGGAGAGAG-3

Rev 5-GTGGGTTGTGGCAGTGAGTC-3

GAPDH Fwd 5-CCATGTTCGTCATGGGTGTGAACCA-3

Rev 5-GCCAGTAGAGGCAGGGATGATGTTC-3

PPAR Fwd 5-GCGGAGATCTCCAGTGATATC-3

Rev 5-TCAGCGACTGGGACTTTTCT-3

potential to partially overcome the iAs induced toxicity and couldreduce the cell death, in vitro. It is reported that arsenic produces

hepatocyte dysfunction including the reduced cell viability (Das

et al., 2010). On the other hand, ROS also plays an important role

in insulin resistance which is a highly prevalent condition impli-

cated in the development of diabetes. We assessed the changes

in free radical accumulation in -cells and hepatocytes and foundan increased level of ROS accumulation in cells intoxicated with

only iAs. On the other hand,treatment with [6]-gingerol along with

iAs intoxication showed lesser amount of ROS accumulation and

significantly increased cell viability in both pancreatic -cells andhepatocytes.

Recently researchers publishedthat arsenic has a profoundtoxic

and cytodegenerative effects on pancreas and liver (Pi et al., 2002;

Santraetal.,2000). Oneof themajor reasons behind arsenic inducedhepatotoxicity is the depletion of antioxidant defense mechanism

which is related to the reductionof anti-oxidative enzymes’ activity

includingSOD,CAT,GPxandGSH(Das etal., 2010). Arsenic in differ-

ent forms also increases the levels of free hydroxyl radicals, which

playan importantrole in the development of genotoxicity(Liuet al.,

2001). Therefore, any agent which by itself is non-toxic, but has

the ability to reduce oxidative stress will be desirable to antago-

nize ROS generation and also thereby genotoxicity. Our findings

Fig. 7. Relative band intensities of immunoblots.The relative intensities of bands were determined using ‘ImageJ’ software.The results shown in histograms arethe average

±SD (N = 6). Significance * p< 0.05 iAs-intoxicated vs. normal control group; significance # p< 0.05 drug-fed vs. iAs-intoxicated group.


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Fig. 8. Reverse transcription polymerase chain reaction analysis of TNF, IL6,

IRS1, I RS2, PI 3K, AKT, PP AR and GA PDH. Lane 1. Normal mic e; La ne 2. iAs-

intoxicated mice, Lane 3. iAs-intoxicated+ [6]-gingerol 50mg/kg bw, Lane 4.

iAs-intoxicated+ [6]-gingerol 75mg/kg bw. Signs indicate that a particular treat-

menthas not (−) been or has been (+)given,respectively.

indicate that the administration of [6]-gingerol reduced the oxida-

tive stress as revealed from the increased activity of antioxidant

biomarkers like CAT, SOD, GPx and GSH. On the other hand, it also

had an anti-hyperglycemic effect as the increased blood glucose

level due to iAs induction was found to decrease up to the normal

level after treatment with [6]-gingerol. In addition, mice treated

with [6]-gingerol also showed significant improvement in oral glu-

cose tolerance. GHbis an importantindex of diabetes management

and we observed a significant fall in GHb after administration of

[6]-gingerol in iAs intoxicated hyperglycemic mice. The in vivo

antioxidant potential of [6]-gingerol in liver tissue was determined

by FRAP assay, which indicates the anti-oxidative potentials of

[6]-gingerol at two different concentrations. The analysis of AAS

data on arsenic content of liver and pancreas of arsenic intoxicated

mice before and after administration of [6]-gingerol confirmed the

ability of this drug to removethe accumulatedarsenic content from

these organs.

According to several authors (Cemek et al., 2008; Singh et al.,

2009), a drug which has the combined anti-oxidative and anti-hyperglycemic properties can make the drug very effective as

an anti-diabetic drug. Pancreatic -cell dysfunction due to iAstoxicity is the main reason behind impaired insulin secretion

which plays an important role in the onset of type 2 diabetes. In

in vivo set of experiment, treatment with both the doses (50mg/kg

and 75mg/kg bw) of [6]-gingerol was found to be associated

with increased plasma insulin concentration in iAs intoxicated

mice.

iAs has been shown to potentially alter some gene expressions

related to glucose homeostasis in body. Uptakeof glucose molecule

inside the cell depends on the translocation of glucose transporter

4 (GLUT4), which plays a very important role in insulin signaling,

by the action of insulin molecule to the plasma membrane. Hence,

we targeted the intracellular GLUT4 content in isolated hepato-cytes (control cells:62.2%) and found that 10M of iAsintoxicationlowers the level of GLUT4 content (31.3%), which was increased

(up to 38.9% and 43%) after incubation with [6]-gingerol. We also

assessed the mRNA and protein level expressions of GLUT4 in iAs

intoxicated mice, treated or un-treated with [6]-gingerol. Auto-

phosphorylation of insulin receptor substrates and activation of

PI3K/AKT signaling pathwayare required for the activation of down

steam signal cascade leading to glucose uptake and metabolism

in cell (Vijayakumar et al., 2005). Therefore we investigated the

expressions of IRS1, IRS2, p85 of PI3K and AKT in liver tissue of

experimental mice.Our results revealed that[6]-gingerol enhanced

the activity and expressions of these genes which were initially

down regulated in iAs intoxicated hyperglycemic mice. TNF- andIL6 levels are closely associated with both hyper-insulinemia and

insulin resistance(Rondinone,2006). Increasedlevels of TNF- andIL6 are also associated with iAs exposure. Therefore, we examined

the changes of expressions of these two genes in iAs intoxicated

mice, and after treatment with [6]-gingerol, reduced expression

levels of thesaid genes were found. This would revealthat thedrug

exerted its effect in combating impaired insulin responsiveness

through regulation of these genes. iAs intoxication can also down

regulate thetranscription factor PPAR whichisverycloselyrelatedto the activity of TNF- and IL6 (Gurnell, 2003). Our investigation

Fig. 9. Relative fluorescence intensities of PCR bands. The relative intensities of bands were determined using ‘Image J’ software. The results shown in histograms are the

average ±SD (N = 6). Significance * p< 0.05 iAs-intoxicated vs. normal control group; significance # p< 0.05 drug-fed vs. iAs-intoxicated group.


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2. Toxicology Letters