Transcript Lead and Arsenic Toxicity

Thymoquinone attenuates cisplatin induced toxicity
and oxidative damage in rat kidney
By: Zeba Farooqui
Department of Biochemistry
Aligarh Muslim University, India
Supervisor
Dr. Farah Khan
Overview
 Cisplatin (cis-diamminedichloroplatinum II, CP) is one of the most effective platinum-based
chemotherapeutic agent with a broad range of antitumor activities.
 Therapeutic utility of CP is limited due to its associated side effects including acute and
chronic nephrotoxicity, hepatotoxicity and ototoxicity.
 Multifactorial mechanisms might be involved in CP induced nephrotoxicity. However, one of
the well known mechanism involved is CP induced reactive oxygen species (ROS) generation.
 Considering the effectiveness of CP, it is necessary to prevent dose limiting side effects that
inhibit its use at tumoricidal doses.
 Since oxidative stress has been implicated in the etiology of CP induced toxicity, renewed
interest has been centred on the role of natural antioxidants having free radical scavenging
and/or antioxidant properties to counteract CP toxicity.
 Thymoquinone, a phytochemical isolated from Nigella sativa seeds, has been tested for its
therapeutic effects in many diseases.
INTRODUCTION
Cisplatin
 Inorganic
complex of platinum, synthesized by Michael
Peyrone in 1845.
 Effectiveness
as anticancer drug was discovered by
Rosenberg and co workers in 1969.
 Approved
by Food and Drug Administration (FDA) in 1978.
 Used
for the treatment of various human solid tumors including those of head, neck,
ovary, testes and breast.
 Health
risks: nephropathy, liver damage, hearing loss, myelosuppression, nausea and
vomiting.
Pathophysiology of cisplatin in kidney
 Adverse morphological changes in the S3 subsegment of renal proximal tubule
 Loss of brush border membrane
 Alters membrane permeability
 Depletes intracellular glutathione and interact with enzyme/protein sulphydryl
groups
 Perturbs antioxidant defense system
 Tubular necrosis
Protection and/or prevention of cisplatin
induced toxicity
Strategies to
ameliorate cisplatin induced
toxicity
Antioxidants: Flaxseed oil, Fish oil, Nigella
sativa oil, melatonin, selenium, curcumin,
silymarin, rutin
Thymoquinone

Thymoquinone (2-Isopropyl-5-methyl-1,4-benzoquinone,
TQ) is a monoterpene isolated from essential oil of Nigella
sativa (NS) seeds.

TQ exhibit promising therapeutic potential against many
diseases such as diabetes, artherosclerosis and cancer.

TQ possesses an antioxidant action to scavenge free radicals and also up regulate the
expression and/or activities of antioxidant enzymes.

The strong antioxidant properties of TQ is related to the redox properties of its quinone
structure and its ability to cross the morpho-physiological barriers and hence easy
access to sub-cellular compartments that facilitates its free radical scavenging activity.
HYPOTHESIS
Anti- tumor
Cisplatin
Nephrotoxicity
Thymoquinone
Thymoquinone would be able to prevent/reduce cisplatin
induced adverse effects on kidney
Experimental design
C : control; CP: cisplatin treated; TQ: Thymoquinone administered; CPTQ: thymoquinone+cisplatin
treated; i.p : intraperitoneal injection ; cisplatin injection ( ) ; normal saline injection ( )
Experiments were conducted to study the effect of CP alone and
in combination with TQ on
o
various biochemical parameters in serum
enzymes of brush border membrane (BBM) in cortical and
medullary homogenates and in cortical BBM vesicles (BBMV)
o
ovarious
oxidative stress parameters in cortical and medullary
homogenates
o
histopathological examination of rat kidney
RESULTS
Table 1. Effect of TQ administration with and without CP treatment on serum
parameters
Groups
Creatinine
(mg/dl)
0.93 ± 0.03
BUN
(mg/dl)
12.83 ± 1.265
Cholesterol
(mg/dl)
54.39 ± 2.64
Phospholipid
(mg/dl)
116.20 ± 5.72
Phosphate
(μmols/ml)
2.50 ± 0.10
Glucose
(mg/dl)
100.42 ± 0.45
CP
1.85 ± 0.29*
(+98.92%)
24.21 ± 1.773*
(+88.69%)
70.06 ± 3.61*
(+28.81%)
211.98 ± 6.28*
(+82.42%)
1.31 ± 0.18*
(-47.6%)
74.29 ± 1.10*
(-26.02%)
TQ
0.99 ± 0.064
(+6.45%)
13.11 ± 1.65
(+2.18%)
49.92 ± 3.15
(-2.74%)
125.9 ± 6.71
(+8.34%)
2.22 ± 0.21
(-11.2%)
117.26 ± 6.69
(+16.76%)
CPTQ
1.13 ± 0.05†
(+21.50%)
15.24 ± 1.20†
(+18.78%)
62.16 ± 2.99
(+14.28%)
173.65 ± 7.73*†
(+49.44%)
1.91 ± 0.38
(-23.6%)
92.48 ± 7.43†
(-7.91%)
Control
CP: cisplatin treated; TQ: Thymoquinone administered; CPTQ: Thymoquinone administered + cisplatin treated; BUN: blood urea nitrogen.
Results are mean ± SEM for three different preparations.
*Significantly different from control.
† Significantly different from CP at p<0.05 by one way ANOVA.
Values in parenthesis represent percent change from control
A
1
†
Specific activity
(µmoles/mg protein/hr)
Specific activity
(µmoles/mg protein/hr)
10
B
0.8
8
*
†
0.6
6
*
0.4
4
2
0.2
0
Control
CP
TQ
Specific activity
(µmoles/mg protein/hr)
70
CPTQ
0
Control
CP
TQ
CPTQ
C
60
50
*
40
30
20
10
0
Control
CP
TQ
CPTQ
Figure 1. Effect of TQ administration with and without CP treatment on ALP activity in (A) cortical
homogenate (B) medullary homogenate and (C) cortical BBMV
A
60
LAP
LAP
Specific activity
(µmoles/mg protein/hr)
Specific activity
(µmoles/mg protein/hr)
40
GGTase
B
GGTase
35
30
†
25
*
20
15
†
10
50
†
40
*
30
†
20
*
10
*
5
0
0
Control
CP
TQ
Specific activity
(µmoles/mg protein/hr)
250
Control
CPTQ
CP
C
TQ
CPTQ
GGTase
LAP
200
†
150
*
100
†
*
50
0
Control
CP
TQ
CPTQ
Figure 2. Effect of TQ administration with and without CP treatment on GGTase and LAP activities in
(A) cortical homogenate (B) medullary homogenate and (C) cortical BBMV
CP
A
Percent change
TQ
100
80
60
40
20
0
-20
-40
-60
-80
CPTQ
LPO
Total-SH
GSH
CP
80
B
TQ
CPTQ
Percent change
60
40
20
0
-20
-40
LPO
Total-SH
GSH
-60
-80
Figure 3. Effect of TQ administration with and without CP treatment on non enzymatic antioxidant
parameters in the homogenates of (A) cortex and (B) medulla
A
SOD
CAT
GSH-Px
TR
GR
GST
CP
TQ
20
CPTQ
Percent change
0
-20
-40
-60
-80
B
40
CP
SOD
CAT
GSH-Px
TR
GR
GST
TQ
CPTQ
Percent change
20
0
-20
-40
-60
-80
Figure 4. Effect of TQ administration with and without CP treatment on enzymatic antioxidant
parameters in the homogenates of (A) cortex and (B) medulla
Histopathology of rat kidney
A. Control group with normal corpuscle and tubular epithelium; B. CP treated group, reveals extensive
damage of both components. Glomerulus appear congested and tubules appear edematous with obvious
interstitial bleeding (all along the lower field); C. TQ alone group, shows histoarchitecture of both renal
corpuscle and renal tubules very similar to control group ; D. CPTQ group, shows reasonably good
preservation of renal corpuscle and tubules. Though, the glomerulus appears mildly congested, tubules are
not edematous and there is no obvious interstitial bleeding.
Figure 5. Histopathology of rat kidney showing glomerular capillary tuft ( ) in the renal corpuscle and
renal tubule ( ).
CONCLUSION
Cisplatin treatment
• Damaged proximal tubular
membrane
• Caused oxidative stress
- Increased lipid peroxidation
- Decreased antioxidant
enzyme activities
 CP elicited deleterious nephrotoxic and
other adverse effects as indicated by
significant increase in serum creatinine and
blood urea nitrogen (BUN), decrease in the
activities of various BBM enzymes and
suppression of antioxidant defense system.
 TQ administration markedly ameliorated
CP induced nephrotoxicity.
have ameliorated the damage
Renal tissue TQ might
caused by CP in the following ways:
• TQ administration increased the levels of
antioxidant enzymes in the renal tissues
Cisplatin + Thymoquinone
resulting in enhanced antioxidant defense
co treatment
against CP generated ROS.
• Owing to its antioxidant/free radical
scavenging property, TQ might have
• Reduced proximal tubular
reduced the CP induced generation of free
damage
radicals and ROS.
• Diminished oxidative stress
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