Professional Documents
Culture Documents
Kidney Injury 1 (Anjeer)
Kidney Injury 1 (Anjeer)
Kidney Injury 1 (Anjeer)
A R T I C LE I N FO A B S T R A C T
Keywords: Background: Cisplatin (CisPt) is one of the most widely used and highly effective drugs for the treatment of
Ficus carica L various solid tumors, unfortunately acute kidney injury (AKI) is considered one of its side effects through several
Gold nanoparticles mechanisms including production of reactive oxygen species (ROS), pro-inflammatory and pro-fibrotic cyto-
Cisplatin kines. Due to the poor effect of AKI therapy, the use of nanoparticles loaded with natural extracts for delivering
Homocysteine
to the kidney molecules are desirable.
Hydroxyproline
Aim: This study aims to investigate the effectiveness of different concentrations of gold nanoparticles (Au-NPs)
as a carrier for Ficus carica L. (Fig) leaves extract against CisPt induced AKI.
Methods: Seventy male albino rats were used and divided into seven groups. After the experimental period,
blood was withdrawn, serum was separated for determination of urea, creatinine, homocystein (Hcy) and folic
acid while reduced glutathione (GSH), nitric oxide (NO), malondialdehyde (MDA), total antioxidant capacity
(TAC) and hydroxyproline content (Hyp) were evaluated in kidney tissue homogenate.
Results: CisPt induced AKI in rats and results in a significant increase in the levels of serum urea, creatinine, Hcy
and kidney Hyp, lipid peroxidation along with a significant reduction of kidney GSH, NO and TAC compared to
the control rats. Treatment with Au-NPs and Fig extract particularly in a ratio of (3:2) respectively was shown to
improve renal functions with efficient capacity in scavenging ROS and reduced AKI severity.
Conclusion: Au-NPs enhanced the anti-oxidative properties of the Fig extract in targeting kidney damaged tissue
and reduced oxidative toxicity induced by CisPt.
1. Introduction clinic have increased drug solubility, reduced off-target side effects, and
provides novel diagnostic tools; there is an increasing cohort of nano-
Cis-diamminedichloroplatinum II (CisPt) is a well-documented drug materials which may have implications for kidney disease [4]. Of these
in the treatment of several malignancy types; while a limitation of its nanomedicines is gold nanoparticles (Au-NPs) which have attracted
use is well observed because CisPt -induced nephrotoxicity (CIN) in intensive interest owing to some features such as being easily prepared,
more than 50% of cases [1]. Several mechanisms were involved in high surface area (more active), low toxicity and can be readily at-
nephrotoxicity including endothelin-1, upregulation of transforming tached to molecules of biological interest.
growth factor-β, attenuation of oxidative stress, also, necrosis and Ficus carica L. (Fig) is a common tree in the Middle East and the
apoptosis in addition to the elevation of macrophage and monocyte Mediterranean region, belonging to the botanical family. Fig is usually
infiltration into the renal cortex [2]. Antioxidants have become a topic consumed its dry or fresh grade [5]. It is an excellent source of minerals
of interest to health and food science researchers and medical experts. and vitamins; it is fat and cholesterol-free and contain a high number of
Food that has antioxidants properties have an effective role in pro- amino acids [6] and a large number of phenolic compounds [7].
tecting against different diseases concerning oxidative stress such as Therefore, the present work was designed to investigate the effec-
liver and kidney diseases in a long duration as chemo preventive agents tiveness of both Au-NPs and Fig leaves extract against CisPt induced
[3]. Nanotechnology can confer significant benefit to medicine, such as AKI. Furthermore, the synergistic behavior of augmenting both agents
the targeted delivery of drugs to specific tissues. Nanomedicines in the was thoroughly studied. Different concentrations of Au-NPs loaded with
⁎
Corresponding author.
E-mail addresses: Mehrez_chem@yahoo.com, mehrezeelnaggar@gmail.com (M.E. El-Naggar).
https://doi.org/10.1016/j.colsurfb.2019.110465
Received 7 March 2019; Received in revised form 22 August 2019; Accepted 27 August 2019
Available online 12 September 2019
0927-7765/ © 2019 Elsevier B.V. All rights reserved.
S.M. El-Sayed, et al. Colloids and Surfaces B: Biointerfaces 184 (2019) 110465
Fig leaves extract were examined in ameliorating CisPt induced ne- ions to Au0. For this purpose, the reduction reaction between tetra-
phrotoxicity in albino rats. chloroauric acid (HAuCl4) and tri-sodium citrate (Na3C6H5O7.2H2O)
was used in an aqueous solution. The second part is the stabilization
2. Materials and methods effect using cationic surfactant; cetyltrimethyl ammonium bromide
(CTAB) to avoid the aggregation of the formed nanoparticles. Finally,
2.1. Materials the prepared Au-NPs with narrow size distribution were obtained.
Typically, 500 mg of tri-sodium citrate was dissolved in 50 ml of deio-
2.1.1. Chemicals nized water, kept under magnetic stirring while the temperature was
Tetrachloroauric Acid (HAuCL4) and Hcy standard {for high per- raised to 70 °C. After complete dissolution, 500 mg of CTAB was added
formance liquid chromatography (HPLC)} were obtained from Sigma- under stirring. The pH of the reaction medium was adjusted to 11 by
Aldrich Company USA. Tri-sodium Citrate was purchased from Fischer using NaOH solution (0.01 M). At the end of mixing, 250 mg of HAuCl4/
Co. Germany. Cisplatin was purchased from Mylan, Saint Priest, France. 5 ml of deionized water was added dropwise to the previous solution
Chloramine T was purchased from Fisher, Fair Lawn, NJ, USA. P-di- and the whole reaction was kept for another 30 min under stirring. At
methylaminobenzaldehyde, and Hyp were obtained from Sigma the end of the reaction, the color of the solution turned to red indicating
Chemical Company (U.S.A). Sodium acetate, sodium hydroxide, acetic the successful preparation of Au-NPs. The solution was kept away from
acid, citric acid, n-propanol and ethanol were of high grade and pur- light for further characterization and application. Techniques such as
chased from Fisher Scientific (USA). All other chemicals were used as UV–vis spectroscopy, TEM and XRD were used to characterize and
received and Deionized water was used for experiments, characteriza- determine the size and size distribution of the synthesized Au-NPs.
tion and application.
2.2.5. Preparation of Fig leaves extract loaded Au-NPs
2.1.2. Plant Different concentrations from both Au-NPs and Fig extract were
Fresh Fig leaves were collected in May 2018 from a farm in prepared as follows: Au-NPs and Fig extract in a ratio of (2:3) v/v, Au-
Menofyia governorate, Egypt. The plant was identified on the NPs and Fig extract in a ratio of (2:2) v/v and Au NPs and Fig extract in
Herbarium of the National Research Centre (NRC) of Egypt. The leaves a ratio of (3:2) v/v. Rats received different concentrations of Au-NPs/
were washed and shade dried at room temperature for one week and Fig extract mixture (0.5 ml/Kg rat) by oral administration daily for 14
then cut into small pieces. Then ground into powder with special days.
electric mill and stored in polyethylene bags until extraction.
2.2.6. Experimental design
2.1.3. Experimental animals Seventy rats were included in this study and equally divided into
Seventy male albino rats, weighing 180 ± 20 g from Animal House, seven groups as follow: Control group: animals received a vehicle, AKI
National Research Centre (NRC) Giza, Egypt was kept in clean cages of group: Animals received a single dose of Cispt and received a vehicle
polypropylene and maintained in controlled room temperature with daily, treated group I: Rats received a single dose of CisPt then received
light and dark cycle, given a standard diet and water ad libitum along Fig leaves extract (400 mg extract /Kg b.w. /day) orally for 14 days.
the experimental period. The experiment was carried out in accordance Treated group II: Rats received a single dose of CisPt then received Au-
with guidelines and protocol approved by the Institutional Animal NPs (1 mg/kg b.w. in aqueous solution/day) orally for 14 days. Treated
Ethics Committee of National Research Centre (NRC), Giza, Egypt. group III: Rats received a single dose of CisPt then received 0.5 ml of Au
NPs/Fig leaves extract mixture as (2:3) v/v respectively orally for 14
2.2. Methods days, treated IV group: Rats received a single dose of CisPt then re-
ceived 0.5 ml of Au-NPs/Fig leaves extract mixture as (2:2) v/v re-
2.2.1. Cisplatin induction spectively orally for 14 days, treated group V: Rats received a single
Rats received a single dose of CisPt (20 mg/kg body weight) by dose of CisPt then received 0.5 ml of Au-NPs/Fig leaves extract mixture
intraperitoneal injection; the dose was modified from a previous study as (3:2) v/v respectively orally for 14 days.
[8]. After finalizing the experimental period, all animals were kept
fasting for12 h for blood sampling that was collected in clean tubes and
2.2.2. Preparation of tissue homogenate centrifuged at 3000 rpm for separation of sera and then stored at
One gram from kidney tissue was cut into tiny pieces and then −20 °C. After that, the biochemical analysis was carried for the sepa-
homogenized with phosphate buffer (pH was adjusted to 7.4). After rated sera while kidney tissues were removed quickly from each rat,
that, the homogenate was centrifuged using cooling centrifuge washed with ice-cold saline and kept at -80 °C until used for determi-
(Laborzentrifugen, 2K15, Sigma, Germany) at 5000 rpm for 10 min at nation of biochemical parameters.
4 °C; clear supernatant was then removed and used for estimation of
different parameters [9]. 2.3. Characterization
2
S.M. El-Sayed, et al. Colloids and Surfaces B: Biointerfaces 184 (2019) 110465
Additionally, the antioxidant activity of the phenol extracts was drops from phosphoric acid ; then filtered twice through a membrane
evaluated according to the method described by Bandoneon et al., [11]. filter (Whatman, 47 mm Dia.,0.45-μm). The M.P was delivered through
The method was carried by mixing 3.9 mL methanolic solution of 2, 2- the column at a flow rate of 1 ml/min a controlled temperature (40 °C)
diphenyl-1-picryl-hydrazyl radical (DPPH) (0.0025 g/100 mL CH3OH) using UV detector that was set at 260 nm. The peak areas of samples
and 0.1 mL of methanolic solutions of phenol extracts in a cuvette and were determined and the concentration of each sample was calculated
the absorbance at 515 nm was measured against methanol using a from the standard curve.
double-beam Uv–vis spectroscopy. On the other hand, the absorbance
at this definite wavelength was measured for the blank sample which 2.3.3.6. Determination of serum folic acid. Serum folic acid was
consists of a mixture containing 0.1 mL methanol and 3.9 mL metha- measured using ELISA Kit (NOVA, Bioneovan Co., Ltd) according to
nolic solution of DPPH). The inhibition percentage of DPPH for the the manufacturer’s instructions.
phenol extract and blank samples were calculated as follow:
the mobile phase (M.P) consisted of equal volumes of 1- sodium Fig (Ficus carica L.) 5.13 ± 0.18 47.34 ± 1.15
phosphate (40 mmol/L). 2- heptanesulfonic acid (8 mmol/L) 3-
methanol (18%) v/v; M.P was adjusted to pH 3.1 by addition of few Mean ± standard deviation, D.W: Dry Weight.
3
S.M. El-Sayed, et al. Colloids and Surfaces B: Biointerfaces 184 (2019) 110465
Scheme 1. Formation of Fig leaves extract, Au-NPs and Fig leaves extract blended with Au-NPs.
exhibited for the identified surface plasmon resonance of Au-NPs. This that the particle shape turned to spherical shape with a marginal in-
identified surface plasmon resonance of Au-NPs results in a strong ab- crease in the particle size due to the coating effect of Fig leaves extract
sorbance band in the visible region; 500 nm–560 nm. As displayed from on the surface of Au-NPs but still in the nanoform state. Thus, TEM was
Fig. 1A that the formed Au-NPs colloidal solution exhibits a peak at effectively used to analyze the shape and diameter of the as synthesized
520 nm confirming the successful conversion of Au ions to Au-NPs. The Au-NPs with and without mixing the Fig leaves extract.
full conversion of ions to nanoparticles is confirmed by the addition of Moving to the second part of the current work to evaluate the hy-
NaCl to the formed colloidal solution of Au-NPs. There is no white brids of Fig leaves extract blended with different concentrations of Au-
precipitate is formed upon the addition of NaCl to Au-NPs solution that NPs against cisplatin induced acute kidney injury. CisPt, the anti-neo-
confirms that there is no noticeable existence of ions at the end of the plastic drug, is used in the treatment of several types of malignancies
reaction. include head, bladder, breast, ovarian and lung cancer. In spite of its
TEM was characteristically used to establish the physical size and role in improvement of patient’s life expectancy [25]. It is considered as
morphological structure of Au-NPs. Fig. 1B shows a TEM image of Au- a strong toxin and the nephrotoxicity is one of its important compli-
NPs reduced via tri-sodium citrate as reductant and CTAB as a capping cations in clinical and also in experimental models that can be pro-
agent. As clearly seen from the onset image of TEM (Fig. 1B), the gressive in a lot of cases [26]. Thus, acute renal toxicity remains the
particles are formed with a very small diameter size. The high-resolu- main problem of CisPt' use [25].
tion image (Fig. 1B) is taken for better understanding the shape of the In the present study, CisPt injection significantly increased blood
particles which reveal that the hexagonal shapes of Au-NPs are formed. urea and serum creatinine in CisPt group in compared with control
For further characterization, XRD was used to study the structural animals (Table 2). Elevation of kidney functions in this work is related
properties of the as-prepared Au-NPs. Fig. 1C shows XRD of Au-NPs in to the deleterious side effects of CisPt that generates ROS as was ob-
its powder form after precipitation of the colloidal solution using cen- served in Table 3, thus it significantly increased lipid peroxidation and
trifuge instrument with 15,000 rpm for 2 h. As presented from XRD, the significantly decreased GSH, NO and TAC in CisPt group compared to
formed Au-NPs exhibit characteristic peaks at 38°, 43.8° and 65° as- control.
cribed to the crystallographic planes (1 1 1), (2 0 0) and (2 2 0), cor- Liberation of reactive oxygen species (ROS) and renal micro-
respondingly. The peak width of Au-NPs from crystalline plane (1 1 1), vasculature vasoconstriction are well dependable on CIN. Moreover,
sizes of the Au0 crystallite were found to be approximately 20 nm. cisplatin selectively injured S3 segment of proximal tubules that is
On the other hand, after mixing Fig leaves extract with Au-NPs in found in the outer stripe of the medulla that is found outside, as evi-
equal amounts and subjected to 30 min of sonication, it is essential to denced by necrosis & apoptosis. It was reported that generation of NOS,
assess the particle shape of the formed hybrids. Fig. 1D displays TE the reactive nitrogen species, have been also involved in CIN; CisPt
image of Fig leaves extract blended with Au-NPs. It is evidently seen exalts the liberation of NO and peroxynitrite in renal tissues; the later
4
S.M. El-Sayed, et al. Colloids and Surfaces B: Biointerfaces 184 (2019) 110465
Fig. 1. (A) Uv–vis, (B) TEM, (C) XRD of Au-NPs and (D) TEM of Au-NPs loaded with Fig leaves extract.
Table 2 tubular cells necrosis that leads to the reduction of glomerular filtration
Blood urea and serum creatinine levels in different studied groups. rate (GFR) and kidney dysfunction [2].
Parameters Groups Urea (mg/dl) Creatinine (mg/dl) Elevation of oxidative stress parameters in this study may be related
to accumulation of CisPt in the kidney by peritubular uptake. Whereas,
Control 25 ± 5.7 0.83 ± 0.14 such this concentration of the drug in the renal cortex is greater than
Cisplatin 113a ± 6.6 2.2a ± 0.20
other organs.
Treated I 70a,b ± 5.0 1.4a,b ± 0.14
Treated II 38.3b ± 7.2 0.7b ± 0.05
The pathological mechanisms of renal toxicity involve elevation of
Treated III 40b,c ± 12.5 0.96b,c ± 0.18 endothelin-1 and transforming growth factor β, oxidative stress, ne-
Treated VI 40b,c ± 2.8 0.66b,c ± 0.21 crosis, apoptosis and increase in macrophage/monocyte infiltration into
Treated V 33.3b,c ± 4.4 0.40b,c ± 0.05 the renal cortex and medulla [2].
Renal dysfunction e.g. chronic kidney disease (CKD) preexisting
P: a significant difference compared to Pa) the control group.
when CisPt chemotherapy is started may induce tubulo-interstitial le-
Pb) CisPt group, Pc) treated I group and Pd)treated II group.
sions, which may further evolve towards interstitial inflammation and
fibrosis. In this study, hydroxyproline content in the kidney was sig-
effectively stimulates the changes of lipid functions and structure, DNA,
nificantly increased by CisPt injection compared with control subjects
proteins and also the reduction of cellular defenses system by oxidation
(Table 4) that appeared the role of CisPt in inducing AKI. In the same
of thiol pools [1].
line, El-Khayat et al., [19] indicated the accumulation of hydroxypro-
In addition, renal toxicity involves kidney free radical production
line content in cirrhotic rat's liver.
and increasing antioxidant defense mechanisms, and also acute renal
Additionally, this study showed a significant increase in serum Hcy
Table 3
Oxidant and antioxidant levels in different studied groups.
Parameters Groups GSH (mg/g tissue) MDA (nmol/g tissue) NO (U/g tissue) TAC (mM/g tissue)
P: a significant difference compared to Pa) the control group, Pb) CisPt group, Pc) treated I group and Pd) treated II group.
5
S.M. El-Sayed, et al. Colloids and Surfaces B: Biointerfaces 184 (2019) 110465
6
S.M. El-Sayed, et al. Colloids and Surfaces B: Biointerfaces 184 (2019) 110465