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Saudi Journal of Kidney Diseases and Transplantation
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Table of Contents   
ORIGINAL ARTICLE  
Year : 2021  |  Volume : 32  |  Issue : 5  |  Page : 1243-1252
Kidney dysfunction and oxidative stress in doxorubicin-induced nephrotic rat: Protective role of sesame oil


1 Department of Physiology, School of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
2 Department of Physiology, School of Medicine; Applied Biomedical Research Center, Mashhad University of Medical Sciences, Mashhad, Iran
3 Department of Physiology, School of Medicine, Iranshahr University of Medical Sciences and Health Services, Iranshahr, Iran

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Date of Web Publication4-May-2022
 

   Abstract 


Doxorubicin (DOX) is an antineoplastic agent which it’s clinical use has been limited due to its major side effects including cardiotoxicity and nephrotic syndrome. Sesame oil (SO) is an important edible oil with many pharmacologic effects. The aim of the present study was to investigate the effect of SO against DOX-induced nephropathy in the rat. In this study, two doses of SO (3 and 6 mL/kg) were administrated orally for six consecutive weeks and DOX (mg/kg) was intravenously injected on the 4th day of the experiment. Blood and urine samples were collected on days 1, 14, 30, and 42 for subsequent measurement of biochemical parameters. The left kidneys were removed for subsequent assessment of total thiol content, malondialdehyde (MDA) concentration, and renal activities of catalase and superoxide dismutase enzymes. DOX caused significant proteinuria, hypoalbuminemia, and hyperlipidemia compared to control group. Significant decrease in antioxidant enzyme activities and total thiol contents and significant increase in MDA levels were also observed following DOX injection when compared to control group. Oral administration of SO significantly reversed DOX-induced proteinuria, hypoalbuminemia, and hyperlipidemia compared to DOX group. Furthermore, compared to DOX group, SO significantly increased total thiols content. MDA concentration significantly decreased following SO administration when compared to DOX group. The current study suggests that SO is able to improve kidney function as well as kidney tissue oxidative damage in DOX-induced nephrotic the rat.

How to cite this article:
Mahzari S, Hosseinian S, Hadjzadeh MA, Mohebbati R, Noshahr ZS, Rad AK. Kidney dysfunction and oxidative stress in doxorubicin-induced nephrotic rat: Protective role of sesame oil. Saudi J Kidney Dis Transpl 2021;32:1243-52

How to cite this URL:
Mahzari S, Hosseinian S, Hadjzadeh MA, Mohebbati R, Noshahr ZS, Rad AK. Kidney dysfunction and oxidative stress in doxorubicin-induced nephrotic rat: Protective role of sesame oil. Saudi J Kidney Dis Transpl [serial online] 2021 [cited 2022 May 25];32:1243-52. Available from: https://www.sjkdt.org/text.asp?2021/32/5/1243/344743



   Introduction Top


Doxorubicin (DOX) is a chemotherapy drug with a broad-spectrum antineoplastic activity. DOX is used for the treatment of a variety of human malignancies including Kaposi’s sarcoma, breast cancer, lymphoma, bladder cancer, and acute lymphocytic leukemia.[1] In spite of potent antitumor efficacy, the clinical use of DOX has been limited due to its major side effects including cardiotoxicity and nephrotic syndrome (NS), characterized by severe proteinuria, hypoalbuminemia, hyperlipidemia, and edema.[2] DOX-induced nephropathy is a widely used experimental model employed for the evaluation of pathophysiologic mechanisms involved in proteinuric nephropathy.[3] Although it was accepted that loss of nephrin due to direct toxic damages caused by DOX to the glomerular structure, is the main cause of proteinuria following DOX administration, the exact mechanism of DOX-induced nephropathy is poorly understood. It was reported that oxidative stress, caused by overproduction of reactive oxygen species, impairs anti-oxidant defense systems and increases lipid peroxidation all leading to DOX-induced renal toxicity. Besides oxidative stress, inflammation plays a central role in DOX-induced nephropathy.[4] It is hypothesized that natural antioxidant and anti-inflammatory agents can effectively improve renal complications produced by DOX.

Sesamum indicum L. belonging to the Pedaliaceae family, is one of the most ancient plants cultivated by humans, and first considered a crop over 4000 years ago in Assyria and Babylon.[5] Sesame oil (SO), extracted from S. indicum seeds, is an important edible oil used as a food ingredient since 2000 years ago.[6] SO contains unsaturated fatty acids, a small amount of free fatty acids and γ-tocopherol. SO also contains considerable amounts of lignans including sesamin, sesamolin, sesamol, sesaminol, pinoresinol and sesamolinol. Lignans and γ-tocopherol provide SO with a good stability against autoxidation.[5] Many pharmacologic properties have been reported for SO including anti-oxidant,[5] anti-inflammatory,[7] antidiabetic[8] and antihypertensive[9] properties. Because of potent antioxidant activity of SO, daily supplementation with SO could protect the kidneys against acute and chronic kidney diseases (CKD) in different animal models.[10],[11] Thus, the aim of the present study was to investigate the effect of daily SO administration against DOX-induced renal injury in rats.


   Method and Materials Top


Chemicals and reagents

DOX was purchased from the Ebewe Pharma Company (Austria). 5, 5′-dithiobis-2-nitrobenzoic acid, 2-thiobarbituric acid, trichloroacetic acid, Tris, HCl, and potassium chloride were obtained from Merck Company (Germany). Biochemical parameters in blood were measured by appropriate diagnostic kits (Pars Azmoon, Tehran, Iran).

Animals

Forty male Wistar albino rats weighing 220 ± 30 g, were obtained from animal house of Mashhad Medical School, Mashhad University of Medical Sciences, Mashhad, Iran. The animals were housed under 12 h/12 h light-dark cycles at 20°C–24°C with free access to food and water ad libitum. All the experiments were approved by Ethics Committee of Mashhad University of Medical Sciences.

Experimental protocol

The animals were randomly divided into the five following groups (8 rats in each): In Group I (control), saline was daily given orally (p.o.) for six consecutive weeks and saline was injected [intravenously (i.v.)] on the 4th day of the experiment. In Group II (DOX), saline was daily given orally (p.o.) for six consecutive weeks and DOX was injected (5 mg/kg, i.v.)[2] on the 4th day of the experiment. In Group III (SO), SO was daily given orally (6 mL/kg, p.o.) for 6 consecutive weeks and saline was injected (i.v.) on the 4th day of the experiment. In groups IV (SO3+DOX) and V (SO6+DOX), SO was daily given orally (3 mL/kg and 6 mL/kg, respectively, p.o.) for six consecutive weeks and DOX was injected (5 mg/kg, i.v.) on the 4th day of the experiment.

Blood samples were collected in heparinized tubes from the orbital sinus under anesthesia induced by ether, on days 1, 14, 30, and 42. Twenty-four-hour urine samples from the rats were taken on the same days while each animal was separately housed in a metabolic cage. Serum was separated by centrifugation at 4000 g for 10 min and stored at -20°C until assayed. At the end of the study (day 42), all animals were anesthetized by an intra-peritoneal injection of ketamine (60 mg/kg) and xylazine (6 mg/kg), and then, both kidneys were quickly removed. The left kidneys were stored at -70°C for subsequent measurement of total thiol content, malondialdehyde (MDA) concentration, and renal activities of catalase and superoxide dismutase (SOD) enzymes. Then, all the animals were humanely killed.

Assessment of biochemical parameters

Serum levels of albumin, triglyceride (TG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) were measured colorimetrically by using commercial diagnostic kits (Pars Azmoon Company, Tehran, Iran). Urine protein concentration was measured using TCA method and used for the calculation of urinary excretion rate of protein based on the following formula:

Urinary excretion rate of protein (mg/day) = urine concentration of protein (mg/mL) × urine output (mL/day)

Assessment of oxidative stress

The peroxidation of lipids, a marker of oxidative stress, was evaluated by measuring MDA in renal tissue homogenates by the method explained by Mihara and Uchiyama.[12] Following formula was used for measurement of MDA concentration (C). C were expressed as nanomole MDA per gram tissue (nmol MDA/g kidney tissue):

C (Molar) = A/1.56 × 105

Total thiol content (thiol concentration - mM) of the renal tissue was estimated using the method explained by Sedlak and Lindsay[13] as per the following equation:

Total thiol concentration (mM) = (A2-A1-B) × 1.07/0.05 × 13.6

Renal catalase activity was determined in tissue homogenates using the calorimetric method described by Aebi based on the disappearance of hydrogen peroxide (H2O2).[14] SOD activity was measured using the calorimetric method described by Madesh and Balasubramanian.[15]


   Statistical Analyses Top


Statistical analyses were carried out using the IBM SPSS Statistics software version 20.0 (IBM Corp., Armonk, NY, USA). All data are expressed as means ± standard error of the mean. Between group comparison was made using the one-way ANOVA followed by LSD post hoc test. Intragroup comparisons were made using the repeated measures. Differences were considered statistically significant at P <0.05.


   Results Top


Effect of doxorubicin and sesame oil on protein excretion rate and serum albumin levels

Changes in urinary excretion rate of protein and serum level of albumin in different groups are summarized in [Table 1]. In the control and SO groups, protein excretion rate did not vary significantly among different days of the study. In the DOX group, there was a significant proteinuria on days 14, 30 and 42 of the study when compared to baseline (P <0.01 - P <0.001). In this group, urinary excretion rate of protein showed significant increases on days 30 and 42 when compared to day 14 (P <0.05 - P <0.001). Compared to day 30, in the DOX group, urinary excretion rate of protein significantly increased (P <0.01). In SO3 + DOX and SO6 + DOX groups, there were significant increases in protein excretion rate on days 30 and 42 when compared to days 1 and 14 (P <0.05 - P <0.001). Serum albumin level was also measured on days 1, 14, 30 and 42 of the experiment. Changes in serum albumin level within the control and SO-treated groups were not significant at different intervals. However, in DOX group, serum albumin showed a significant decrease on day 30 when compared to days 1 and 14 (P <0.05 - P<0.01). Furthermore, on the last day of the study (day 42), serum level of albumin significantly decreased when compared to days 1, 14, and 30 of the study (P <0.05 - P <0.01).
Table 1: The effect of sesame oil on sequential changes in protein excretion rate and serum albumin level in all experimental groups (n = 8).

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Effect of doxorubicin and sesame oil on serum lipid profile

Alterations in serum lipid profile are summarized in [Figure 1]. Administration of DOX caused significant increase in serum level of TC, TG, and LDL-C, and significant decrease in HDL-C, when compared to control group (P <0.001). At the end of the study (day 42), SO-treated groups had a significant lower levels of serum TC, TG, LDL-C, and significant higher level of HDL-C, compared to DOX group (P <0.001) [Figure 1]. Furthermore, the level of LDL-C in SO6 + DOX group was significantly higher than that of SO3 + DOX group (P <0.05) [Figure 1]c.
Figure 1: Serum lipid profile in all experimental groups of animal (n = 8). Values are the mean ± standard error of the mean. The data were analyzed using one-way ANOVA and post hoc LSD. A significant difference was considered at P <0.05.
***P <0.001 significant difference from control group, +++P <0.01 significant difference from DOX group, #P <0.01 significant difference from SO3+DOX group.
CO: Control, DOX: Doxorubicin, SO: Sesame oil.


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Effect of doxorubicin and sesame oil on renal MDA concentration

The changes in the renal levels of MDA are summarized in [Figure 2]. The level of MDA in DOX group significantly increased compared to the control group (P <0.001). Oral administration of two doses of SO to DOX-treated rats caused a significant decrease in renal MDA concentration compared to DOX group (P <0.01 for all).
Figure 2: Kidney tissue MDA concentration in all experimental groups of animal (n = 8). Values are the mean ± standard error of the mean. The data were analyzed using one-way ANOVA and y LSD. A significant difference was considered at P <0.05.
***P <0.001 significant difference from control group, ++P <0.01 significant difference from DOX group.
CO: control, DOX: Doxorubicin, SO: Sesame oil, MAD: Malondialdehyde.


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Effect of doxorubicin and sesame oil on antioxidant defense system

Alterations in renal total thiol content and activities of two antioxidant enzyme, SOD and catalase, are shown in [Figure 3], [Figure 4], [Figure 5]. Total thiol content in DOX group was significantly lowered compared to control group (P <0.05). Administration of SO (6 mL/kg) to DOX-treated rats significantly increased the levels of total thiols compared to DOX group (P <0.05) [Figure 3]. Renal activities of SOD and catalase significantly decreased in DOX-treated rats when compared to the control animals (P <0.05 for both) (Figures 4 and 5). Catalase and SOD activities showed 15% and 70% increase in SO6 + DOX group respectively, when compared to DOX group, but they were not statistically significant. Interestingly, in SO6 + DOX group, catalase activity significantly increased when compared to those of SO3 + DOX group (P <0.01) [Figure 4].
Figure 3: Kidney tissue total thiol content in all experimental groups of animal (n = 8). Values are the mean ± standard error of the mean. The data were analyzed using one-way ANOVA and post hoc LSD. A significant difference was considered at P <0.05.
*P <0.05 significant difference from control group, +P <0.05 significant difference from DOX group.
CO: Control, DOX: Doxorubicin, SO: Sesame oil.


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Figure 4: Kidney tissue catalase activity in all experimental groups of animal (n = 8). Values are the mean ± standard error of the mean. The data were analyzed using one-way ANOVA and post hoc LSD. A significant difference was considered at P <0.05.
*P <0.05, **P <0.01 significant difference from control group, ##P <0.01 significant difference from SO3+DOX group.
CO: Control, DOX: Doxorubicin, SO: Sesame oil.


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Figure 5: Kidney tissue SOD activity in all experimental groups of animal (n = 8). Values are the mean ± standard error of the mean. The data were analyzed using one-way ANOVA and post hoc LSD. A significant difference was considered at P <0.05.
*P <0.05 significant difference from control group, +++P <0.001 significant difference from DOX group.
CO: Control; DOX: Doxorubicin; SO: Sesame oil, SOD: Superoxide dismutase.


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   Discussion Top


In the present study, we showed that SO significantly improves proteinuria, changes in serum lipid profile and renal oxidative status in DOX-injected rats. DOX-induced NS in rodents, is a well-known experimental model which mimics proteinuria and pathological changes as observed in human minimal change disease and focal segmental glomerulosclerosis.[3] NS begins with changes in the glomerular filtration barrier, and consequent increased permeability to plasma proteins.[16] The results of the current study, in line with previous studies, showed that DOX administration causes a typical pattern of NS which was manifested by proteinuria as the major characteristic of NS beginning 10 days after a single intravenous injection. Our results also reflected occurrence of significant hypoalbuminemia and dyslipidemia. These findings confirmed accurate induction of NS in the present experiment. We observed that oral administration of two doses of SO (i.e. 3 and 6 mL/kg) could significantly reverse severe proteinuria and hypoalbuminemia, both caused by DOX injection. This finding was consistent with that reported by Liu et al showing attenuation of albuminuria in DOCA/salt-induced CKD in rats.[10] In the present study, DOX resulted in a significant lipid peroxidation manifested by increased levels of MDA. Also, antioxidant defense system was significantly impaired by DOX, as shown by decreased activity of SOD and catalase antioxidant enzymes and diminished total thiol content. Similar results were also observed in earlier studies.[17],[18],[19] It is believed that DOX-induced nephrotoxicity may be mediated via membrane lipid peroxidation, free radical formation, protein oxidation and iron-dependent oxidative damage of biological macromolecules.[20] In our study, elevated lipid peroxidation significantly decreased following administration of SO 3 and 6 mL/kg. However, total thiol content and SOD and catalase activities showed increase only following treatment with SO 6 mL/kg. This may indicate that antioxidant effect of SO is dose-dependent. Previous studies indicated that SO decreases levels of hydroxyl radical and peroxynitrite and prevents lipid peroxidation through elevating activities of antioxidant enzyme.[5],[21],[22] It was reported that the outstanding antioxidant activity of SO might be due to the presence of tocopherols and lignans (e.g. sesamol, sesamin, episesamin, sesaminol, and sesamolin). Furthermore, different studies found that the antioxidant property of SO is mainly because of synergistic activities of the above-noted active components instead of the activity of each one alone.[5] Different studies indicated that free radicals generation and subsequent inflammation, play critical roles in pathophysiology of DOX-induced kidney damage.[17],[23] Thus, the beneficial effects of SO against renal tissue damage induced by DOX, may be related to its antioxidant and anti-inflammatory properties. In the present study, SO administration was found to decrease serum level of TC, TG and LDL-C in DOX treated rats, suggesting that SO plays an important role in regulating lipid metabolism. This finding was consistent with that of Zhang et al showed that Sesamin, a lignin isolated from sesame seeds and present in SO, could ameliorate renal injury caused by lipid metabolism disorders in a rat model of hyperlipidemia.[24] Ramesh also showed that SO consumption beneficially influences lipid parameters in streptozotocin-induced diabetic rats.[25] Lipid peroxidation, as demonstrated with MDA augmentation, may exert a toxic effect on the glomerular basement membrane, and make proteinuria worse following DOX administration. As well as, filtered serum lipids may accumulate on glomeruli and cause transformation of glomerular mesangial cells to a proliferative phenotype. Hyperlipidemia, may also change phenotype of tubular epithelial cells which both could result in glomerular and tubular fibrosis. A recent study showed that Sesamin via hypolipidemic effect ameliorated mesangial cell proliferation, and reduced interstitial fibrosis in hyperlipidemic rats.[24] Hypolipidemic effect of SO might be due to the presence of monounsaturated fatty acids, polyunsaturated fatty acids and lignans (sesamin and sesamolin) in the oil.[25] Thus, it can be concluded that ameliorating effects of SO against nephrotic rats may possibly be via its antioxidant and hypolipidenic effects.

In summary, we investigated the effect of SO against DOX-induced kidney damage in rats. In this study, 42 day administration of SO markedly improved proteinuria, kidney tissue damage and renal oxidative stress in DOX-injected rats. It seems that potent antioxidant and anti-inflammatory effects of SO most possibly contribute to the therapeutic potential of SO in DOX-induced nephropathy. However, further investigations are necessary to clarify the exact mechanisms through which SO exerts its beneficial actions against kidney damage induced by DOX.


   Acknowledgment Top


This study was financially supported by Research Council of Mashhad University of Medical Sciences, Mashhad, Iran.

Conflict of interest: None declared.



 
   References Top

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Katzung BG, Masters SB, Trevor AJ. Basic and Clinical Pharmacology. 12th ed. New York: McGraw-Hill; 2012.  Back to cited text no. 1
    
2.
Naji Ebrahimi Yazd Z, Hosseinian S, Shafei MN, et al. Protection against doxorubicin-induced nephropathy by plantago major in rat. Iran J Kidney Dis 2018;12:99-106.  Back to cited text no. 2
    
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Okuda S, Oh Y, Tsuruda H, Onoyama K, Fujimi S, Fujishima M. Adriamycin-induced nephropathy as a model of chronic progressive glomerular disease. Kidney Int 1986;29:502-10.  Back to cited text no. 3
    
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Szalay CI, Erdélyi K, Kökény G, et al. Oxidative/nitrative stress and inflammation drive progression of doxorubicin-induced renal fibrosis in rats as revealed by comparing a normal and a fibrosis-resistant rat strain. PLoS One 2015;1:e0127090.  Back to cited text no. 4
    
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Wan Y, Li H, Fu G, Chen X, Chen F, Xie M. The relationship of antioxidant components and antioxidant activity of sesame seed oil. J Sci Food Agric 2015;95:2571-8.  Back to cited text no. 5
    
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Minar Mahmoud M. Hassanien MM, Abdel-Razek AG. Improving the stability of edible oils by blending with roasted sesame seed oil as a source of natural antioxidants. J Appl Sci Res 2012;8:4074-83.  Back to cited text no. 6
    
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Monteiro EM, Chibli LA, Yamamoto CH, et al. Antinociceptive and anti-inflammatory activities of the sesame oil and sesamin. Nutrients 2014;6:1931-44.  Back to cited text no. 7
    
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Sankar D, Ali A, Sambandam G, Rao R. Sesame oil exhibits synergistic effect with anti-diabetic medication in patients with type 2 diabetes mellitus. Clin Nutr 2011;30:351-8.  Back to cited text no. 8
    
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Karatzi K, Stamatelopoulos K, Lykka M, et al. Acute and long-term hemodynamic effects of sesame oil consumption in hypertensive men. J Clin Hypertens (Greenwich) 2012;14:630-6.  Back to cited text no. 9
    
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Liu CT, Chien SP, Hsu DZ, Periasamy S, Liu MY. Curative effect of sesame oil in a rat model of chronic kidney disease. Nephrology (Carlton) 2015;20:922-30.  Back to cited text no. 10
    
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Hsu DZ, Liu CT, Li YH, Chu PY, Liu MY. Protective effect of daily sesame oil supplement on gentamicin-induced renal injury in rats. Shock 2010;33:88-92.  Back to cited text no. 11
    
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Mihara M, Uchiyama M. Determination of malonaldehyde precursor in tissues by thiobarbituric acid test. Anal Biochem 1978;86: 271-8.  Back to cited text no. 12
    
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Sedlak J, Lindsay RH. Estimation of total, protein-bound, and nonprotein sulfhydryl groups in tissue with Ellman’s reagent. Anal Biochem 1968;25:192-205.  Back to cited text no. 13
    
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Aebi H. Catalase in vitro. Methods Enzymol 1984;105:121-6.  Back to cited text no. 14
    
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Madesh M, Balasubramanian KA. Microtiter plate assay for superoxide dismutase using MTT reduction by superoxide. Indian J Biochem Biophys 1998;35:184-8.  Back to cited text no. 15
    
16.
Pereira Wde F, Brito-Melo GE, de Almeida CA, et al. The experimental model of nephrotic syndrome induced by Doxorubicin in rodents: An update. Inflamm Res 2015;64:287-301.  Back to cited text no. 16
    
17.
Mohebbati R, Shafei MN, Soukhtanloo M, et al. Adriamycin-induced oxidative stress is prevented by mixed hydro-alcoholic extract of Nigella sativa and Curcuma longa in rat kidney. Avicenna J Phytomed 2016;6:86-94.  Back to cited text no. 17
    
18.
Lahoti TS, Patel D, Thekkemadom V, Beckett R, Ray SD. Doxorubicin-induced in vivo nephrotoxicity involves oxidative stress-mediated multiple pro- and anti-apoptotic signaling pathways. Curr Neurovasc Res 2012; 9:282-95.  Back to cited text no. 18
    
19.
Entezari Heravi N, Mohebbati R, Naji Ebrahimi Yazd Z, et al. Effect of Plantago major extract on Doxorubicin-induced nephropathy in rat. Physiol Pharmacol 2018;22:269-78.  Back to cited text no. 19
    
20.
Ayla S, Seckin I, Tanriverdi G, Cengiz M, Eser M, Soner BC, et al. Doxorubicin induced nephrotoxicity: Protective effect of nicotinamide. Int J Cell Biol 2011;2011: 390238.  Back to cited text no. 20
    
21.
Zhou L, Lin X, Abbasi AM, Zheng B. Phytochemical contents and antioxidant and antiproliferative activities of selected black and white sesame seeds. Biomed Res Int 2016; 2016:8495630.  Back to cited text no. 21
    
22.
Hsu DZ, Chiang PJ, Chien SP, Huang BM, Liu MY. Parenteral sesame oil attenuates oxidative stress after endotoxin intoxication in rats. Toxicology 2004;196:147-53.  Back to cited text no. 22
    
23.
Entezari Heravi N, Hosseinian S, Naji Ebrahimi Yazd Z, et al. Doxorubicin-induced renal inflammation in rats: Protective role of Plantago major. Avicenna J Phytomed 2018; 8:179-87.  Back to cited text no. 23
    
24.
Zhang R, Yu Y, Deng J, et al. Sesamin ameliorates high-fat diet-induced dyslipidemia and kidney injury by reducing oxidative stress. Nutrients 2016;8:E276.  Back to cited text no. 24
    
25.
Ramesh B. Beneficial effect of substitution of sesame oil on hepatic redox status and lipid parameters in streptozotocin diabetic rats. Int J Sci Nat 2011;2:488-93.  Back to cited text no. 25
    

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Correspondence Address:
Abolfazl Khajavi Rad
Department of Physiology, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad
Iran
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/1319-2442.344743

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