1. Introduction
Chemotherapy is an effective treatment for various kinds of cancer diseases, but it causes multiple side effects. The side effects of chemotherapy are different for each patient depending on the type of cancer, location, drug, dose, and the patient’s immune system. Most of the drugs including anticancer and their metabolites cleared from the renal pathway and causes major damage to the kidney [ 1 ]. Cyclophosphamide is one of the anticancer drugs with a wide spectrum of clinical uses, such as sarcoma, lymphoma, neuroblastoma, multiple myeloma, leukemia, breast cancer, ovarian and lung cancer, etc. Its typical side effects are multiple complications, including nephrotoxicity [ 2 ].
The metabolic activation of CP by the hepatic enzyme is 4-hydroxy cyclophosphamide, which converts into two toxic metabolites such as scarlein and phosphoramide [ 3 ]. This metabolite makes a covalent bond with DNA and proteins, resulting in cell death. A small fraction of CP is removed by the kidney, while the tubular reabsorption of this drug is very high [ 4 ]. Various researchers have reported that oxidative stress plays an important role in CP-induced renal injury [ 5 , 6 , 7 ]. Researchers have reported that CP-induced reactive oxygen species (ROS) play a key role in the pathogenesis of CP-independent renal damage. In this way, ROS-induced oxidative stress in the proximal tubules of the kidney mainly affects the cells, leading to nephrotoxicity [ 8 ]. Thus, natural scavengers of free radicals and antioxidants can be used to treat kidney damage caused by CP. It is now possible to reduce or eliminate the toxic effects of cyclophosphamide by combining it with various detoxifying and protective agents [ 9 ].
Therefore, palliative care as free radical scavengers and antioxidants is always necessary for cancer treatment, including CP. Natural resources have been used as a therapeutic medicine for thousands of years. Nevertheless, the majority of the population of a developing country relies on herbal medicine for primary health care [ 10 ]. The seeds of sesame plants and their oil have been used in human diets for thousands of years as a healthy food source [ 11 ]. Sesamin is an active ingredient of sesame seeds ( Sesamum indicum ) and sesame oil known for various physiological actions. Sesamin is a major lignin that is absorbed proficiently and circulated throughout the body. It is highly distributed in the liver and kidney as sesamin metabolites and plays various physiological effects such as antioxidant [ 12 ], anti-inflammatory [ 13 ], a protective role against mitochondrial toxicity [ 14 ], a lowering effect on serum cholesterol, lipid level, and blood pressure [ 15 , 16 ], a protective effect against alcohol-induced liver injury [ 17 ] and anti-allergic effect [ 18 ], etc. A recent study published by B H Ali (2020) [ 19 ] indicated sesamin ameliorated cisplatin-induced nephrotoxicity, but cyclophosphamide-induced nephrotoxicity has not been explored yet. Therefore, this idea encourages us to explore the defending action of Sesamin against cyclophosphamide-induced nephrotoxicity.
2. Results
2.1. Kidney Function Test
Table 1 showed blood urea nitrogen (BUN), uric acid, and creatinine content that increased significantly ( p < 0.001) in the CP administered group as compared to the normal control group. However, SM10 and SM20 treatment showed remarkable improvement and restoration in these active markers of the kidney. Significant improvement of these markers was seen in pretreatment with SM20 than SM10 as compared to CP administration, but no significant changes were seen in alone as positive control SM20 treatment vs. normal control.
2.2. Oxidative Stress Markers
Table 2 represents the MDA level that was remarkably amplified in the CP administration group than in the normal control group. The CP caused a significant decrease ( p < 0.05) in reduced glutathione (GSH), catalase (CAT), and superoxide dismutase (SOD) content compared to the normal control group (CNT). Pre-treatment with SM10 and SM20 significantly improved these levels (GSH, CAT, and SOD) and reduced the level of MDA. It was also noticed that SM20 was more effective and highly significant than SM10. The administration of SM10 failed to increase the level of SOD as compared to CP administration. Pre-treatment with SM20 alone as a positive control did not find any significant changes in these markers.
2.3. Inflammation and Apoptosis
Figure 1 and Figure 2 showed a significant ( p < 0.0001) increment in pro-inflammatory cytokines (IL-1β, and TNFα) after CP administration in comparison to CNT. In contrast, seven days of continuous pre-treatment with SM10 and SM20 drastically lowered these inflammatory cytokines in comparison to the CP alone administered group. However, these cytokines were not significant between SM20 alone and normal control groups. Figure 3 represents the effects of Sesamin on apoptosis markers. Interestingly, caspase 3 was increased significantly ( p < 0.0001) in the CP administered group, while pre-treatment of SM10 and SM20 remarkably decreased the level of caspase 3 in the SM10 + CP and SM20 + CP groups. It was noticed that no significant difference was found between SM20 alone and CNT.
2.4. Histopathology
Figure 4 B shows the harmful effects of cyclophosphamide (CP) on kidney histology in the CP treatment group in comparison to the normal control group (CNT). The impact of CP toxicities was very prompt with glomerulus nephritis, missing capillary loops, open Bowman’s space, edema, hemorrhage, and vacuolization, in the CP administered group than in CNT. However, pre-treatment with SM10 showed minor improvement in glomerulus morphology ( Figure 4 C), but remarkable regaining was observed in the SM20 group ( Figure 4 D) compared to CP alone. In the sesamin higher dose (SM20) treated group ( Figure 4 E), no lesions were observed. The normal control group (CNT) showed no sign of inflammation or necrosis ( Figure 4 A).
3. Discussion
Chemotherapeutic drugs improve cancer patient survival, but nephrotoxicity remains a critical complication and challenge. Cyclophosphamide is a chemotherapeutic drug that causes renal injuries at a different level that reflects in serum markers. In a previous report, cyclophosphamide therapy was reported to develop renal toxicity in cancer patients [ 20 , 21 ]. Current cancer therapeutic strategies are needed to avoid direct or indirect renal toxicity by using natural or complementary antioxidants. Sesamin was used in this study to explore the possible mechanism linked to antioxidant markers, inflammation markers, and apoptosis markers behind renal protection. Our findings showed that sesamin attenuates the CP-based nephrotoxicity in rats.
The progression of renal toxicity after CP administration is characterized by increases in serum BUN, uric acid, and creatinine [ 22 , 23 , 24 ]. The current outcomes showed a significant elevation in the BUN, uric acid, and creatinine in the CP administration group. The elevated serum markers may be due to renal damage, as evidenced by decreased creatinine clearance and low glomerular filtration rate. The measurement of these markers proves helpful in diagnosing renal toxicities [ 25 ].
In the past few decades, experimental animal research has shown that CP nephrotoxicity is linked with the free radical induced oxidative stress that mediates extensive renal impairments [ 26 ]. Cyclophosphamide causes renal toxicity by alkylation of the renal cell by the Cys sulfhydryl group of acrolein, one of CPs active toxic metabolites. As per scientific evidence, tubular reabsorption of CP is very high; therefore, a minimal amount is excreted from the kidneys [ 27 ]. CP experiences metabolic activation through liver enzymes and forms 4-hydroxycyclophosphamide that further converts into two metabolites, acrolein and phosphoramide mustard, which is cytotoxic [ 28 ]. Acrolein is an electrophilic and highly reactive aldehyde that interacts with the antioxidant defense system [ 29 ]. The acrolein can bind with reduced glutathione (GSH) and reduce its renal cell level. Thus, acrolein hampered the antioxidant system by producing more free radical generation, which is responsible for oxidative stress and damaging the cells of the kidney. The renal alkylation process reduces the glomerular filtration rate and causes tubular dysfunction [ 30 ].
The current observation indicates that a single dose of CP caused lipid peroxidation and MDA content were increased compared to CNT, while antioxidant enzymes were significantly reduced. These observations are supported by the previous investigator where CP produced lipid peroxidation and caused renal damage in rats [ 31 , 32 ]. Sesamin treatment remarkably restores these antioxidant defense systems in the present study. Sesamin has antioxidant [ 33 ], anti-inflammatory [ 34 ], and anti-apoptotic [ 35 ] properties, which are already proven. It has been suggested that sesamin’s metabolites (monocatechol and dicatechol) are responsible for various physiological effects. These metabolites exhibited antioxidant activity in the liver cells [ 36 ]. Sesamin was sequentially metabolized by cytochrome p450 and UDP glucuronosyltransferase or sulfotransferase. Sesamin metabolism was mainly mediated by CYP2C9 in the human liver [ 37 ]. A previous study indicates that sesamin suppresses increased NADPH oxidase-dependent oxidative stress in diabetic mice. NADPH inhibition may therefore result in various anti-inflammatory effects. As a result of oxidative stress, pro-inflammatory cytokines are produced as well as different signaling pathways are stimulated, resulting in inflammation, fibrosis, and apoptosis [ 38 , 39 ].
Inflammation shows a vital role in CP-involved renal pathogenesis by involving several molecular and cellular factors. As a result of CP-induced oxidative stress, the transcription factor NF-κB is activated in the nucleus, promoting a series of inflammatory changes and IL-1 and TNF-expression [ 40 ]. Thus, increased concentration of pro-inflammatory cytokines leads to activation of NF-κB, which results in increased transcription of the pro-apoptotic genes and eventually causes apoptotic cell death [ 41 ].
In the present study, these cytokines (IL-1β and TNFα) levels were increased in the kidneys of rats after exposure to CP, and their effects were significantly inhibited by the treatment with Sesamin. In addition, pro-inflammatory cytokines are also responsible for activating the p38-MAPK pathway that acts as a serious role in cellular apoptosis [ 42 , 43 ]. Apoptosis is programmed cell death associated with the activation of caspases, which is called the initiator or executioner of apoptosis [ 44 ].
Caspase 3 plays an important role in apoptosis in renal tubules [ 45 , 46 ]. This study found that caspase 3 levels increased after CP exposure and decreased after sesamin treatment.
In addition to the biochemical findings discussed previously, histopathological findings confirm that sesamin can attenuate renal injury induced by CP. In the presence of CP, infiltration of inflammatory cells and significant changes in renal architecture were noted, but sesamin ameliorated the effect and improved renal structure and function. Overall, the protective and antioxidant properties of sesamin contribute to the normal histological architecture of renal tissue.
4. Materials and Methods
4.1. Chemicals
Sesamin, cyclophosphamide, 5,5dithiobis-(2-nitrobenzoic acid), reduced GSH, TBA, were obtained from Sigma chemicals (3300 S 2nd St #3306 St. Louis, MO 63118, USA). Biochemical assay kits for BUN, Uric Acid, and Creatinine were obtained from Randox, Crumlin, UK. Elisa estimation kits for IL-1β, TNFα, and Caspase-3 were purchased from Abcam, Cambridge, UK.
4.2. Animals
Male Wistar rats weighing 150–160 g purchased from the MRC, Jazan University, Saudi Arabia. The rats were adapted under normal laboratory conditions of 12 h dark and light cycle, temperature (20 ± 2 °C), and humidity (50 ± 15%). The animal experiments were carried out after ethical approval no REC41/1-032 from the Research Ethical Committee, Jazan University.
4.3. Experimental Design
The rats were grouped into five test groups having six rats in each. Normal control (CNT) received the same number of vehicles as saline only. Test groups were further classified into four groups as rats received a single dose of Cyclophosphamide with saline i.p. on day four, represented as CP alone. The third and fourth groups of rats received two oral doses of sesamin (SM10 and 20 mg/kg) [ 47 ] from day one to day seven along with a single dose of CP (150 mg/kg) on day four, represented as SM10 + CP and SM20 + CP, for seven days respectively. Another group of rats received only sesamin (SM20 mg/kg) p.o. daily for seven days. Rats of all groups except normal and sesamin control were used as single dose administration intraperitoneally of CP (150 mg/kg body weight) [ 48 ] on day four.
After seven days of treatment (i.e., on day 8), rats fasted for a minimum of 12 h. All rats of each group were anesthetized with chloral hydrate, and blood was collected immediately from them through the retro-orbital plexus. After that, the animal was sacrificed, and kidneys were removed from each rat and divided into two parts. The first parts were kept at −80 °C for tissue preparation and biochemical analysis. In contrast, another part of the kidney was immersed in a 10% formalin solution for histological investigation by a light microscope. Samples of blood were centrifuged at 300 rpm for 15 min; serum was extracted and stored at −20 °C for kidney function tests. The tissue was homogenized in 10% child PB solution (0.1 M and pH 7.4). The homogenized tissue was centrifuged at 3000 rpm for 15 min at 4 °C, and the upper layer was used to estimate oxidative stress and cytokine assay.
4.4. Assessment of Renal Function Test in Serum
Determination of renal function tests (blood urea nitrogen, uric acid, and creatinine) was carried out as per the standard procedure of Randox UK using a photometer.
4.5. Assessment of Oxidative Stress Markers
Assessment of malondialdehyde (MDA) in renal tissue was carried out by Ohkawa et al. (1979). In a nutshell, the mixture contained 1.5 mL of 20% glacial acetic acid, 1.5 mL of aqueous solution of TBA, 0.2 mL of 8% SDS, and 0.8 mL of sample. This mixture was further heated at 95 °C for 60 min. After being cooled with tap water, the solution was agitated vigorously before 5 mL of a 15:1 butanol and pyridine combination and 1 mL of distilled water were added. The mixture was then centrifuged at 2000× g for 10 min, and the supernatant’s absorbance at 532 nm was then determined. The standard curve is used to calculate the amount of thiobarbituric reactive chemicals produced, and the result is represented in nmole per mg protein [ 49 ].
The Sedlak and Lindsay approach was used to measure reduced glutathione (GSH) (1968). The sample was precipitated with trichloroacetic acid after being homogenized in 5 mL of infant KCl (1%). (TCA). The reaction mixture contains 2 mL of Tris buffer (pH 8.9), 0.5 mL of DTNB, and 0.5 mL of supernatant. At 412 nm, the solution was read [ 50 ].
Claiborne’s 1975 approach was used to measure the catalase activity. The reaction mixture, which had a final volume of 3 mL, was made up of 2 mL of phosphate buffer, 0.95 mL of H 2 O 2 (20 mM), and 0.05 mL of sample PMS (10%). At intervals of 30 s, changes in absorbance were recorded at 240 nm. Using the molar extinction coefficient 43.6 M −1 CM −1 , catalase activity was determined as nmole H 2 O 2 consumed per minute per milligram protein [ 51 ].
Stevens et al. (2000) [ 52 ] approach was used to measure the superoxide dismutase (SOD) through measuring the auto-oxidation of epinephrine for 3 min at 480 nm wavelength. The enzymatic reaction of SOD was expressed in nmol epinephrine protected from oxidation/min/mg protein through using extinction co-efficient of 4.02 × 10 3 M −1 CM −1 .
4.6. Assessment of Inflammation and Apoptosis
Assessment of pro-inflammatory cytokines (IL-1β, and TNFα) and apoptosis (Caspase 3) were carried out as per the standard protocol of Abcam UK by using Elisa reader.
4.7. Histopathological Inspection
Formalin buffer solution (10%) was used to fix kidney tissues overnight, after which they were waxed and sectioned to 5 µm lengths. Hematoxylin and eosin (H and E) were used to stain the slides, and a 400× light microscope was used for the histopathological examination.
5. Conclusions
The current study concludes that sesamin (SM) is effective in attenuating oxidative stress and inflammatory cytokines associated with CP-induced nephrotoxicity in rats. No apparent adverse effects were noted during treatment with SM.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Author Contributions
Conceptualization, S.A. and M.F.A.; methodology, M.F.A., S.A.; dosing and sample collection, H.M.A.T., A.M.A.Z. and T.A.; formal analysis, T.A., A.M.J., Y.N., R.A.A., S.S.M. and M.F.A.; histopathology, M.F.A. and S.A.; data analysis, S.A., M.Q., R.A.A. and A.M.J.; writing and reviewing, M.F.A. and S.A. All authors have read and agreed to the published version of the manuscript.
Institutional Review Board Statement
Animals were used in this study according to guidelines approved by the Research Ethical Committee, Jazan University, Saudi Arabia (Aproval number REC41/1-032).
Data Availability Statement
The data presented in this study are available withing the article.
Conflicts of Interest
The authors declare no conflict of interest.
References
1. Lameire N. Nephrotoxicity of recent anti-cancer agents Clin. Kidney J. 2014 7 11 22 10.1093/ckj/sft135 25859345 PMC4389154
2. Khan T.S. Sundin A. Juhlin C. Wilander E. Öberg K. Eriksson B. Cisplatin, teniposide, and cyclophosphamide combination in the treatment of recurrent or meta-static adrenocortical cancer Med. Oncol. 2004 21 167 177 10.1385/MO:21:2:167 15299189
3. Hales F.B. Comparison of the mutagenicity and teratogenicity of cyclophosphamide and its active metabolites, 4-hydroxycyclophosphamide, phosphoramide mustard, and acrolein Cancer Res. 1982 42 3016 7046914
4. Milsted A.V.R. Jarman M. Metabolism of high doses of cyclophosphamide Cancer Chemother. Pharmacol. 1982 8 311 313 10.1007/BF00254056 7127662
5. Barry H. Free radicals, antioxidants, and human disease curiosity, cause, or consequence? Lancet 1994 344 721 724 7915779 10.1016/s0140-6736(94)92211-x
6. Devasagayam T.P.A. Tilak J.C. Boloor K.K. Sane K.S. Ghaskadbi S.S. Lele R.D. Free radicals and antioxidants in human health, Current status and future prospects J. Assoc. Phys. India 2004 52 794 804 15909857
7. Dumontet C. Drai J. Thieblemont C. Hequet O. Espinouse D. Bouafia F. Salles G. Coiffier B. The superoxide dismutase content in erythrocytes predicts short-term toxicity of high-dose cyclophosphamide Br. J. Haematol. 2001 112 405 409 10.1046/j.1365-2141.2001.02595.x 11167839
8. Ratliff B.B. Abdulmahdi W. Pawar R. Wolin M.S. Oxidant mechanisms in renal injury and disease Antioxid. Redox Signal. 2016 25 119 146 10.1089/ars.2016.6665 26906267 PMC4948213
9. Neboh E.E. Ufelle S.A. Myeloprotective activity of crude methanolic leaf extract of Cassia occidentalis in cyclophosphamideinduced bone marrow suppression in Wistar rats Adv. Biomed. Res. 2015 6 4 5 10.4103/2277-9175.148285 PMC4300593 25625111
10. Khan M.S.A. Ahmad I. Chapter 1—Herbal Medicine: Current Trends and Future Prospects New Look to Phytomedicine Khan M.S.A. Ahmad I. Chattopadhyay D. Academic Press Cambridge, MA, USA 2019 3 13 10.1016/B978-0-12-814619-4.00001-X 9780128146194
11. Alshahrani S. Al Sreaya A.A. Mashyakhi M.Y. Alqahtani S. Sivakumar S.M. Alhazmi H.A. Rehman Z. Alam F. Chemical characterization and antibacterial efficacy of Saudi sesame oil against human pathogenic bacteria Environ. Conserv. J. 2020 21 19 29 10.36953/ECJ.2020.211203
12. Fan D. Yang Z. Liu F. Jin Y.-G. Zhang N. Ni J. Yuan Y. Liao H.-H. Wu Q.-Q. Xu M. Sesamin protects against cardiac remodeling via Sirt3/ROS Pathway Cell Physiol. Biochem. 2020 44 2212 2227 10.1159/000486026 29248930
13. Fan D. Yang Z. Yuan Y. Wu Q.Q. Xu M. Jin Y.G. Tang Q.Z. Sesamin prevents apoptosis and inflammation after experimental myocardial infarction by JNK and NF-κB pathways Food Funct. 2017 8 2875 2885 10.1039/C7FO00204A 28726929
14. Takada S. Kinugawa S. Matsushima S. Takemoto D. Furihata T. Mizushima W. Fukushima A. Yokota T. Ono Y. Shibata H. Sesamin prevents decline in exercise capacity and impairment of skeletal muscle mitochondrial function in mice high-fat diet-induced diabetes Exp. Physiol. 2015 100 1319 1330 10.1113/EP085251 26300535 PMC5054862
15. Penalvo J.L. Hopia A. Adlercreutz H. Effect of sesamin on serum cholesterol and triglycerides levels in LDL receptor-deficient mice Eur. J. Nutr. 2006 45 439 444 10.1007/s00394-006-0617-8 17039285 PMC1705523
16. Kita S. Matsumura Y. Morimoto S. Akimoto K. Furuya M. Oka N. Tanaka T. Antihypertensive effect of sesamin. II. Protection against two-kidney, one-clip renal hypertension and cardiovascular hypertrophy Biol. Pharm. Bull. 1995 18 1283 1285 10.1248/bpb.18.1283 8845824
17. Akimoto K. Kitagawa Y. Akamatsu T. Hirose N. Sugano M. Shimizu S. Yamada H. Protective effects of sesamin against liver damage caused by alcohol or carbon tetrachloride in rodents Ann. Nutr. Metab. 1993 37 218 224 10.1159/000177771 8215239
18. Li L.C. Piao H.M. Zheng M.Y. Lin Z.H. Li G. Yan G.H. Sesamin attenuates mast cell-mediated allergic responses by suppressing the activation of p38 and nuclear factor-κB Mol. Med. Rep. 2016 13 536 542 10.3892/mmr.2015.4546 26573554
19. Ali B.H. Al Salam S. Al Suleimani Y. Al Za’abi M. Ashique M. Manoj P. Nemmar A. Ameliorative effect of sesamin in cispla-tin-induced nephrotoxicity in rats by suppressing inflammation, oxidative/nitrosative stress, and cellular damage Physiol. Res. 2020 69 61 72 10.33549/physiolres.934142 31852200 PMC8565953
20. Sinanoglu O. Yener A. Ekici S. Midi A. Aksungar F. The Protective effects of spirulina in cyclophosphamide induced nephrotoxicity and urotoxicity in rats Urology 2012 80 1392 1396 10.1016/j.urology.2012.06.053 22951000
21. Lameire N. Kruse V. Rottey S. Nephrotoxicity of anticancer drugs—An underestimated problem Acta Clin. Belg. 2011 66 337 345 22145268 10.2143/ACB.66.5.2062585
22. Cagle C.S. Appel S. Skelly A.H. Carter-Edwards L. Mid-life African-American women with type 2 diabetes, influence on work and the multicare giver role Ethn. Dis. 2002 12 555 566 12477143
23. Gines S. Seong I.S. Fossale E. Ivanova E. Trettel F. Gusella J.F. Wheeler V.C. Persichetti F. MacDonald M.E. Specific progressive cAMP reduction implicates energy deficit in presymptomatic Huntington’s disease knock-in mice Hum. Mol. Genet. 2003 12 497 508 10.1093/hmg/ddg046 12588797
24. Rehman J. Zhang H.J. Toth P.T. Zhang Y. Marsboom G. Hong Z. Salgia R. Husain A.N. Wietholt C. Archer S.L. Inhibition of mitochondrial fission prevents cell cycle progression in lung cancer FASEB J. 2012 26 2175 2186 10.1096/fj.11-196543 22321727 PMC3336787
25. Fuchs T.C. Hewitt P. Biomarkers for drug-induced renal damage and nephrotoxicity-an overview for applied toxicology AAPS J. 2011 13 615 631 10.1208/s12248-011-9301-x 21969220 PMC3231866
26. Ebrahimi B. Eirin A. Li Z. Zhu X. Zhang X. Lerman A. Textor S.C. Lerman L.O. Mesenchymal stem cells improve medullary inflammation and fibrosis after revascularization of swine atherosclerotic renal artery stenosis PLoS ONE 2013 8 e67474 10.1371/journal.pone.0067474 23844014 PMC3701050
27. Boddy A.V. Furtun Y. Sardas S. Sardas O. Idle J.R. Individual variation in the activation and inactivation of metabolic pathways of cyclophosphamide J. Nat. Cancer Inst. 1992 84 1744 1748 10.1093/jnci/84.22.1744 1433359
28. Mirkes P.E. Greenaway J.C. Rogers J.G. Brundrett R.B. Role of acrolein in cyclophosphamide teratogenicity in rat embryos in vitro Toxicol. Appl. Pharmacol. 1984 72 281 291 10.1016/0041-008X(84)90313-2 6546458
29. Senthilkumar S. Yogeeta S.K. Subashini R. Devaki T. Attenuation of cyclophosphamide induced toxicity by squalene in experimental rats Chem. Biol. Interact. 2006 160 252 260 10.1016/j.cbi.2006.02.004 16554041
30. Santos M.L.C. de Brito B.B. da Silva F.A.F. Botelho A.C.D.S. de Melo F.F. Nephrotoxicity in cancer treatment, An overview World J. Clin. Oncol. 2020 11 190 204 10.5306/wjco.v11.i4.190 32355641 PMC7186234
31. Haque R. Bin-Hafeez B. Parvez S. Pandey S. Aqueous extract of walnut ( Juglans regia L.) protects mice against cyclophosphamide induced biochemical toxicity Hum. Exp. Toxicol. 2003 22 473 480 10.1191/0960327103ht388oa 14580007
32. Manda K. Bhatia A.L. Prophylactic action of melatonin against cyclophosphamide induced oxidative stress in mice Cell Biol. Toxicol. 2003 19 367 372 10.1023/B:CBTO.0000013342.17370.16 15015761
33. 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 2578 10.1002/jsfa.7035 25472416
34. Monteiro É.M.H. Chibli L.A. Yamamoto C.H. Pereira M.C.S. Vilela F.M.P. Rodarte M.P. De Oliveira Pinto M.A. Da Penha Henriques do Amaral M. Silvério M.S. De Matos Araújo A.L.S. Antinociceptive and anti-inflammatory activities of the sesame oil and sesamin Nutrients 2014 6 1931 1944 10.3390/nu6051931 24824289 PMC4042560
35. Chen X. Ying X. Chen L. Zhang W. Zhang Y. Protective effects of sesamin on liver fibrosis through antioxidative and anti-inflammatory activities in rats Immunopharmacol. Immunotoxicol. 2015 37 465 472 10.3109/08923973.2015.1085064 26466645
36. Kiso Y. Antioxidative roles of sesamin, a functional lignan in sesame seed, and it’s effect on lipid- and alcohol-metabolism in the liver: A DNA microarray study Biofactors 2004 21 191 196 10.1002/biof.552210139 15630196
37. Sakaki T. Yasuda K. Nishikawa M. Ikushiro S. Metabolism of Sesamin and Drug-Sesamin Interaction Yakugaku Zasshi 2018 138 357 363 (In Japanese) 10.1248/yakushi.17-00191-4 29503429
38. Sun Y. Ito S. Nishio N. Tanaka Y. Chen N. Isobe K.-I. Acrolein induced both pulmonary inflammation and the death of lung epithelial cells Toxicol. Lett. 2014 229 384 392 10.1016/j.toxlet.2014.06.021 24999835
39. Mythili Y. Sudharsan P.T. Selvakumar E. Varalakshmi P. Protective effect of DL-alpha-lipoic acid on cyclophosphamide induced oxidative cardiac injury Chem. Biol. Interact. 2004 151 13 19 10.1016/j.cbi.2004.10.004 15607758
40. El-Kholy A.A. Elkablawy M.A. El-Agamy D.S. Lutein mitigates cyclophosphamide induced lung and liver injury via NF-κB/MAPK dependent mechanism Biomed. Pharmacother. 2017 92 519 527 10.1016/j.biopha.2017.05.103 28575809
41. Kumar A. Takada Y. Boriek A.M. Aggarwal B.B. Nuclear factor-κB, its role in health and disease J. Mol. Med. 2004 82 434 448 10.1007/s00109-004-0555-y 15175863
42. Alhoshani A.R. Hafez M.M. Husain S. Al-sheikh A.M. Alotaibi M.R. Al Rejaie S.S. Alshammari M.A. Almutairi M.M. Al-Shabanah O.A. Protective effect of rutin supplementation against cisplatin-induced Nephrotoxicity in rats BMC Nephrol. 2017 18 194 10.1186/s12882-017-0601-y 28619064 PMC5472980
43. Galluzzi L. Vitale I. Abrams J.M. Alnemri E.S. Baehrecke E.H. Blagosklonny M.V. Dawson T.M. Dawson V.L. El-Deiry W.S. Fulda S. Molecular definitions of cell death subroutines, recommendations of the Nomenclature Committee on Cell Death 2012 Cell Death Differ. 2012 19 107 120 10.1038/cdd.2011.96 21760595 PMC3252826
44. Temel Y. Kucukler S. Yıldırım S. Caglayan C. Kandemir F.M. Protective effect of chrysin on cyclophosphamide-induced hepatotoxicity and nephrotoxicity via the inhibition of oxidative stress, inflammation, and apoptosis Naunyn-Schmiedeberg’s Arch Pharmacol. 2020 393 325 337 10.1007/s00210-019-01741-z 31620822
45. Nafees S. Rashid S. Ali N. Hasan S.K. Sultana S. Rutin ameliorates cyclophosphamide induced oxidative stress and inflammation in Wistar rats, role of NFκB/MAPK pathway Chem. Biol. Interact. 2015 231 98 107 10.1016/j.cbi.2015.02.021 25753322
46. Liu Q. Lin X. Li H. Yuan J. Peng Y. Dong L. Dai S. Paeoniflorin ameliorates renal function in cyclophosphamide-induced mice via AMPK suppressed inflammation and apoptosis Biomed. Pharmacother. 2016 84 1899 1905 10.1016/j.biopha.2016.10.097 27829543
47. Lv D. Zhu C.Q. Liu L. Sesamin ameliorates oxidative liver injury induced by carbon tetrachloride in rat Int. J. Clin. Exp. Pathol. 2015 8 5733 5738 26191289 PMC4503160
48. Abraham P. Isaac B. The effects of oral glutamine on cyclophosphamide-induced nephrotoxicity in rats Hum. Exp. Toxicol. 2011 30 616 623 10.1177/0960327110376552 20621952
49. Ohkawa H. Ohishi N. Yagi K. Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction Anal. Biochem. 1979 95 351 358 10.1016/0003-2697(79)90738-3 36810
50. Sedlak J. Lindsay R.H. Estimation of total, proteinbound, and nonprotein sulfhydryl groups in tissue with Ellman’s reagent Anal. Biochem. 1968 25 192 205 10.1016/0003-2697(68)90092-4 4973948
51. Claiborne A.L. Catalase activity CRC Handbook of Methods for Oxygen Radical Research CRC Boca Raton, FL, USA 1985 Volume 1 283 284
52. Stevens M.J. Obrosova I. Cao X. Van Huysen C. Greene D.A. Effects of DL-alpha-lipoic acid on peripheral nerve conduction, blood flow, energy metabolism and oxidative, blood flow, energy metabolism, and oxidative stress in experimental diabetic neuropathy Diabetes 2000 49 1006 1015 10.2337/diabetes.49.6.1006 10866054