|Year : 2013 | Volume
| Issue : 2 | Page : 230-234
Ayurvedic formulations ameliorate cisplatin-induced nephrotoxicity: Preclinical studies on Brahma Rasayana and Chyavanaprash
Aditya Menon, Cherupally Krishnan Krishnan Nair
Deptartment of Radiation Biology, Pushpagiri Institute of Medical Sciences and Research Centre, Thiruvalla, Kerala, India
|Date of Web Publication||13-Jun-2013|
Cherupally Krishnan Krishnan Nair
Deptartment of Radiation Biology, Pushpagiri Institute of Medical Sciences and Research Centre, Thiruvalla - 689 101, Kerala
Source of Support: Board of Research in Nuclear Sciences, Department of
Atomic Energy., Conflict of Interest: None
Aim of Study: To explore the ability of two Ayurvedic formulations, Brahma Rasayana (BRM) and Chyavanaprash (CHM) in alleviating Cisplatin (Cis-dichlorodiammineplatinum [II] CDDP) induced acute nephrotoxicity.
Materials and Methods: Swiss albino mice were administered with CDDP (12 mg/kg, i.p) and two doses of BRM or CHM (1 and 2 g/kg). Various antioxidant parameters in the kidney as well as release of marker enzymes in the serum were assayed. Histology of the kidney was also performed to check for CDDP induced damages.
Results: Administration of either BRM or CHM (1 and 2 g/kg) maintained the antioxidant status in the kidney thereby preventing tissue damage as well as the release of marker enzymes. CDDP induced variation of renal architecture was also prevented by BRM and CHM administration.
Conclusion: BRM and CHM administration could prevent CDDP induced acute renal toxicity.
Keywords: Antioxidant status, Brahma Rasayana, Chyavanaprash, cisplatin, nephrotoxicity
|How to cite this article:|
Menon A, Krishnan Nair CK. Ayurvedic formulations ameliorate cisplatin-induced nephrotoxicity: Preclinical studies on Brahma Rasayana and Chyavanaprash. J Can Res Ther 2013;9:230-4
|How to cite this URL:|
Menon A, Krishnan Nair CK. Ayurvedic formulations ameliorate cisplatin-induced nephrotoxicity: Preclinical studies on Brahma Rasayana and Chyavanaprash. J Can Res Ther [serial online] 2013 [cited 2020 May 28];9:230-4. Available from: http://www.cancerjournal.net/text.asp?2013/9/2/230/113363
| > Introduction|| |
Cisplatin (Cis-dichlorodiammineplatinum [II] CDDP) represents a class of antineoplastic drugs containing a heavy metal, platinum. It is effective against several human tumors, e.g. testis, ovary, head, neck and lung. , Though high doses of CDDP is preferred for tumour therapy,  its optimal clinical use is limited due to the numerous side effects it produces like nephrotoxicity, neurotoxicity, bone marrow toxicity, gastrointestinal toxicity and ototoxicity. ,, Among these, nephrotoxicity is the major side effect of CDDP since kidney accumulates cisplatin to a greater degree than other organs and is the major route for its excretion. ,
Recent evidences suggest the involvement of free radicals, depletion of GSH and other cellular antioxidant enzymes, in the induction of nephrotoxicity by CDDP. ,, Since free radicals bring out the basic mechanism of CDDP induced nephrotoxicity, it is rational to consider antioxidants as therapeutics to mitigate its toxicity. Various studies have reported the use of antioxidants in mitigating nephrotoxicity caused by CDDP. ,,
Ayurveda, the Indian holistic healthcare system of traditional medicines made using natural ingredients  have several potent antioxidant formulations. Two such formulations Brahma Rasayana (BRM) and Chyavanaprash (CHM) are evaluated in the present study for their nephroprotective ability since several ingredients of BRM and CHM are proved to be potent antioxidants. , These formulations have shown promise in alleviating various free radicals induced physiological conditions. , Also these are reported to confer tissue regeneration capability  and antitumour activity. ,, These findings have encouraged us to further investigate its potential during cancer therapy. As an initial step, the present study envisages to find out the ability of these Ayurvedic formulations in reducing nephrotoxicity induced by CDDP, which is a widely used chemotherapeutic agent, in mice model.
| > Materials and Methods|| |
Brahma Rasayana and Chyavanaprash, the two rasayana formulations prepared according to Sahasrayogam  were from reputed manufacturers available locally. All the other chemicals and reagents used in this study were of analytical grade.
Swiss albino mice of 8-10 weeks old, weighing 22-25 g was obtained from the Small Animal Breeding Section (SABS), Kerala Agricultural University, Mannuthy, Thrissur, Kerala. They were kept under standard conditions of temperature and humidity in the Centre's Animal House Facility. The animals were provided with standard mouse chow (Sai Durga Feeds and Foods, Bangalore, India) and water ad libitum. All animal experiments in this study were carried out with the prior approval of the Institutional Animal Ethics Committee (IAEC) and were conducted strictly adhering to the guidelines of Committee for the purpose of Control and Supervision of Experiments on Animals (CPCSEA) constituted by the Animal Welfare Division of Government of India.
Animals were randomly divided into 6 groups of five each as detailed below. Group I was kept as the untreated control and all the other groups received CDDP (12 mg/kg, i.p). Group III and Group IV were administered with BRM or CHM (2 g/kg, p.o) after 1 hour of CDDP injection, i.p.
CDDP (12 mg/kg, i.p).
CDDP (12 mg/kg, i.p) + BRM (1 g/kg, p.o).
CDDP (12 mg/kg, i.p) + BRM (2 g/kg, p.o).
CDDP (12 mg/kg, i.p) + CHM (1 g/kg, p.o).
CDDP (12 mg/kg, i.p) + CHM (2 g/kg, p.o).
Assessment of nephrotoxicity
72 hours after CDDP treatment, blood was collected by cardiac puncture and serum was separated for biochemical analysis. Kidney was excised and washed with ice-cold phosphate buffered saline (PBS) and homogenates 10% (w/v) were prepared in PBS. Serum creatinine level was determined by alkaline picric acid method and serum urea level was determined by diacetylmonoxime (DAM) reagent (Agappe Diagnostic Pvt. Ltd.; Ernakulam, Kerala, India). Level of GSH was assayed by the method of Moron (1974),  based on the reaction with DTNB. GPx activity was measured based on the method of Hafeman (1974),  based on the degradation of H 2 O 2 . Activity of SOD was measured by NBT reduction method of Mc Cord and Fridovich (1969).  Protein levels in the tissue were measured by following the method of Lowry (1951).  Levels in peroxidation of membrane lipids were done based on the method of Buege and Aust (1987).  For histopathological studies kidney was fixed in 10% formalin solution immediately after sacrifice, dehydrated in graded ethanol, cleared in xylene, and embedded in paraffin. Five-micron-thick sections made using a microtome were mounted on glass slides, dewaxed, rehydrated with distilled water, and stained with hematoxylin and eosin and mounted in DPX.  The slides were observed in light microscope under oil immersion microscope (100 X) and photographed.
The results are presented as Mean ± SD of the studied group. Statistical analyses of the results were performed using ANOVA with Tukey-Kramer multiple comparisons test.
| > Results|| |
[Table 1] gives the cellular antioxidant levels (GSH, GPx and SOD). The results indicated that a single dose of CDDP (12 mg/kg) significantly decreased the tissue antioxidants. Oral administration of BRM or CHM not only helped in maintaining these antioxidants at normal levels, but also improved the total antioxidant status when compared to the control untreated animals.
|Table 1: Effect of BRM or CHM on antioxidants level in CDDP induced nephrotoxicity in mice. Values are expressed as mean ± SD (n = 5)|
Click here to view
Peroxidation of membrane lipids
As presented in [Figure 1], CDDP (12 mg/kg) injection resulted in an increase in the levels of peroxidation of membrane lipids from 4.78 ± 1.56 to 8.34 ± 1.98 nMoles/mg protein. Administration of BRM or CHM helped in reducing the extent of peroxidation. BRM administration at a dose of 1 g/kg and 2 g/kg restored the level of peroxidation to 2.95 ± 1.17 and 2.15 ± 0.32 respectively. CHM administration at a dose of 1 g/kg and 2 g/kg restored the level of peroxidation to 3.01 ± 1.02 and 2.25 ± 0.43 respectively.
|Figure 1: Effect of BRM or CHM on peroxidation of membrane lipids in CDDP induced nephrotoxicity in mice. Values are expressed as mean ± SD (n = 5). ***indicates 'P < 0.001' when compared with respective controls|
Click here to view
Serum biochemical parameters
The levels of various serum parameters presented in [Table 2] suggest that CDDP (12 mg/kg) administration increased these enzyme levels while the administration of BRM or CHM prevented the increase in the serum enzyme levels following CDDP treatment.
|Table 2: Effect of BRM or CHM on serum marker enzymes in CDDP induced nephrotoxicity in mice. Values are expressed as mean ± SD (n = 5)|
Click here to view
Histopathological investigation showed that, in CDDP (12 mg/kg) treated mice kidney there is a decreased cellularity of the glomeruli and edema of the lining of epithelial cells in the renal tubules. More over the nuclei of the lining cells show vacoulation. The interstitial tissue also showed oedema as can be evident from [Figure 2]b. The renal tissues of CDDP treated mice when administered with BRM or CHM after the CDDP treatment, showed near normal architecture with normal glomerular, renal tubules and interstitial tissue appearance [Figure 2]c-f.
|Figure 2: Effect of BRM and CHM various doses (1 and 2 g/kg) on CDDP induced renal damage in mice (× 40)|
Click here to view
| > Discussion|| |
There are 2 suggested mechanisms by which CDDP inflicts nephrotoxicity.  One mechanism is by the cleavage of GSH by the enzyme gamma glutamyl transpeptidase (GGT) thereby releasing cysteinyl-glycine, which is further cleaved into cysteine and glycine by diaminopeptidase N.  Depletion of tissue GSH is a major factor which weakens the cell's defence against ROS mediated peroxidative cell injury. 
The other mechanism of renal toxicity is due to the formation of nephrotoxic metabolites of CDDP through a GSH-conjugate intermediate, a majority of which occurs in the kidney.  It has also been shown that inhibition of GSH reduces the toxicity of CDDP. ,
CDDP elicits its action against tumour cells by the activation of apoptotic pathways.  In recent years, kidney tubular apoptosis has been identified as a common pathway in response to cellular stress applied at intensity below the threshold for necrosis,  and this holds true in case of CDDP toxicity also. 
Cisplatin is known to accumulate in mitochondria of renal epithelial cells and induces ROS production, resulting in reduction of antioxidant status of kidney along with peroxidation of membrane lipids. , Various antioxidant molecules have promise in mitigating the nephrotoxicity induced by CDDP. ,, The present work on the attempt to find the ability of two antioxidant Ayurvedic formulations in mitigating CDDP induced nephrotoxicity have produced interesting results, since 2 over the counter formulations are able to mitigate the adverse effects of a potent anticancer drug. These formulations have been reported to have antioxidant activity under in vitro conditions and also under in vivo conditions in various tissues, following consumption.  Administration of BRM or CHM prevented the CDDP induced nephrotoxicity by restoring the total antioxidant content. BRM and CHM are formulations used in 'rasayana therapy' in Ayurveda, which is a dedicated treatment modality for immune promotive, antidegenerative and rejuvenative health care.  The capacity to repair and regenerate damaged tissue, may also have contributed to the nephroprotective property of these formulations.
In the present study, a single dose of CDDP (12 mg/kg) induced nephrotoxicity was manifested biochemically by the increase in the levels of serum creatinine and urea. These results were consistent with previous studies reported by other investigators. , Administration of BRM and CHM prevented the alterations in renal haemodynamics by CDDP administration.
The present study convincingly proved that BRM or CHM administration prevents the nephrotoxic effect of CDDP. Screening the vast repertoire of Ayurveda can yield in the discovery of formulations which can be useful in preventing the side effects of chemotherapeutic drugs. Further studies are to be conducted to know whether these formulations can offer preferential protection to normal tissues without affecting the antineoplastic activity of CDDP.
| > Acknowledgement|| |
The authors thank Board of Research in Nuclear Sciences, Department of Atomic Energy for the financial support.
| > References|| |
|1.||Loehrer PJ, Einhorn LH. Drugs five years later. Cisplatin. Ann Intern Med 1984;100:704-13. |
|2.||Meyer KB, Medias E. Cisplatin nephrotoxicity. Miner Electrolyte Metab 1994;20:201-13. |
|3.||Di Re F, Bohm S, Oriana S, Spatti GB, Zunino F. Efficacy and safety of high-dose cisplatin and cyclophosphamide with glutathione protection in the treatment of bulky advanced epithelial ovarian cancer. Cancer Chemother Pharmacol 1990;25:355-60. |
|4.||Antunes LM, Darin JD, Bianchi MD. Protective effect of vitamin C against cisplatin-induced nephrotoxicity and lipid peroxidation in adult rats: A dose dependant study. Pharmacol Res 2000;41:405-11. |
|5.||Lynch ED, Gu R, Pierce C, Kil J. Reduction of acute cisplatin ototoxicity and nephrotoxicity in rats by oral administration of allopurinol and ebselen. Hear Res 2005;201:81-9. |
|6.||Choie DD, Longnecker DS, del Campo AA. Acute and chronic cisplatin nephropathy in rats. Lab Invest 1981;44:397-402. |
|7.||Mansour MA, Mostafa AM, Nagi MN, Khattab MM, Al-Shabanah OA. Protective effect of aminoguanidine against nephrotoxicity induced by cisplatin in normal rats. Comp Biochem Physiol C Toxicol Pharmacol 2002;132:123-8. |
|8.||Yao X, Panichpisal K, Kurtzman N, Nugent K. Cisplatin Nephrotoxicity: A review. Am J Med Sci 2007;334:115-24. |
|9.||Matsushima H, Yonemura K, Ohishi K, Hishida A. The role of oxygen free radicals in cisplatin-induced acute renal failure in rats. J Lab Clin Med 1998;131:518-26. |
|10.||Yildirim Z, Sogut S, Odaci E, Iraz M, Ozyurt H, Kotuk M, et al. Oral erdosteine administration attenuates cisplatin-induced renal tubular damage in rats. Pharmacol Res 2003;47:149-56. |
|11.||Dekant W. Bioactivation of nephrotoxins and renal carcinogens by glutathione S-conjugate formation. Toxicol Lett 1993;67:151-60. |
|12.||Saad SY, Al-Rikabi AC. Protection effects of Taurine supplementation against cisplatin-induced nephrotoxicity in rats. Chemotherapy 2002;48:42-8. |
|13.||Al-Majed AA, Abd-Allah AR, Al-Rikabi AC, Al-Schbanah OA, Mostafa AM. Effect of oral administration of Arabic gum on cisplatin-induced nephrotoxicity in rats. J Biochem Mol Toxicol 2003;17:146-53. |
|14.||Antunes LM, Darin JD, Bianchi Nde L. Effects of the antioxidants curcumin or selenium on cisplatin-induced nephrotoxicity and lipid peroxidation in rats. Pharmacol Res 2001;43:145-50. |
|15.||Patwardhan B, Vaidya AD, Chorghade M, Joshi SP. Reverse Pharmacology and Systems Approaches for Drug Discovery and Development. Curr Bioact Compd 2008;4:201-12. |
|16.||Ramnath V, Rekha PS, Sujatha KS. Amelioration of heat stress induced disturbances of antioxidant defence system in chicken by Brahma Rasayana. Evid Based Complement Alternat Med 2008;5:77-84. |
|17.||Parle M, Bansal N. Traditional medicinal formulation, Chyawanprash - A review. Indian J Tradit Know 2006;5:484-8. |
|18.||Jose JK, Kuttan R. Hepatoprotective activity of Emblica officinalis and Chyavanaprash. J Ethnopharmacol 2000;72:135-40. |
|19.||Rekha PS, Kuttan G, Kuttan R. Antioxidant activity of brahma rasayana. Indian J Exp Biol 2001;39:447-52. |
|20.||Singh RH, Narsimhamurthy K, Singh G. Neuronutrient impact of Ayurvedic Rasayana therapy in brain aging. Biogerontology 2008;9:369-74. |
|21.||Jose JK, Kuttan G, Kuttan R. Antitumour activity of Emblica officinalis. J Ethnopharmacol 2001;75:65-9. |
|22.||Praveenkumar V, Kuttan R, Kuttan G. Effect of Rasayanas on normal and tumor-bearing animals. J Exp Clin Cancer Res 1994;13:67-70. |
|23.||Rekha PS, Kuttan G, Kuttan R. Effect of Brahma Rasayana on antioxidant systems and cytokine levels in mice during cyclophosphamide administration. J Exp Clin Cancer Res 2001;20:219-23. |
|24.||Krishna Vaidyan KV, Gopalapillai S, Sahasrayogam (Sanskrit). Alappuzha: Vidyarambam Press and Books Depot; 1969. |
|25.||Moron MS, Depierre JW, Mannervick B. Levels of glutathione, glutathione-S- transferase actovoties in rat liver. Biochim Biophys Acta 1979;582:67-78. |
|26.||Hafeman DG, Sundae RA, Houestru WG. Effect of dietary selenium on erythrocyte and liver glutathione peroxidase in rat. J Nutr 1974;104:580-7. |
|27.||Mc Cord JM, Fridovich I. Super oxide dismutase enzyme function for erythrocaprein. J Biol Chem 1969;244:6049-55. |
|28.||Lowry OH, Rosenblum NJ, Farr AL, Randall RJ. Protein measurement with Folin phenol reagent. J Biol Chem 1951;193:265-75. |
|29.||Buege JA, Aust SD. Microsomal lipid peroxidation. Methods Enzymol 1978;52:302-10. |
|30.||Galeano M, Altavilla D, Cucinotta D, Russo GT, Calò M, Bitto A, et al. Recombinant human erythropoietin stimulates angiogenesis and wound healing in the genetically diabetic mouse. Diabetes 2004;53:2509-17. |
|31.||Hanigan MH, Devarajan P. Cisplatin nephrotoxicity: Molecular mechanisms. Cancer Ther 2003;1:47-61. |
|32.||McIntyre T, Curthoys NP. Renal catabolism of glutathione: Characterization of a particulate rat renal dipeptidase that catalyzes the hydrolysis of cysteinylglycine. J Biol Chem 1982;257:11915-21. |
|33.||DeLeve LD, Kaplowitz N. Importance and regulation of hepatic glutathione. Semin Liver Dis 1990;10:251-66. |
|34.||Mistry P, Lee C, McBrien DC. Intracellular metabolites of cisplatin in the rat kidney. Cancer Chemother Pharmacol 1989;24:73-9. |
|35.||Sadzuka Y, Shimizu Y, Takino Y. Role of glutathione S-transferase isoenzymes in cisplatin-induced nephrotoxicity in the rat. Toxicol Lett 1994;70:211-22. |
|36.||Sadzuka Y, Shimizu Y, Takino Y, Hirota S. Protection against cisplatin-induced nephrotoxicity in the rat by inducers and an inhibitor of glutathione S-transferase. Biochem Pharmacol 1994;48:453-9. |
|37.||Friesen C, Fulda S, Debatin KM. Cytotoxic drugs and the CD95 pathway. Leukemia 1999;13:1854-8. |
|38.||Ueda N, Kaushal GP, Shah SV. Apoptotic mechanisms in acute renal failure. Am J Med 2000;108:403-15. |
|39.||Lieberthal W, Triaca V, Levine J. Mechanisms of death induced by cisplatin in proximal tubular epithelial cells: Apoptosis vs. necrosis. Am J Physiol 1996;39:F700-8. |
|40.||Huang Q, Dunn RT 2 nd , Jayadev S, DiSorbo O, Pack FD, Farr SB, et al. Assessment of cisplatin-induced nephrotoxocity by microarray technology. Toxicol Sci 2001;63:196-207. |
|41.||Safirstein R, Miller P, Guttenplan JB. Uptake and metabolism of cisplatin by rat kidney. Kidney Int 1984;25:753-8. |
|42.||Mingeot-Leclerq M, Tulkens PM. Aminoglycosides: Nephrotoxicity. Antimicrob Agents Chemother 1999;43:1003-12. |
|43.||Badary OA, Abdel-Maksoud S, Ahmed A, Owieda GH. Naringenin attenuates cisplatin nephrotoxicity in rats. Life Sci 2005;76:2125-35. |
|44.||Badary OA, Nagi MN, Al-Sawaf HA, Al-Harbi M, Al-Beikairi AM. Effect of L-histidinol on cisplatin nephrotoxicity in rat. Nephron 1997;77:435-9. |
|45.||Chulet R, Pradhan P. A review on rasayana. Pharmacogn Rev 2009;3:229-34. |
|46.||Atessahin A, Yilmaz S, Karahan I, Ceribasi AO, Karaoglu A. Effects of lycopene against cisplatin- induced nephrotoxicity and oxidative stress in rats. Toxicology 2005;212:116-23. |
|47.||Mansour HH, Hafez HF, Fahmy NM. Silymarin modulates cisplatin-induced oxidative stress and hepatotoxicity in rats. J Biochem Mol Biol 2006;39:656-61. |
[Figure 1], [Figure 2]
[Table 1], [Table 2]