The Effect of Whey Proteins on the Brain and Small Intestine Nitric Oxide Levels: Protein Profiles in Methotrexate-Induced Oxidative Stress

Abstract

Objectives: The aim of this study was to determine the effects of whey proteins on methotrexate (MTX)-induced brain and small intestine damage.

Materials and Methods: 30 Sprague Dawley rats (200-300 g) were divided into four groups: Control, control + whey, MTX, and MTX+whey. MTX was administered at 20 mg/kg (single dose) intraperitoneally to the MTX group rats, and 2 mg/kg of whey protein were administered by oral gavage for 10 days to the whey groups. Lipid peroxidation, glutathione, and nitric oxide (NO) levels, as well as glutathione-S-transferase and superoxide dismutase activities were measured in the brain and small intestine. SDS-polyacrylamide gel electrophoresis of the brain and intestine tissues were also carried out.

Results: While MTX treatment caused oxidative damage in the brain and small intestine, whey protein administration ameliorated MTX-induced oxidative stress. MTX administration did not change the brain’s NO level, while an increase in intestinal NO level was detected.

Conclusion: MTX induced oxidative stress in the brain and small intestine changed the protein metabolism in these tissues regardless of reduced food intake. Consecutive 10-day administration of whey proteins has shown its therapeutic effect on MTX-induced brain and small intestine oxidative damage.

Keywords

• Methotrexate
• whey protein
• brain
• small intestine
• nitric oxide
• oxidative stress

References

1. 1. Hannoodee M, Mittal M. Methotrexate. StatPearls (Internet): StatPearls Publishing; 2021. google scholar
2. 2. Letertre MP, Munjoma N, Wolfer K, Pechlivanis A, McDonald JA, Hardwick RN, et al. A two-way interaction between methotrexate and the gut microbiota of male sprague-dawley rats. J Proteome Res 2020; 19(8): 3326-39. [CrossRef] google scholar
3. 3. Maeda T, Miyazono Y, Ito K, Hamada K, Sekine S, Horie T. Oxidative stress and enhanced paracellular permeability in the small intestine of methotrexate-treated rats. Cancer Chemother Pharmacol 2010; 65(6): 1117-23. [CrossRef] google scholar
4. 4. Jahovic N, Şener G, Çevik H, Ersoy Y, Arbak S, Yeğen BÇ. Amelioration of methotrexate-induced enteritis by melatonin in rats. Cell Biochem Funct. 2004; 22(3): 169-78. [CrossRef] google scholar
5. 5. Miyazono Y, Gao F, Horie T. Oxidative stress contributes to methotrexate-induced small intestinal toxicity in rats. Scand J Gastroenterol 2004; 39(11): 1119-27. [CrossRef] google scholar
6. 6. Welbat JU, Naewla S, Pannangrong W, Sirichoat A, Aranarochana A, Wigmore P. Neuroprotective effects of hesperidin against methotrexate-induced changes in neurogenesis and oxidative stress in the adult rat. Biochem Pharmacol 2020; 178: 114083. [CrossRef] google scholar
7. 7. Vezmar S, Becker A, Bode U, Jaehde U. Biochemical and Clinical Aspects of Methotrexate Neurotoxicity. Chemotherapy 2003; 49(1-2): 92-104. [CrossRef] google scholar
8. 8. Townsend DM, Tew KD. The role of glutathione-S-transferase in anti-cancer drug resistance. Oncogene 2003; 22(47): 7369-75. [CrossRef] google scholar

9. 9. Tsurui K, Kosakai Y, Horie T, Awazu S. Vitamin A protects the small intestine from methotrexate-induced damage in rats. J Pharmacol Exp Ther 1990; 253(3): 1278-84. google scholar
10. 10. Horie T, Matsumoto H, Kasagi M, Sugiyama A, Kikuchi M, Karasawa C, et al. Protective effect of aged garlic extract on the small intestinal damage of rats induced by methotrexate administration. Planta Med 1999; 65(06): 545-8. [CrossRef] google scholar
11. 11. Corrochano AR, Arranz E, De Noni I, Stuknyte M, Ferraretto A, Kelly PM, et al. Intestinal health benefits of bovine whey proteins after simulated gastrointestinal digestion. J Funct Foods 2018; 49: 52635. [CrossRef] google scholar
12. 12. Sprong RC, Schonewille AJ, van der Meer R. Dietary cheese whey protein protects rats against mild dextran sulfate sodium-induced colitis: role of mucin and microbiota. J Dairy Sci 2010; 93(4): 136471. [CrossRef] google scholar
13. 13. Akal C. Benefits of Whey Proteins on Human Health. Chapter 28; In: Watson RR, Collier RJ, Preedy VR, editors. Dairy in Human Health and Disease Across the Lifespan: Academic Press; 2017. p. 363-72. [CrossRef] google scholar
14. 14. Aykaç A, Becer E, Özbeyli D, Şener G, Başer K. Protective effects of Origanum onites essential oil in the methotrexate-induced rat model: role on apoptosis and hepatoxicity. Rec Nat Prod 2020; 14(6): 395-404. [CrossRef] google scholar
15. 15. Shimizu Y, Hara H, Hira T. Glucagon-like peptide-1 response to whey protein is less diminished by dipeptidyl peptidase-4 in comparison with responses to dextrin, a lipid and casein in rats. Br J Nutr 2021; 125(4): 398-407. [CrossRef] google scholar
16. 16. Ledwozyw A, Michalak J, Stepien A, Kadziolka A. The relationship between plasma triglycerides, cholesterol, total lipids and lipid peroxidation products during human atherosclerosis. Clin Chim Acta 1986; 155(3): 275-83. [CrossRef] google scholar
17. 17. Mylroie AA, Collins H, Umbles C, Kyle J. Erythrocyte superoxide dismutase activity and other parameters of copper status in rats ingesting lead acetate. Toxicol Appl Pharmacol 1986; 82(3): 51220. [CrossRef] google scholar
18. 18. Habig WH, Jakoby WB. Assays for differentiation of glutathione S-transferases. Methods Enzymol 1981; 77: 398-405. [CrossRef] google scholar
19. 19. Miranda KM, Espey MG, Wink DA. A rapid, simple spectrophotometric method for simultaneous detection of nitrate and nitrite. Nitric Oxide 2001; 5(1): 62-71. [CrossRef] google scholar
20. 20. Beutler E. Red cell metabolism: a manual of biochemical methods. 3rd edition, New York, 1984. google scholar
21. 21. Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 1970; 227(5259): 680-5. [CrossRef] google scholar
22. 22. Faul F, Erdfelder E, Lang A-G, Buchner A. G* Power 3: A flexible statistical power analysis program for the social, behavioral, and biomedical sciences. Behav Res Methods 2007; 39(2): 175-91. [CrossRef] google scholar
23. 23. Leeanantsaksiri P. Gut-Brain Axis Association to the Brain Function. Inter J Formal Sci: Curr Future Res Trend 2022; 13(1): 167-81. google scholar
24. 24. Nayak RR, Alexander M, Deshpande I, Stapleton-Gray K, Rimal B, Patterson AD, et al. Methotrexate impacts conserved pathways in diverse human gut bacteria leading to decreased host immune activation. Cell Host Microbe 2021; 29(3): 362-77.e11. [CrossRef] google scholar
25. 25. Modun D, Giustarini D, Tsikas D. Nitric oxide-related oxidative stress and redox status in health and disease. Oxidative Medicine and Cellular Longevity, Hindawi; 2014. [CrossRef] google scholar
26. 26. Lanas A. Role of nitric oxide in the gastrointestinal tract. Arthritis Res Ther 2008; 10 Suppl 2(Suppl 2): S4. [CrossRef] google scholar
27. 27. Dijkstra G, van Goor H, Jansen P, Moshage H. Targeting nitric oxide in the gastrointestinal tract. Curr Opin Investig Drugs 2004; 5: 52936. google scholar
28. 28. Cronstein BN, Aune TM. Methotrexate and its mechanisms of action in inflammatory arthritis. Nat Rev Rheumatol 2020; 16(3): 145-54. [CrossRef] google scholar
29. 29. Keri Marshall N. Therapeutic applications of whey protein. Altern Med Rev 2004; 9(2): 136-56. google scholar
30. 30. Walzem R, Dillard C, German JB. Whey components: millennia of evolution create functionalities for mammalian nutrition: What we know and what we may be overlooking. Crit Rev Food Sci Nutr 2002; 42(4): 353-75. [CrossRef] google scholar