Elsevier

Clinical Nutrition

Volume 23, Issue 2, April 2004, Pages 183-193
Clinical Nutrition

Original article
l-Ergothioneine modulates oxidative damage in the kidney and liver of rats in vivo: studies upon the profile of polyunsaturated fatty acids

https://doi.org/10.1016/S0261-5614(03)00108-0Get rights and content

Abstract

Background & aims: l-Ergothioneine is a fungal metabolite exhibiting antioxidant functions in cells. The aim of this study was to assess the effect of oral administration of l-Ergothioneine on the oxidative damage in vivo caused by the Fenton reagent ferric-nitrilotriacetate.

Methods: Rats were supplemented with l-Ergo prior to the administration of acute dose of ferric-nitrilotriacetate. Kidney and liver levels of l-ergothioneine, glutathione, α-tocopherol, polyunsaturated fatty acids and conjugated dienes were assessed.

Results: Oral administration of 70 mg l-Ergo/kg body weight of rats for 7 days prior to the injection of ferric-nitrilotriacetate protected the fatty acids against oxidation, with notable protections directed to: 20:5 (eicosapentaenoic acid) (23%), 22:6 (docosahexaenoinic acid) (30%), 20:3 n6 (eicosatrienoic acid) (22%), 20:4 (arachidonic acid) (25%), 18:2 linoleic acid (25%) and 18:1 oleic acid (14%) in the kidney. The protection of 20:5, 20:3 n6 and 18:1 in the liver by 32%, 20% and 11%, respectively, were statistically significant. l-Ergothioneine significantly reduced kidney and liver levels of conjugated dienes and conserved the concentrations of α-tocopherol and glutathione in the kidney and liver in the ferric-nitrilotriacetate/l-ergothioneine treated rats.

Conclusion: Supplementation with l-ergothioneine not only protects the organs against the lipid peroxidation but conserves the consumption of endogenous glutathione and α-tocopherol. However consumption of mushrooms may have better promise as dietary sources of l-ergothioneine to humans.

Introduction

The benefits of antioxidants continue to be advocated in the context of health and disease management.1., 2., 3., 4., 5., 6. Although antioxidant actions may either involve direct inhibition of the generation or scavenging of reactive oxygen species, direct reproduction of efficacy in vivo are often hard to achieve, this is driven in part by the lack of a reliable in vivo model with which to assess oxidative stress. Thus challenges continue to emerge from difficulties associated with methods used in evaluating antioxidant actions in vivo.7., 8., 9., 10., 11. In order to establish bio-efficacy of antioxidants, ‘‘markers’’ of baseline oxidative damage in vivo need to be examined as a function of their modulation by dietary antioxidants.2., 5. To this end, Deiana et al.11 developed a Fenton chemistry model, using the iron complex of the chelating agent nitrilotriacetic acid (Fe-NTA), based on the profile of polyunsaturated fatty acids for the assessment of antioxidant actions in vivo. Intraperitoneal injection of ferric nitrilotriacetate (Fe-NTA) induces renal proximal tubular damage associated with oxidative damage that eventually leads to a high incidence of renal cell carcinoma in rodents after repeated administration.12., 13., 14., 15. In the kidney, Fe-NTA can be filtered through the glomeruli into the lumen of the renal proximal tubule where Fenton chemistry could occur mediating oxidative damage. Intraperitoneal administration of sub-lethal dose of Fe-NTA to rats induces a time-dependent reduction in the levels of polyunsaturated fatty acids (PUFA), together with an increase of conjugated dienes value and a decrease of cellular antioxidants α-tocopherol and glutathione.11 Fig. 1 presents the suggested scheme of the model which argues that the peak of the levels of the PUFAs at 3 h following the injection of acute dose of Fe-NTA may serve as a sampling point if this model is to be used for the assessment of antioxidant actions based on PUFA profiles.11

l-Ergo is a naturally occurring 2-thio-imidazole amino acid which in aqueous solution exists predominantly in the thione form (Fig. 2). Thus unlike glutathione (GSH), at physiologic pH l-Ergo does not auto-oxidize and is therefore very stable in aqueous solution. l-Ergo is extremely hydrophilic with a solubility limit of 0.9 M at room temperature. It is an excellent chelator of divalent metals especially copper and remarkably stable to strong alkali, properties which further differentiate it from other biological thiols. l-Ergo is formed in some bacteria and fungi but not in animals.16., 17., 18., 19. In humans, l-Ergo is only absorbed through consumption of plant diet, primarily by consumption of edible mushrooms and meat where upon the grazing animals would have absorbed l-Ergo from consumption of grazing grasses. The concentrations of l-Ergo in almost every species investigated are in the micromolar range. For man, blood concentrations have been estimated to be in the range 1–4 mg/100 ml blood which approximates to concentrations of 46–183 μM20 while bovine and porcine ocular tissue concentrations are reported to be 2.96+0.2  and 8.69+1.57 μmol/mg tissue, respectively.19 Kumosani21 reported the blood levels of l-Ergo in males to be 2.8 mg/100 ml blood at the age of 51. Concentrations of l-Ergo was reported to be age dependent with levels of 1.5–2.0 mg/100 ml blood at 1–10 years of age, reaching a maxima of 3.7 mg/100 ml blood at 11–18 years of age and on average, 2.3–3.0 mg/100 ml blood at 19–50 years of age.21 These data are consistent with the earlier work of Hunter in 1951 as discussed by Mann and Leone22 from which blood concentrations in the range 1.8–1.95 mg/100 ml blood was reported. l-Ergo is taken up by erythrocytes.23., 24. The biological role for l-Ergo is gradually being defined, presently l-Ergo is radioprotective,25 scavenges singlet oxygen, hydroxyl radicals,26., 27., 28. hypochlorous acid (HOCl), peroxyl radicals,28., 29. inhibits peroxynitrite dependent nitration of amino acids tyrosine and DNA and confers cellular homeostasis in neuronal cells,30 protects retinal neurones against N-methyl-d-aspartate (NMDA) excitotoxicity in vivo,31 protects against diabetic embyropathy in pregnant rats,32 induces S-nitrosoglutathione (GSNO) decomposition.33 Kawano et al.34 have shown that l-Ergo inhibits hepatic injury and significantly decreases the levels of lipid peroxide-induced by phenobarbital or ethionine. The high concentrations of l-Ergo in boar serum22 is known to prevent Cu2+ inhibition of sperm motility and fructolysis, thus interactions of l-Ergo with metalloenzymes35 is an important metabolic function. l-Ergo inhibits the UVA-stimulated ATM (the gene mutated in ataxia telangiectasia) kinase activity,36 suggesting possible protection by l-Ergo of the ATM/apoptosis induced by UVA. In this paper, it was of interest to study the ability of l-Ergo to modulate the oxidative damage caused by the ferric-nitrilotriacetic acid.

Section snippets

Chemicals

All solvents used were HPLC grade (Merck, Darmstadt, Germany). Ferric nitrate nonahydrate, nitrilotriacetic acid disodium salt and fatty acids standards were purchased from Sigma Chemical, St. Louis, MO, USA. Desferal (deferoxamine methanesulfonate) was purchased from CIBA-Geigy, Basel, Switzerland. α-Tocopherol was purchased from Fluka AG, Switzerland. All other reagents and chemicals were of the highest available purity. l-Ergo was obtained from OXIS International Inc. Portland, Oregon, USA.

Preparation of Fe-NTA

Results

A number of methods have been discussed for the extraction of l-Ergo and its measurement by HPLC.20., 45. This paper reports a simultaneous extraction of l-Ergo and total lipids using a slight modification of the Folch et al. procedure,38 without any effect on the recovery of lipids. Fig. 3 shows the peak for l-Ergo measured in the tissue homogenates, compared to the standard compound. The quali-quantitative analyses of the compound was performed by HPLC. In the methanol/water phase solution,

Discussion

Early detection of oxidative stress-dependent damage, use of antioxidants and the development of effective preventive strategies are critical in combating ROS-mediated diseases and in management of health. Indeed kidney toxicity comprises a significant portion of admissions to hospitals and this has associated high healthcare cost. Kidney function can be affected by numerous factors including oxidative stress. The Fenton chemistry Fe-NTA model is a unique model for studying the ability of

Acknowledgements

We wish to thank Giacomo Satta for his precious technical assistance in animal handling. This work was supported by: Regione Autonoma della Sardegna-Programma Comunitario Interreg (PIC-INTERREG II). OXIS International is thanked for provision of ergothioneine.

References (52)

  • D. Akanmu et al.

    The antioxidant action of l-Ergo

    Arch Biochem Biophys

    (1991)
  • O.I. Aruoma et al.

    The antioxidant action of l-Ergo. Assessment of its ability to scavenge peroxynitrite

    Biochem Biophys Res Commun

    (1997)
  • O.I. Aruoma et al.

    Protection against oxidative damage and cell death by natural antioxidant l-Ergo

    Food Chem Toxicol

    (1999)
  • J.A. Moncaster et al.

    l-Ergo treatment protects neurons against N-methyl-d-aspartate excitotoxicityin an in vivo retinal model

    Neurosci Lett

    (2002)
  • M.V. Guijarro et al.

    Effects of l-Ergo on diabetic embropathy in pregnant rats

    Food Chem Toxicol

    (2002)
  • Y. Zhang et al.

    Requirement of ATM in UVA-induced signaling and apoptosis

    J Biol Chem

    (2002)
  • J. Folch et al.

    A simple method for the isolation and purification of total lipid from animal tissues

    J Biol Chem

    (1957)
  • S. Banni et al.

    Characterization of conjugated diene fatty acids in milk, dairy products and lamb tissues

    J Nutr Biochem

    (1996)
  • F.P. Corongiu et al.

    Detection of conjugated dienes by second derivative ultraviolet spectrophotometry

    Methods Enzymol

    (1994)
  • T.A. Zainal et al.

    Localization of 4-hydroxy-2-nonenal modified proteins in kidney following iron overload

    Free Rad Biol Med

    (1999)
  • C.P. Diggle

    In vitro studies on the relationship between polyunsaturated fatty acids, cancertumour or tissue specific effects?

    Prog Lipid Res

    (2002)
  • O.I. Aruoma

    Free radicals, oxidative stress and antioxidants in health and disease

    J Am Oil Chem Soc

    (1998)
  • B. Halliwell

    Anti oxidants in human health and disease

    Annu Rev Nutr

    (1996)
  • Bray T, Schoene N. Models and Methods in Cell Signalling and Gene Expression. Applications to oxidative stress...
  • Aruoma OI, Halliwell B. Molecular Biology of Free Radicals in Human Diseases. London: OICA International,...
  • H.R. Griffith et al.

    Biomarkers

    Mol Aspect Med

    (2002)
  • Cited by (76)

    • L-ergothioneine and its combination with metformin attenuates renal dysfunction in type-2 diabetic rat model by activating Nrf2 antioxidant pathway

      2021, Biomedicine and Pharmacotherapy
      Citation Excerpt :

      L-egt (as shown in Fig. 1) possesses antioxidant and anti-inflammatory properties, while its accumulation at the sites of tissue injury has been hypothesized as an adaptive mechanism of protecting tissues at risk of damage and regenerating injured tissues [21,42]. Also, L-egt has been reported to activate the Nrf2 (nuclear factor erythroid-2 related factor-2) antioxidant signaling pathway to protect against cellular injury, enhance glutathione level and reduce oxidative damage in the kidney as well as activates sirt1 and 6 to protect against high glucose-induced cell senescence [12,13,22]. Furthermore, coadministration of L- egt with existing therapy (such as melatonin and hispidin) significantly increases treatment benefits both in-vitro and in-vivo [52,53], suggesting that L-egt may improve the efficacy of available treatment options.

    • The potential therapeutic effects of ergothioneine in pre-eclampsia

      2018, Free Radical Biology and Medicine
      Citation Excerpt :

      It is a dietary water-soluble amino acid, derived from histidine, which is synthesised mainly by non-yeast fungi (especially basidiomycetes), actinobacteria [24], methylotrophs [29] and cyanobacteria [30]. Numerous physiological roles of ERG have been proposed including cation chelation (Cu2+ in particular) [31,32], immune regulation, regulation of gene expression and most widely as a direct anti-oxidant due to its preferential concentration in high O2 stress organs: liver, kidneys, erythrocytes, eye lens and seminal fluid [24,26,32–53]. Despite ERG's high concentration and ubiquitous presence, all mammalian ERG is derived from dietary sources with typical whole blood concentrations of 66 ± 2.2 µmol/L [54] (Table 1).

    View all citing articles on Scopus
    View full text