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Antioxidant fractions of Khaya grandifoliola C.DC. and Entada africana Guill. et Perr. induce nuclear translocation of Nrf2 in HC-04 cells

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Cell Stress and Chaperones Aims and scope

Abstract

The in vitro antioxidant properties, cytoprotective activity, and ability to induce nuclear translocation of nuclear factor E2-related factor-2 (Nrf-2) of five solvent fractions of the methylene chloride/methanol (1:1 v/v) extract of Khaya grandifoliola (Meliaceae) and Entada africana (Fabaceae) were evaluated. Five antioxidant endpoints were used in the antioxidant activity investigation. The total phenolic content of the fractions was assessed as to the Folin–Ciocalteu method and the profile of interesting fractions analyzed by high-performance liquid chromatography (HPLC). The cytoprotective activity of fractions was determined by H2O2-induced oxidative stress in HC-04 cells by measuring lactate dehydrogenase (LDH) leakage into culture medium. HC-04 cells were used to investigate the ability to induce nuclear translocation of Nrf2. For both plants, the methylene chloride/methanol (90/10; v/v) fraction (F10), methylene chloride/methanol (75/25; v/v) (F25), and the methanolic fraction (F100) were found to have the highest total polyphenol content and exhibited high antioxidant activity strongly correlated with total polyphenol content. The cytoprotective activity of fraction F25 from both plants was comparable to that of quercetin (3.40 ± 0.05 μg/mL), inhibiting LDH leakage with a low half inhibition concentration (IC50) of 4.05 ± 0.03 and 3.8 ± 0.02 μg/mL for K. grandifoliola and E. africana, respectively. Lastly, fraction F25 of K. grandifoliola significantly (P < 0.05) induced nuclear Nrf2 translocation by sixfold, whereas that from E. africana and quercetin was only twofold. The results indicate for the first time that fraction F25 of the studied plants is more antioxidant and cytoprotective and induces nuclear translocation of Nrf2 in a human hepatocyte cell line.

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Abbreviations

CE:

Plant crude extracts

F0:

Methylene chloride fraction

F5:

Methylene chloride/methanol (95/5; v/v) fraction

F10:

Methylene chloride/methanol (90/10; v/v) fraction

F25:

Methylene chloride/methanol (75/25; v/v) fraction

F100:

Methanolic fraction

RC:

Reference compound

PE:

Plant extracts

AA:

Ascorbic acid

DPPH 2:

2-Diphenyl-picryl-hydrazyl (DPPH) free-radical scavenging assay

LP:

Inhibition of rat liver lipid peroxidation assay

HR:

Hydroxyl radical scavenging assay

LDH:

Lactate dehydrogenase

HPLC-UV:

High-performance liquid chromatography-ultraviolet

ROS:

Reactive oxygen species

Nrf2:

Nuclear factor E2-related factor-2

DMEM:

Dulbecco’s modified Eagle’s medium

DMSO:

Dimethyl sulfoxide

MTS:

3-(4, 5-Dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium salt

CAE:

Chlorogenic acid equivalents

AAE:

Ascorbic acid equivalent

HEPES:

N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid

EDTA:

Ethylenediaminetetraacetic acid

DTT:

Dithiotreitol

PVDF:

Polyvinylidene difluoride

SDS-PAGE:

Sodium dodecyl sulfate polyacrylamide gel electropheresis

ANOVA:

Analysis of variance

References

  • Aleksunes LM, Slitt AL, Maher JM, Dieter MZ, Knight TR, Goedken M, Cherrington NJ, Chan JY, Klaassen CD, Manautou JE (2006) Nuclear factor-E2-related factor 2 expression in liver is critical for induction of NAD(P)H:quinone oxidoreductase 1 during cholestasis. Cell Stress Chaperones 11(4):356–363

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Amarowicz R, Pegg RB, Rahimi-Moghaddam P, Barl B, Weil JA (2004) Free-radical scavenging capacity and antioxidant activity of selected plant species from the Canadian prairies. Food Chem 84:551–562

    Article  CAS  Google Scholar 

  • Battu CR, Ethadi SR, Vede PG, Swathi PK, Chandrika K, Rao VA (2011) Evaluation of antioxidant and anti-inflammatory activity of Euphorbia heyneana Spreng. Asian Pac J Trop Biomed 1:191–194

    Article  Google Scholar 

  • Blažeković B, Vladimir-Knežević S, Brantner A, Štefan MB (2010) Evaluation of antioxidant potential of Lavandula x intermedia Emeric ex Loisel ‘Budrovka’: a comparative study with L. angustifolia Mill. Molecules 15:5971–5987

    Article  PubMed  Google Scholar 

  • Bryan HK, Olayanju A, Goldring CE, Park BK (2013) The Nrf2 cell defence pathway: Keap1-dependent and -independent mechanisms of regulation. Biochem Pharmacol 85:705–717

    Article  CAS  PubMed  Google Scholar 

  • Casarett LJ, Doull J, Klaassen CD (2008) Casarett and Doull’s toxicology: the basic science of poisons, 7th edn. McGraw-Hill, New York

    Google Scholar 

  • Chanda S, Dave R (2009) In vitro models for antioxidant activity evaluation and some medicinal plants possessing antioxidant properties: an overview, review. Afr J Microbiol Res 3(13):981–996

    Google Scholar 

  • Copple IM, Goldring CE, Jenkins RE, Chia AJL, Randle LE, Hayes JD, Kitteringham NR, Park BK (2008) The hepatotoxic metabolite of acetaminophen directly activates the Keap1-Nrf2 cell defense system. Hepatology 48:1292–1301

    Article  CAS  PubMed  Google Scholar 

  • Erlank H, Elmann A, Kohen R, Kanner J (2011) Polyphenols activate Nrf2 in astrocytes via H2O2, semiquinones, and quinines. Free Radical Bio Med 51:2319–2327

    Article  CAS  Google Scholar 

  • Esterbauer H (1993) Cytotoxicity and genotoxicity of lipid-oxidation products. Am J Clin Nutr 57:779–785, discussion 785S-786S

    Google Scholar 

  • Germanò MP, Certo G, D’Angelo V, Sanogo R, Malafronte N, De Tommasi N, Rapisarda A (2014) Anti-angiogenic activity of Entada africana root. Nat Prod Res 6:1–6. doi:10.1080/14786419.2014.987773

    Google Scholar 

  • Han WC, Kwun MJ, HaKim K, Choi JY, Oh SR, Ahn KS, Lee JH, Joo M (2013) Ethanol extract of Alismatis rhizoma reduces acute lung inflammation by suppressing NF-kB and activating Nrf2. J Ethnopharmacol http://dx.doi.org/10.1016/j.jep.2013.01.010.i

  • Harbone J (1976) Phytochemical methods. A guide to modern techniques of plants analysis. Chapman and Hall, London, p 150

    Google Scholar 

  • Inoue H, Satoko A, Maeda-Yamamoto M, Nesumi A, Tanaka T, Murakami A (2011) High-dose green tea polyphenols induce nephrotoxicity in dextran sulfate sodium-induced colitis mice by down-regulation of antioxidant enzymes and heat-shock protein expressions. Cell Stress Chaperones 16:653–662. doi:10.1007/s12192-011-0280-8

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Kumar R, Hemalatha S (2011) In-vitro antioxidant activity of alcoholic leaf extract and subfractions of Alangium lamarckii Thwaites. J Chem Pharm Res 3(1):259–267

    Google Scholar 

  • Long LH, Clement MV, Halliwell B (2000) Artifacts in cell culture: rapid generation of hydrogen peroxide on addition of (−)-epigallocatechin, (−)-epigallocatechin gallate, (+)-catechin, and quercetin to commonly used cell culture media. Biochem Bioph Res Co 273:50–53

    Article  CAS  Google Scholar 

  • Mohammadzadeh M, Halabian R, Gharehbaghian A, Amirizadeh N, Jahanian-Najafabadi A, Roushandeh AM (2012) Roudkenar MH (2012) Nrf-2 overexpression in mesenchymal stem cells reduces oxidative stress-induced apoptosis and cytotoxicity. Cell Stress Chaperones 17:553–565. doi:10.1007/s12192-012-0331-9

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Moyo M, Ndhlala AR, Finnie JF, Staden JV (2010) Phenolic composition, antioxidant and acetylcholinesterase inhibitory activities of Sclerocarya birrea and Harpephyllum caffrum (Anacardiaceae) extracts. Food Chem 123:69–76

    Article  CAS  Google Scholar 

  • Njayou FN, Aboudi ECE, Tandjang MK, Tchana AK, Ngadjui BT, Moundipa PF (2013a) Hepatoprotective and antioxidant activities of stem bark extract of Khaya grandifoliola (Welw) C.DC and Entada africana Guill. et Perr. J Nat Prod 6:73–80

    Google Scholar 

  • Njayou FN, Ngoungoure FP, Tchana A, Moundipa PF (2013b) Protective effect of Khaya grandifoliola C.DC. stem bark extract on carbon tetrachloride-induced hepatotoxicity in rats. Int J Ind Med Plants 29(1):1161–1166

    Google Scholar 

  • Njayou FN, Tietcheu BRG, Ngadjui BT, Moundipa PF (2013c) Hepatoprotective activity of three Cameroonian medicinal plants against oxidative damages induced by paracetamol and CCl4 in primary rat hepatocytes. Int J Pharma Photon 104:306–314

    Google Scholar 

  • Odebeyi O, Sofowora (1978) Phytochemical screening of Nigerian medicinal plants II Llyordia 41:234–246

  • Owona BA, Njayou NF, Laufer SA, Schluesener HJ, Moundipa PF (2013a) Entada africana fraction CH2Cl2/MeOH 5% inhibits inducible nitric oxide synthetase and proinflammatory cytokines gene expression induced by lipopolysacharide in microglia. BMC Complement Altern Med 13:254, http://www.biomedcentral.com/ 1472-6882/13/254

    Article  PubMed Central  PubMed  Google Scholar 

  • Owona BA, Njayou NF, Laufer SA, Schluesener HJ, Moundipa PF (2013b) A fraction of stem bark extract of Entada africana suppresses lipopolysaccharide-induced inflammation in RAW264.7cells. J Ethnopharmacol http://dx.doi.org/10.1016/j.jep.2013.06.016.i.

  • Quideau S, Deffieux D, Douat-Casassus C, Pouysegu L (2011) Plant polyphenols: chemical properties, biological activities, and synthesis. Review, natural products. Angew Chem Int Edit 50:586–621

    Article  CAS  Google Scholar 

  • Shin SM, Yang JH, Ki SH (2013) Role of the Nrf2-ARE pathway in liver diseases. Oxid Med Cell Longev 1–9

  • Su XY, Wang ZY, Liu W (2009) In vitro and in vivo antioxidant activity of Pinus Koraiensis seed extract containing phenolic compounds. Food Chem 117:681–686

    Article  CAS  Google Scholar 

  • Subramaniam SR, Ellis EM (2011) Esculetin-induced protection of human hepatoma HepG2 cells against hydrogen peroxide is associated with the Nrf2-dependent induction of the NAD(P)H: quinone oxidoreductase 1 gene. Toxicol Appl Pharm 250:130–136

    Article  CAS  Google Scholar 

  • Tang W, Jiang YF, Ponnusamy M, Diallo M (2014) Role of Nrf2 in chronic liver disease. World J Gastroenterol 20(36):13079–13087. doi:10.3748/wjg.v20.i36.13079

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Tietcheu BRG, Sass G, Njayou NF, Mkounga P, Tiegs G, Moundipa PF (2014) Anti-hepatitis C virus activity of crude extract and fractions of Entada africana in genotype 1b replicon systems. T Am J Chin Med 42(4):853–868

    Article  Google Scholar 

  • Tosi MER, Bocanegra V, Manucha W, Lorenzo AG, Vallés PG (2011) The Nrf2–Keap1 cellular defense pathway and heat shock protein 70 (Hsp70) response. Role in protection against oxidative stress in early neonatal unilateral ureteral obstruction (UUO). Cell Stress Chaperones 16:57–68. doi:10.1007/s12192-010-0221-y

    Article  CAS  Google Scholar 

  • Varshneya C, Kant V, Mehta D (2012) Total phenolic contents and free radical scavenging activities of different extracts of seabuckthorn (Hippophae rhamnoides) pomace without seeds. Int J Food Sci Nutr 63(2):153–9

    Article  CAS  PubMed  Google Scholar 

  • Wong SP, Leong LP, Koh JHW (2006) Antioxidant activities of aqueous extracts of selected plants. Food Chem 99:775–783

    Article  CAS  Google Scholar 

  • Xu D, Zhou C (2013) Antioxidative effects of berberine pre-treatment on hydrogen peroxide-induced PC12 cell toxicity. Neural Regen Res 5(18):1391–1395

    Google Scholar 

  • Zhao CR, Gao ZH, Qu XJ (2010) Nrf2–ARE signaling pathway and natural products for cancer chemoprevention. Cancer Epidemiol 34:523–533

    Article  CAS  PubMed  Google Scholar 

  • Zhu R, Wang Y, Zhang L, Guo Q (2012) Oxidative stress and liver disease. Hepatol Res 42(8):741–749

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

Grants from the International Foundation for Sciences and the Organisation for the Prohibition of Chemical Weapons (F/4223-2) and National Institute of Health (DK069557) awarded to Drs. Njayou and Manautou, respectively, supported this work. Dr. Njayou thanks the Fulbright Program for the award which allowed him to carry out experiments on HC-04 cells in the laboratory of Dr. Manautou at the School of Pharmacy, Connecticut, USA.

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Authors and Affiliations

  1. Laboratory of Pharmacology and Toxicology, Department of Biochemistry, Faculty of Science, University of Yaoundé 1, P.O. Box 812, Yaounde, Cameroon

    Frédéric Nico Njayou, Atsama Marie Amougou, Romeo Fouemene Tsayem, Jacqueline Njikam Manjia & Paul Fewou Moundipa

  2. Department of Food Science, University of Wisconsin-Madison, Madison, WI, 53706, USA

    Bolling Bradley

  3. Department of Pharmaceutical Sciences, University of Connecticut, Storrs, Connecticut, 06269, USA

    Frédéric Nico Njayou, Swetha Rudraiah & José Enrique Manautou

Authors
  1. Frédéric Nico Njayou
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  2. Atsama Marie Amougou
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  3. Romeo Fouemene Tsayem
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  4. Jacqueline Njikam Manjia
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  6. Bolling Bradley
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  7. José Enrique Manautou
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  8. Paul Fewou Moundipa
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Correspondence to Paul Fewou Moundipa.

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Njayou, F.N., Amougou, A.M., Fouemene Tsayem, R. et al. Antioxidant fractions of Khaya grandifoliola C.DC. and Entada africana Guill. et Perr. induce nuclear translocation of Nrf2 in HC-04 cells. Cell Stress and Chaperones 20, 991–1000 (2015). https://doi.org/10.1007/s12192-015-0628-6

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