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Planta Med 2015; 81(05): 363-372
DOI: 10.1055/s-0035-1545839
Biological and Pharmacological Activity
Original Papers
Georg Thieme Verlag KG Stuttgart · New York

In Vitro Effects of Panax ginseng in Aristolochic Acid-Mediated Renal Tubulotoxicity: Apoptosis versus Regeneration

Valérian Bunel
1   Laboratory of Experimental Nephrology, Faculty of Medicine, Université Libre de Bruxelles (ULB), Brussels, Belgium
2   Laboratory of Pharmacognosy, Bromatology and Human Nutrition, Faculty of Pharmacy, Université Libre de Bruxelles (ULB), Brussels, Belgium
,
Marie-Hélène Antoine
1   Laboratory of Experimental Nephrology, Faculty of Medicine, Université Libre de Bruxelles (ULB), Brussels, Belgium
,
Joëlle Nortier
1   Laboratory of Experimental Nephrology, Faculty of Medicine, Université Libre de Bruxelles (ULB), Brussels, Belgium
,
Pierre Duez
2   Laboratory of Pharmacognosy, Bromatology and Human Nutrition, Faculty of Pharmacy, Université Libre de Bruxelles (ULB), Brussels, Belgium
3   Department of Therapeutical Chemistry and Pharmacognosy, Université de Mons (UMONS), Mons, Belgium
,
Caroline Stévigny
2   Laboratory of Pharmacognosy, Bromatology and Human Nutrition, Faculty of Pharmacy, Université Libre de Bruxelles (ULB), Brussels, Belgium
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Publikationsverlauf

received 12. März 2014
revised 19. Januar 2015

accepted 29. Januar 2015

Publikationsdatum:
23. März 2015 (online)

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Abstract

This in vitro study aimed to determine the effects of a Panax ginseng extract on aristolochic acid-mediated toxicity in HK-2 cells. A methanolic extract of ginseng (50 µg/mL) was able to reduce cell survival after treatment with 50 µM aristolochic acid for 24, 48, and 72 h, as evidenced by a resazurin reduction assay. This result was confirmed by a flow cytometric evaluation of apoptosis using annexin V-PI staining, and indicated higher apoptosis rates in cells treated with aristolochic acid and P. ginseng extract compared with aristolochic acid alone. However, P. ginseng extract by itself (5 and 50 µg/mL) increased the Ki-67 index, indicating an enhancement in cellular proliferation. Cell cycle analysis excluded a P. ginseng extract-mediated induction of G2/M cell cycle arrest such as the one typically observed with aristolochic acid. Finally, β-catenin acquisition was found to be accelerated when cells were treated with both doses of ginseng, suggesting that the epithelial phenotype of renal proximal tubular epithelial cells was maintained. Also, ginseng treatment (5 and 50 µg/mL) reduced the oxidative stress activity induced by aristolochic acid after 24 and 48 h. These results indicate that the ginseng extract has a protective activity towards the generation of cytotoxic reactive oxygen species induced by aristolochic acid. However, the ginseng-mediated alleviation of oxidative stress did not correlate with a decrease but rather with an increase in aristolochic acid-induced apoptosis and death. This deleterious herb-herb interaction could worsen aristolochic acid tubulotoxicity and reinforce the severity and duration of the injury. Nevertheless, increased cellular proliferation and migration, along with the improvement in the epithelial phenotype maintenance, indicate that ginseng could be useful for improving tubular regeneration and the recovery following drug-induced kidney injury. Such dual activities of ginseng certainly warrant further in vivo studies.

Key words

apoptosis - aristolochic acids - HK-2 cells - Panax ginseng - Araliaceae - proliferation - regeneration
 

  • References

  • 1 Price PM, Safirstein RL, Megyesi J. The cell cycle and acute kidney injury. Kidney Int 2009; 76: 604-613
    CrossrefPubMedGoogle Scholar
  • 2 Megyesi J, Andrade L, Vieira jr. JM, Safirstein RL, Price PM. Coordination of the cell cycle is an important determinant of the syndrome of acute renal failure. Am J Physiol Renal Physiol 2002; 283: F810-F816
    CrossrefPubMedGoogle Scholar
  • 3 Wynn TA. Fibrosis under arrest. Nat Med 2010; 16: 523-525
    CrossrefPubMedGoogle Scholar
  • 4 Vanherweghem JL, Depierreux M, Tielemans C, Abramowicz D, Dratwa M, Jadoul M, Richard C, Vandervelde D, Verbeelen D, Vanhaelen-Fastre R, Vanhaelen M. Rapidly progressive interstitial renal fibrosis in young women: association with slimming regimen including Chinese herbs. Lancet 1993; 341: 387-391
    CrossrefPubMedGoogle Scholar
  • 5 Debelle FD, Vanherweghem JL, Nortier JL. Aristolochic acid nephropathy: a worldwide problem. Kidney Int 2008; 74: 158-169
    CrossrefPubMedGoogle Scholar
  • 6 Yang L, Su T, Li XM, Wang X, Cai SQ, Meng LQ, Zou WZ, Wang HY. Aristolochic acid nephropathy: variation in presentation and prognosis. Nephrol Dial Transplant 2012; 27: 292-298
    CrossrefPubMedGoogle Scholar
  • 7 Daoudi A, Benboubker H, Bousta D, Aarab L. Screening of fourteen Moroccan medicinal plants for immunomodulating activities. Moroccan J Biol 2008; 1: 24-30
    PubMedGoogle Scholar
  • 8 Heinrich M, Chan J, Wanke S, Neinhuis C, Simmonds MS. Local uses of Aristolochia species and content of nephrotoxic aristolochic acid 1 and 2 – a global assessment based on bibliographic sources. J Ethnopharmacol 2009; 125: 108-144
    CrossrefPubMedGoogle Scholar
  • 9 Rueda DC, Zaugg J, Quitschau M, Reich E, Hering S, Hamburger M. Discovery of GABAA receptor modulator aristolactone in a commercial sample of the Chinese herbal drug “Chaihu” (Bupleurum chinense roots) unravels adulteration by nephrotoxic Aristolochia manshuriensis roots. Planta Med 2012; 78: 207-210
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 10 Bensky D, Gamble A, Kaptchuk T, Bensky L. Chinese herbal medicine: Materia Medica, 4th edition. Seattle: Eastland Press; 1986
    Google Scholar
  • 11 Nocerino E, Amato M, Izzo AA. The aphrodisiac and adaptogenic properties of ginseng. Fitoterapia 2000; 71 (Suppl. 01) S1-S5
    CrossrefPubMedGoogle Scholar
  • 12 Baek SH, Piao XL, Lee UJ, Kim HY, Park JH. Reduction of cisplatin-induced nephrotoxicity by ginsenosides isolated from processed ginseng in cultured renal tubular cells. Biol Pharm Bull 2006; 29: 2051-2055
    CrossrefPubMedGoogle Scholar
  • 13 Xie XS, Yang M, Liu HC, Zuo C, Li Z, Deng Y, Fan JM. Influence of ginsenoside Rg1, a panaxatriol saponin from Panax notoginseng, on renal fibrosis in rats with unilateral ureteral obstruction. J Zhejiang Univ Sci B 2008; 9: 885-894
    CrossrefPubMedGoogle Scholar
  • 14 Xie XS, Yang M, Liu HC, Zuo C, Li HJ, Fan JM. Ginsenoside Rg1, a major active component isolated from Panax notoginseng, restrains tubular epithelial to myofibroblast transition in vitro . J Ethnopharmacol 2009; 122: 35-41
    CrossrefPubMedGoogle Scholar
  • 15 Zhang MH, Fan JM, Xie XS, Chen YP, Zhen R, Li J, Cheng Y, Wen J. Ginsenoside-Rg1 protects podocytes from complement mediated injury. J Ethnopharmacol 2011; 137: 99-107
    CrossrefPubMedGoogle Scholar
  • 16 Varjas T, Nowrasteh G, Budan F, Nadasi E, Horvath G, Makai S, Gracza T, Cseh J, Ember I. Chemopreventive effect of Panax ginseng . Phytother Res 2009; 23: 1399-1403
    CrossrefPubMedGoogle Scholar
  • 17 Kang KS, Kim HY, Yamabe N, Nagai R, Yokozawa T. Protective effect of sun ginseng against diabetic renal damage. Biol Pharm Bull 2006; 29: 1678-1684
    CrossrefPubMedGoogle Scholar
  • 18 Kim DH. Chemical diversity of Panax ginseng, Panax quinquefolium, and Panax notoginseng . J Ginseng Res 2012; 36: 1-15
    CrossrefPubMedGoogle Scholar
  • 19 Ryan MJ, Johnson G, Kirk J, Fuerstenberg SM, Zager RA, Torok-Storb B. HK-2: an immortalized proximal tubule epithelial cell line from normal adult human kidney. Kidney Int 1994; 45: 48-57
    CrossrefPubMedGoogle Scholar
  • 20 Yang L, Besschetnova TY, Brooks CR, Shah JV, Bonventre JV. Epithelial cell cycle arrest in G2/M mediates kidney fibrosis after injury. Nat Med 2010; 16: 535-543 1 p following 143
    CrossrefPubMedGoogle Scholar
  • 21 Li J, Zhang L, Jiang Z, Shu B, Li F, Bao Q. Toxicities of aristolochic acid I and aristololactam I in cultured renal epithelial cells. Toxicol In Vitro 2010; 24: 1092-1097
    CrossrefPubMedGoogle Scholar
  • 22 Hong SJ, Wan JB, Zhang Y, Hu G, Lin HC, Seto SW, Kwan YW, Lin ZX, Wang YT, Lee SM. Angiogenic effect of saponin extract from Panax notoginseng on HUVECs in vitro and zebrafish in vivo . Phytother Res 2009; 23: 677-686
    CrossrefPubMedGoogle Scholar
  • 23 Wang P, Wei X, Zhou Y, Wang YP, Yang K, Zhang FJ, Jiang R. Effect of ginsenoside Rg1 on proliferation and differentiation of human dental pulp cells in vitro . Aust Dent J 2012; 57: 157-165
    CrossrefPubMedGoogle Scholar
  • 24 Choi S. Epidermis proliferative effect of the Panax ginseng ginsenoside Rb2. Arch Pharmacal Res 2002; 25: 71-76
    CrossrefPubMedGoogle Scholar
  • 25 Lin T, Liu Y, Shi M, Liu X, Li L, Zhao G. Promotive effect of ginsenoside Rd on proliferation of neural stem cells in vivo and in vitro . J Ethnopharmacol 2012; 142: 754-761
    CrossrefPubMedGoogle Scholar
  • 26 Sakurai M. Perspective: Herbal dangers. Nature 2011; 480: S97
    CrossrefPubMedGoogle Scholar
  • 27 Jia L, Zhao Y. Current evaluation of the millennium phytomedicine – ginseng (I): etymology, pharmacognosy, phytochemistry, market and regulations. Curr Med Chem 2009; 16: 2475-2484
    CrossrefPubMedGoogle Scholar
  • 28 Kitts D, Hu C. Efficacy and safety of ginseng. Public Health Nutr 2000; 3: 473-485
    PubMedGoogle Scholar
  • 29 Elmer GW, Lafferty WE, Tyree PT, Lind BK. Potential interactions between complementary/alternative products and conventional medicines in a Medicare population. Ann Pharmacother 2007; 41: 1617-1624
    CrossrefPubMedGoogle Scholar
  • 30 Dickman KG, Sweet DH, Bonala R, Ray T, Wu A. Physiological and molecular characterization of aristolochic acid transport by the kidney. J Pharmacol Exp Ther 2011; 338: 588-597
    CrossrefPubMedGoogle Scholar
  • 31 Martinek V, Kubickova B, Arlt VM, Frei E, Schmeiser HH, Hudecek J, Stiborova M. Comparison of activation of aristolochic acid I and II with NADPH : quinone oxidoreductase, sulphotransferases and N-acetyltranferases. Neuro Endocrinol Lett 2011; 32 (Suppl. 01) S57-S70
    PubMedGoogle Scholar
  • 32 Jerabek P, Martinek V, Stiborova M. Theoretical investigation of differences in nitroreduction of aristolochic acid I by cytochromes P450 1A1, 1A2 and 1B1. Neuro Endocrinol Lett 2012; 33 (Suppl. 03) S25-S32
    PubMedGoogle Scholar
  • 33 Havasi A, Borkan SC. Apoptosis and acute kidney injury. Kidney Int 2011; 80: 29-40
    CrossrefPubMedGoogle Scholar
  • 34 Urruticoechea A, Smith IE, Dowsett M. Proliferation marker Ki-67 in early breast cancer. J Clin Oncol 2005; 23: 7212-7220
    CrossrefPubMedGoogle Scholar
  • 35 Mizui M, Isaka Y, Takabatake Y, Mizuno S, Nakamura T, Ito T, Imai E, Hori M. Electroporation-mediated HGF gene transfer ameliorated cyclosporine nephrotoxicity. Kidney Int 2004; 65: 2041-2053
    CrossrefPubMedGoogle Scholar
  • 36 Docherty NG, OʼSullivan OE, Healy DA, Murphy M, OʼNeill A J, Fitzpatrick JM, Watson RW. TGF-beta1-induced EMT can occur independently of its proapoptotic effects and is aided by EGF receptor activation. Am J Physiol Renal Physiol 2006; 290: F1202-F1212
    PubMedGoogle Scholar
  • 37 Zeisberg M, Neilson EG. Biomarkers for epithelial-mesenchymal transitions. J Clin Invest 2009; 119: 1429-1437
    CrossrefPubMedGoogle Scholar
  • 38 Carew RM, Wang B, Kantharidis P. The role of EMT in renal fibrosis. Cell Tissue Res 2011; 347: 103-116
    PubMedGoogle Scholar
  • 39 Saotome K, Morita H, Umeda M. Cytotoxicity test with simplified crystal violet staining method using microtitre plates and its application to injection drugs. Toxicol In Vitro 1989; 3: 317-321
    CrossrefPubMedGoogle Scholar
  • 40 Mosmann T. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J Immunol Methods 1983; 65: 55-63
    CrossrefPubMedGoogle Scholar
  • 41 Bunel V, Ouedraogo M, Nguyen AT, Stévigny C, Duez P. Methods applied to the in vitro primary toxicology testing of natural products: state of the art, strengths, and limits. Planta Med 2014; 80: 1210-1226
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 42 Council of Europe. European Pharmacopoeia, 8th edition. Strasbourg: Tec & Doc; 2013
    Google Scholar
  • 43 Geback T, Schulz MM, Koumoutsakos P, Detmar M. TScratch: a novel and simple software tool for automated analysis of monolayer wound healing assays. Biotechniques 2009; 46: 265-274
    PubMedGoogle Scholar
  • 44 Valcu M, Valcu CM. Data transformation practices in biomedical sciences. Nat Methods 2011; 8: 104-105
    CrossrefPubMedGoogle Scholar