Advertisement

Apoptosis

, Volume 11, Issue 10, pp 1801–1811 | Cite as

PPARγ is a key target of butyrate-induced caspase-3 activation in the colorectal cancer cell line Caco-2

  • Markus Schwab
  • Veerle Reynders
  • Sandra Ulrich
  • Nadine Zahn
  • Jürgen Stein
  • Oliver SchröderEmail author
Report

Abstract

Background: Butyrate, a potent histone deacetylase inhibitor, belongs to a promising new class of antineoplastic agents with the capacity to induce apoptosis of cancer cells. However, the underlying mechanisms of action have yet not been elucidated. Aim: To further investigate the molecular events involved in butyrate-induced caspase-3 activation in Caco-2 wild-type, empty-vector and dominant-negative PPARγ mutant cells along the signalling pathway. In this context, the involvement and up-regulation of PPARγ was examined. Results: Stimulation of cells with butyrate resulted in increased expression of PPARγ mRNA, protein, and activity as well as phospho-p38 MAPK protein expression and caspase-3 activity. Arsenite, a direct stimulator of p38 MAPK, also led to an increased PPARγ expression, thereby mimicking the effects of butyrate. In contrast, butyrate-mediated up-regulation of PPARγ was counteracted by co-incubation with the p38 MAPK inhibitor SB203580. Treatment of cells with butyrate resulted in both increased caspase-8 and -9 activity and reduced expression of XIAP and survivin. However, butyrate-mediated effects on these apoptosis-regulatory proteins leading to caspase-3 activation were almost completely abolished in Caco-2 dominant-negative PPARγ mutant cells. Conclusions: Our data clearly unveil PPARγ as a key target in the butyrate-induced signalling cascade leading to apoptosis via caspase-3 in Caco-2 cells.

Keywords

Apoptosis Butyrate Caspase-3 p38 MAPK PPARγ 

Abbreviations

DFF

DNA Fragmentation Factor

DMEM

Dulbecco’s modified Eagle’s medium

DMSO

Dimethyl sulfoxide

ECACC

European Collection of cell cultures

EGF

Epithelial growth factor

ERK

Extracellular signal-regulated kinase

FCS

Foetal calf serum

FLIP

FLICE-inhibitory protein

HDAC

Histone deacetylase

IAP

Inhibitor of apoptosis protein

JNK

cJun N-terminal kinase

LDH

Lactate dehydrogenase

MAPK

Mitogen-activated protein kinase

PARP

Poly(ADP-ribose) Polymerase

PBS

Phosphate buffered saline

PKC-δ

Protein kinase C-δ

PPARγ

Peroxisome proliferator-activated receptor γ

SCFA

Short chain fatty acid

STAT3

Signal transducer and activator of transcription-3

TBS-T

Tris-buffered saline containing 0.05% Tween 20

TNF

Tumor necrosis factor

XIAP

X linked inhibitor of apoptosis

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Nakaji S, Ishiguro S, Iwane S, et al (2004) The prevention of colon carcinogenesis in rats by dietary cellulose is greater than the promotive effect of dietary lard as assessed by repeated endoscopic observation. J Nutr 134:935–939PubMedGoogle Scholar
  2. 2.
    Wachtershauser A, Stein J (2000) Rationale for the luminal provision of butyrate in intestinal diseases. Eur J Nutr 39:164–171PubMedCrossRefGoogle Scholar
  3. 3.
    Klurfeld DM (1999) Nutritional regulation of gastrointestinal growth. Front Biosci 4:D299–302PubMedGoogle Scholar
  4. 4.
    Gaschott T, Steinhilber D, Milovic V, Stein J (2001) Tributyrin, a stable and rapidly absorbed prodrug of butyric acid, enhances antiproliferative effects of dihydroxycholecalciferol in human colon cancer cells. J Nutr 131:1839–1843PubMedGoogle Scholar
  5. 5.
    Wachtershauser A, Stein J (2001) Butyrate-induced differentiation of Caco-2 cells occurs independently from p27. Biochem Biophys Res Commun 281:295–299PubMedCrossRefGoogle Scholar
  6. 6.
    Galfi P, Neogrady S, Kutas F, Veresegyhazy T (1985) Influence of sodium butyrate on HeLa cell morphology and proliferation. Life Sci 37:2257–2268PubMedCrossRefGoogle Scholar
  7. 7.
    Cherbut C (1995) Role of gastrointestinal motility in the delay of absorption by dietary fibre. Eur J Clin Nutr 49(Suppl 3): S74–80PubMedGoogle Scholar
  8. 8.
    Toscani A, Soprano DR, Soprano KJ (1988) Molecular analysis of sodium butyrate-induced growth arrest. Oncogene Res 3:223–238PubMedGoogle Scholar
  9. 9.
    Rozental R, Faharani R, Yu Y, Johnson JM, Chan SO, Chiu FC (2004) Sodium butyrate induces apoptosis in MSN neuroblastoma cells in a calcium independent pathway. Neurochem Res 29:2125–2134PubMedCrossRefGoogle Scholar
  10. 10.
    Velazquez OC, Rombeau JL (1997) Butyrate. Potential role in colon cancer prevention and treatment. Adv Exp Med Biol 427:169–181PubMedGoogle Scholar
  11. 11.
    Dashwood RH, Myzak MC, Ho E (2006) Dietary HDAC inhibitors: time to rethink weak ligands in cancer chemoprevention? Carcinogenesis 27:344–349PubMedCrossRefGoogle Scholar
  12. 12.
    Ruemmele FM, Schwartz S, Seidman EG, Dionne S, Levy E, Lentze MJ (2003) Butyrate induced Caco-2 cell apoptosis is mediated via the mitochondrial pathway. Gut 52:94–100PubMedCrossRefGoogle Scholar
  13. 13.
    Shao Y, Gao Z, Marks PA, Jiang X (2004) Apoptotic and autophagic cell death induced by histone deacetylase inhibitors. Proc Natl Acad Sci USA 101:18030–18035PubMedCrossRefGoogle Scholar
  14. 14.
    Li CJ, Elsasser TH (2005) Butyrate-induced apoptosis and cell cycle arrest in bovine kidney epithelial cells: involvement of caspase and proteasome pathways. J Anim Sci 83:89–97PubMedGoogle Scholar
  15. 15.
    McMillan L, Butcher SK, Pongracz J, Lord JM (2003) Opposing effects of butyrate and bile acids on apoptosis of human colon adenoma cells: differential activation of PKC and MAP kinases. Br J Cancer 88:748–753PubMedCrossRefGoogle Scholar
  16. 16.
    Wesierska-Gadek J, Gueorguieva M, Wojciechowski J, Tudzarova-Trajkovska S (2004) In vivo activated caspase-3 cleaves PARP-1 in rat liver after administration of the hepatocarcinogen N-nitrosomorpholine (NNM) generating the 85 kDa fragment. J Cell Biochem 93:774–787PubMedCrossRefGoogle Scholar
  17. 17.
    Liu X, Zou H, Slaughter C, Wang X (1997) DFF, a heterodimeric protein that functions downstream of caspase-3 to trigger DNA fragmentation during apoptosis. Cell 89:175–184PubMedCrossRefGoogle Scholar
  18. 18.
    Kumar S (1995) ICE-like proteases in apoptosis. Trends Biochem Sci 20:198–202PubMedCrossRefGoogle Scholar
  19. 19.
    Nagata S (1997) Apoptosis by death factor. Cell 88:355–365PubMedCrossRefGoogle Scholar
  20. 20.
    Cory S, Adams JM (2002) The Bcl2 family: regulators of the cellular life-or-death switch. Nat Rev Cancer 2:647–656PubMedCrossRefGoogle Scholar
  21. 21.
    Zhang XD, Gillespie SK, Borrow JM, Hersey P (2003) The histone deacetylase inhibitor suberic bishydroxamate: a potential sensitizer of melanoma to TNF-related apoptosis-inducing ligand (TRAIL) induced apoptosis. Biochem Pharmacol 66:1537–1545PubMedCrossRefGoogle Scholar
  22. 22.
    Wachtershauser A, Loitsch SM, Stein J (2000) PPAR-gamma is selectively upregulated in Caco-2 cells by butyrate. Biochem Biophys Res Commun 272:380–385PubMedCrossRefGoogle Scholar
  23. 23.
    Ulrich S, Wachtershauser A, Loitsch S, von Knethen A, Brune B, Stein J (2005) Activation of PPARgamma is not involved in butyrate-induced epithelial cell differentiation. Exp Cell Res 310:196–204PubMedCrossRefGoogle Scholar
  24. 24.
    Date M, Fukuchi K, Morita S, Takahashi H, Ohura K (2003) 15-Deoxy-delta12,14-prostaglandin J2, a ligand for peroxisome proliferators-activated receptor-gamma, induces apoptosis in human hepatoma cells. Liver Int 23:460–466PubMedCrossRefGoogle Scholar
  25. 25.
    Gurnell M, Wentworth JM, Agostini M, et al (2000) A dominant-negative peroxisome proliferator-activated receptor gamma (PPARgamma) mutant is a constitutive repressor and inhibits PPARgamma-mediated adipogenesis. J Biol Chem 275:5754–5759PubMedCrossRefGoogle Scholar
  26. 26.
    Gupta RA, Brockman JA, Sarraf P, Willson TM, DuBois RN (2001) Target genes of peroxisome proliferator-activated receptor gamma in colorectal cancer cells. J Biol Chem 276:29681–29687PubMedCrossRefGoogle Scholar
  27. 27.
    Miller SJ (2004) Cellular and physiological effects of short-chain fatty acids. Mini Rev Med Chem 4:839–845PubMedGoogle Scholar
  28. 28.
    Salminen A, Tapiola T, Korhonen P, Suuronen T (1998) Neuronal apoptosis induced by histone deacetylase inhibitors. Brain Res Mol Brain Res 61:203–206PubMedCrossRefGoogle Scholar
  29. 29.
    Emanuele S, D’Anneo A, Bellavia G, et al (2004) Sodium butyrate induces apoptosis in human hepatoma cells by a mitochondria/caspase pathway, associated with degradation of beta-catenin, pRb and Bcl-XL. Eur J Cancer 40:1441–1452PubMedCrossRefGoogle Scholar
  30. 30.
    Avivi-Green C, Polak-Charcon S, Madar Z, Schwartz B (2002) Different molecular events account for butyrate-induced apoptosis in two human colon cancer cell lines. J Nutr 132:1812–1818PubMedGoogle Scholar
  31. 31.
    Maher P (1999) p38 mitogen-activated protein kinase activation is required for fibroblast growth factor-2-stimulated cell proliferation but not differentiation. J Biol Chem 274:17491–17498PubMedCrossRefGoogle Scholar
  32. 32.
    Cho SD, Ahn NS, Jung JW, et al (2006) Critical role of the c-JunNH2-terminal kinase and p38 mitogen-activated protein kinase pathways on sodium butyrate-induced apoptosis in DU145 human prostate cancer cells. Eur J Cancer Prev 15:57–63PubMedCrossRefGoogle Scholar
  33. 33.
    Nick JA, Young SK, Brown KK, et al (2000) Role of p38 mitogen-activated protein kinase in a murine model of pulmonary inflammation. J Immunol 164:2151–2159PubMedGoogle Scholar
  34. 34.
    Nagata Y, Takahashi N, Davis RJ, Todokoro K (1998) Activation of p38 MAP kinase and JNK but not ERK is required for erythropoietin-induced erythroid differentiation. Blood 92:1859–1869PubMedGoogle Scholar
  35. 35.
    Daniel C, Schroder O, Zahn N, Gaschott T, Stein J (2004) p38 MAPK signaling pathway is involved in butyrate-induced vitamin D receptor expression. Biochem Biophys Res Commun 324:1220–1226PubMedCrossRefGoogle Scholar
  36. 36.
    Witt O, Sand K, Pekrun A (2000) Butyrate-induced erythroid differentiation of human K562 leukemia cells involves inhibition of ERK and activation of p38 MAP kinase pathways. Blood 95:2391–2396PubMedGoogle Scholar
  37. 37.
    Zhou J, Tang Y, Liu W, Sun H, Hu J, Gong J (2002) Molecular mechanism of enhanced apoptotic response in U937 cells mediated by sodium butyrate. Zhonghua Zhong Liu Za Zhi 24:320– 322PubMedGoogle Scholar
  38. 38.
    Ruemmele FM, Dionne S, Qureshi I, Sarma DS, Levy E, Seidman EG (1999) Butyrate mediates Caco-2 cell apoptosis via up-regulation of pro-apoptotic BAK and inducing caspase-3 mediated cleavage of poly-(ADP-ribose) polymerase (PARP). Cell Death Differ 6:729–735PubMedCrossRefGoogle Scholar
  39. 39.
    Moss SF, Agarwal B, Arber N, et al (1996) Increased intestinal Bak expression results in apoptosis. Biochem Biophys Res Commun 223:199–203PubMedCrossRefGoogle Scholar
  40. 40.
    Clem RJ, Cheng EH, Karp CL, et al (1998) Modulation of cell death by Bcl-XL through caspase interaction. Proc Natl Acad Sci USA 95:554–559PubMedCrossRefGoogle Scholar
  41. 41.
    Wang J, Friedman E (2000) Downregulation of p53 by sustained JNK activation during apoptosis. Mol Carcinog 29:179–188PubMedCrossRefGoogle Scholar
  42. 42.
    Auwerx J (2002) Nuclear receptors. I. PPAR gamma in the gastrointestinal tract: gain or pain? Am J Physiol Gastrointest Liver Physiol 282:G581–585PubMedGoogle Scholar
  43. 43.
    Theocharis S, Margeli A, Vielh P, Kouraklis G (2004) Peroxisome proliferator-activated receptor-gamma ligands as cell-cycle modulators. Cancer Treat Rev 30:545–554PubMedCrossRefGoogle Scholar
  44. 44.
    Kim EJ, Park KS, Chung SY, et al (2003) Peroxisome proliferator-activated receptor-gamma activator 15-deoxy-Delta12,14-prostaglandin J2 inhibits neuroblastoma cell growth through induction of apoptosis: association with extracellular signal-regulated kinase signal pathway. J Pharmacol Exp Ther 307:505–517PubMedCrossRefGoogle Scholar
  45. 45.
    Li MY, Deng H, Zhao JM, Dai D, Tan XY (2003) PPARgamma pathway activation results in apoptosis and COX-2 inhibition in HepG2 cells. World J Gastroenterol 9:1220–1226PubMedGoogle Scholar
  46. 46.
    Sarraf P, Mueller E, Smith WM, et al (1999) Loss-of-function mutations in PPAR gamma associated with human colon cancer. Mol Cell 3:799–804PubMedCrossRefGoogle Scholar
  47. 47.
    Bull AW, Steffensen KR, Leers J, Rafter JJ (2003) Activation of PPAR gamma in colon tumor cell lines by oxidized metabolites of linoleic acid, endogenous ligands for PPAR gamma. Carcinogenesis 24:1717–1722PubMedCrossRefGoogle Scholar
  48. 48.
    Natoni F, Diolordi L, Santoni C, Gilardini Montani MS (2005) Sodium butyrate sensitises human pancreatic cancer cells to both the intrinsic and the extrinsic apoptotic pathways. Biochim Biophys Acta 1745:318–329PubMedCrossRefGoogle Scholar
  49. 49.
    Kim EH, Kim HS, Kim SU, Noh EJ, Lee JS, Choi KS (2005) Sodium butyrate sensitizes human glioma cells to TRAIL-mediated apoptosis through inhibition of Cdc2 and the subsequent downregulation of survivin and XIAP. Oncogene 24:6877–6889PubMedCrossRefGoogle Scholar
  50. 50.
    Cheng Y, Chang LW, Tsou TC (2005) Mitogen-activated protein kinases mediate arsenic-induced down-regulation of survivin in human lung adenocarcinoma cells. Arch Toxicol 1–9Google Scholar
  51. 51.
    Qiu L, Wang Q, Di W, et al (2005) Transient activation of EGFR/AKT cell survival pathway and expression of survivin contribute to reduced sensitivity of human melanoma cells to betulinic acid. Int J Oncol 27:823–830PubMedGoogle Scholar
  52. 52.
    Johnson ME, Howerth EW (2004) Survivin: a bifunctional inhibitor of apoptosis protein. Vet Pathol 41:599–607PubMedCrossRefGoogle Scholar
  53. 53.
    Sanna MG, da Silva Correia J, Ducrey O, et al (2002) IAP suppression of apoptosis involves distinct mechanisms: the TAK1/JNK1 signaling cascade and caspase inhibition. Mol Cell Biol 22:1754–1766PubMedCrossRefGoogle Scholar
  54. 54.
    Kim Y, Suh N, Sporn M, Reed JC (2002) An inducible pathway for degradation of FLIP protein sensitizes tumor cells to TRAIL-induced apoptosis. J Biol Chem 277:22320–22329PubMedCrossRefGoogle Scholar
  55. 55.
    Hernandez A, Thomas R, Smith F, et al (2001) Butyrate sensitizes human colon cancer cells to TRAIL-mediated apoptosis. Surgery 130:265–272PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science + Business Media, LLC 2006

Authors and Affiliations

  • Markus Schwab
    • 1
  • Veerle Reynders
    • 1
  • Sandra Ulrich
    • 1
  • Nadine Zahn
    • 1
  • Jürgen Stein
    • 1
  • Oliver Schröder
    • 1
    Email author
  1. 1.First Department of Medicine-ZAFESJohann Wolfgang Goethe-University FrankfurtFrankfurt am MainGermany

Personalised recommendations