International Journal of Colorectal Disease

, Volume 22, Issue 4, pp 401–409 | Cite as

The anti-apoptotic and growth stimulatory actions of leptin in human colon cancer cells involves activation of JNK mitogen activated protein kinase, JAK2 and PI3 kinase/Akt

  • Olorunseun O. Ogunwobi
  • Ian L. P. BealesEmail author
Original Article


Background and aims

Obesity is a major risk factor for the development of colon cancer. Secretion of the hormone leptin from adipocytes is increased in obesity, and serum levels are proportional to body fat mass. Serum leptin levels are an independent risk factor for colon cancer. Leptin receptors are expressed in normal, premalignant and malignant colonic epithelia. We have investigated the effects of leptin on proliferation and apoptosis of colonic cancer cells and the early signalling events involved.


Proliferation of HT-29 colon cancer cells in response to leptin was assessed by 3-[4, 5-dimethylthiazol-2-y-l]-2, 5-diphenyltetrazolium bromide (MTT) assay, and apoptosis was quantified by enzyme-linked immunosorbent assay (ELISA) for intracellular nucleosomes. Signalling pathways involved were determined by using specific inhibitors, quantification of phosphorylated active intermediates and ELISA of active nuclear-translocated transcription factors.


Leptin stimulated HT-29 cell proliferation and inhibited both serum-starvation and celecoxib-induced apoptosis. The proliferative and anti-apoptotic effects of leptin were abolished by inhibition of JAK2 with AG490, phosphatidylinositol 3′-kinase (PI3 kinase) with LY294002 and c-Jun NH2-terminal kinase (JNK) with SP600125. Stimulation of HT-29 cells with leptin increased phosphorylation of JAK2, Akt and JNK. Activation of JAK2 was upstream of PI3 kinase/Akt but not of JNK. Activation of JAK2 was followed by activation and nuclear translocation of STAT3 and JNK activation led to increased activator protein 1 (AP-1) transcriptional activity.


Leptin stimulates proliferation and inhibits apoptosis in human colon cancer cells and may be an important factor in the increased incidence of colon cancer in obesity. This effect involves JAK2, PI3 kinase and JNK and activation of the oncogenic transcription factors signal transducer and activator of transcription (STAT)3 and AP-1.


Leptin Apoptosis Proliferation Colon cancer Janus tyrosine kinase 2 Akt c-Jun NH2-terminal kinase Activator protein 1 STAT3 protein Celecoxib 



activator protein 1




Dulbecco’s modified Eagle medium


dimethyl sulphoxide


enzyme-linked immunosorbent assay


foetal bovine serum


Janus tyrosine kinase


c-Jun NH2-terminal kinase


mitogen-activated protein kinase


3-[4, 5-dimethylthiazol-2-y-l]-2, 5-diphenyltetrazolium bromide


phosphate-buffered saline (pH 7.4)

PI3 kinase

phosphatidylinositol 3′-kinase


signal transducer and activator of transcription



This work was funded by the Norfolk and Norwich University Hospital Bicentenary Trust in the form of a research studentship to Dr. Ogunwobi. Further financial support was provided by The Royal Society, The Peel Medical Research Trust, The Mason Medical Research Foundation, The Institute of Biomedical Science, the Norfolk and Norwich University Hospital Inflammatory Bowel Disease Research Fund and National Health Service (NHS) Research and Development. Part of this work was presented in abstract form to the British Society of Gastroenterology, March 2005.


  1. 1.
    Tuynman JB, Peppelenbosch MP, Richel DJ (2004) COX-2 inhibition as a tool to treat and prevent colorectal cancer. Crit Rev Oncol Hematol 52:81–101PubMedGoogle Scholar
  2. 2.
    Giovannucci E, Ascherio A, Rimm EB, Colditz GA, Stampfer MJ, Willett WC (1995) Physical activity, obesity, and risk for colon cancer and adenoma in men. Ann Intern Med 122:327–334PubMedGoogle Scholar
  3. 3.
    Pan SY, Johnson KC, Ugnat AM, Wen SW, Mao Y (2004) Association of obesity and cancer risk in Canada. Am J Epidemiol 159: 259–268PubMedCrossRefGoogle Scholar
  4. 4.
    Considine RV, Sinha MK, Heiman ML, Kriauciunas A, Stephens TW, Nyce MR, Ohannesian JP, Marco CC, McKee LJ, Bauer TL et al (1996) Serum immunoreactive-leptin concentrations in normal-weight and obese humans. N Engl J Med 334:292–295PubMedCrossRefGoogle Scholar
  5. 5.
    Stattin P, Lukanova A, Biessy C, Soderberg S, Palmqvist R, Kaaks R, Olsson T, Jellum E (2004) Obesity and colon cancer: does leptin provide a link? Int J Cancer 109:149–152PubMedCrossRefGoogle Scholar
  6. 6.
    Attoub S, Noe V, Pirola L, Bruyneel E, Chastre E, Mareel M, Wymann MP, Gespach C (2000) Leptin promotes invasiveness of kidney and colonic epithelial cells via phosphoinositide 3-kinase-, rho-, and rac-dependent signaling pathways. FASEB J 14:2329–2338PubMedCrossRefGoogle Scholar
  7. 7.
    Hardwick JC, van Den Brink GR, Offerhaus GJ, van Deventer SJ, Peppelenbosch MP (2001) Leptin is a growth factor for colonic epithelial cells. Gastroenterology 121:79–90PubMedCrossRefGoogle Scholar
  8. 8.
    Myers MG Jr (2004) Leptin receptor signaling and the regulation of mammalian physiology. Recent Prog Horm Res 59:287–304PubMedCrossRefGoogle Scholar
  9. 9.
    Saxena NK, Titus MA, Ding X, Floyd J, Srinivasan S, Sitaraman SV, Anania FA (2004) Leptin as a novel profibrogenic cytokine in hepatic stellate cells: mitogenesis and inhibition of apoptosis mediated by extracellular regulated kinase (Erk) and Akt phosphorylation. FASEB J 18:1612–1614PubMedGoogle Scholar
  10. 10.
    Kim GS, Hong JS, Kim SW, Koh JM, An CS, Choi JY, Cheng SL (2003) Leptin induces apoptosis via ERK/cPLA2/cytochrome c pathway in human bone marrow stromal cells. J Biol Chem 278:21920–21929PubMedCrossRefGoogle Scholar
  11. 11.
    Brunet A, Bonni A, Zigmond MJ, Lin MZ, Juo P, Hu LS, Anderson MJ, Arden KC, Blenis J, Greenberg ME (1999) Akt promotes cell survival by phosphorylating and inhibiting a Forkhead transcription factor. Cell 96:857–868PubMedCrossRefGoogle Scholar
  12. 12.
    Thompson JE, Thompson CB (2004) Putting the rap on Akt. J Clin Oncol 22:4217–4226PubMedCrossRefGoogle Scholar
  13. 13.
    Somasundar P, Frankenberry KA, Skinner H, Vedula G, McFadden DW, Riggs D, Jackson B, Vangilder R, Hileman SM, Vona-Davis LC (2004) Prostate cancer cell proliferation is influenced by leptin. J Surg Res 118:71–82PubMedCrossRefGoogle Scholar
  14. 14.
    Stepan VM, Sawada M, Todisco A, Dickinson CJ (1999) Glycine-extended gastrin exerts growth-promoting effects on human colon cancer cells. Mol Med 5:147–159PubMedGoogle Scholar
  15. 15.
    Ogunwobi O, Beales I (2005) Glycine-extended gastrin stimulates proliferation and inhibits apoptosis in colon cancer cells via multiple pathways (abstract). Gut 54(Suppl 11):A57Google Scholar
  16. 16.
    Onuma M, Bub JD, Rummel TL, Iwamoto Y (2003) Prostate cancer cell–adipocyte interaction: leptin mediates androgen-independent prostate cancer cell proliferation through c-Jun NH2-terminal kinase. J Biol Chem 278:42660–42667PubMedCrossRefGoogle Scholar
  17. 17.
    Sakai I, Kraft AS (1997) The kinase domain of Jak2 mediates induction of bcl-2 and delays cell death in hematopoietic cells. J Biol Chem 272:12350–12358PubMedCrossRefGoogle Scholar
  18. 18.
    Kloek C, Haq AK, Dunn SL, Lavery HJ, Banks AS, Myers MG Jr (2002) Regulation of Jak kinases by intracellular leptin receptor sequences. J Biol Chem 277:41547–41555PubMedCrossRefGoogle Scholar
  19. 19.
    Pai R, Lin C, Tran T, Tarnawski A (2005) Leptin activates STAT and ERK2 pathways and induces gastric cancer cell proliferation. Biochem Biophys Res Commun 331:984–992PubMedCrossRefGoogle Scholar
  20. 20.
    Vaisse C, Halaas JL, Horvath CM, Darnell JE Jr, Stoffel M, Friedman JM (1996) Leptin activation of Stat3 in the hypothalamus of wild-type and ob/ob mice but not db/db mice. Nat Genet 14:95–97PubMedCrossRefGoogle Scholar
  21. 21.
    Ma XT, Wang S, Ye YJ, Du RY, Cui ZR, Somsouk M (2004) Constitutive activation of Stat3 signaling pathway in human colorectal carcinoma. World J Gastroenterol 10:1569–1573PubMedGoogle Scholar
  22. 22.
    Steinbach G, Lynch PM, Phillips RK, Wallace MH, Hawk E, Gordon GB, Wakabayashi N, Saunders B, Shen Y, Fujimura T et al (2000) The effect of celecoxib, a cyclooxygenase-2 inhibitor, in familial adenomatous polyposis. N Engl J Med 342:1946–1952PubMedCrossRefGoogle Scholar
  23. 23.
    Corpet DE, Pierre F (2005) How good are rodent models of carcinogenesis in predicting efficacy in humans? A systematic review and meta-analysis of colon chemoprevention in rats, mice and men. Eur J Cancer 41:1911–1922PubMedCrossRefGoogle Scholar
  24. 24.
    Maier TJ, Schilling K, Schmidt R, Geisslinger G, Grosch S (2004) Cyclooxygenase-2 (COX-2)-dependent and -independent anticarcinogenic effects of celecoxib in human colon carcinoma cells. Biochem Pharmacol 67:1469–1478PubMedCrossRefGoogle Scholar
  25. 25.
    Beales I, Calam J (1997) Stimulation of IL-8 production in human gastric epithelial cells by Helicobacter pylori, IL-1β and TNF-α requires tyrosine kinase activity, but not protein kinase C. Cytokine 9:514–520PubMedCrossRefGoogle Scholar
  26. 26.
    Beales IL (2004) Gastrin and interleukin-1beta stimulate growth factor secretion from cultured rabbit gastric parietal cells. Life Sci 75:2983–2995PubMedCrossRefGoogle Scholar
  27. 27.
    Micoud F, Mandrand B, Malcus-Vocanson C (2001) Comparison of several techniques for the detection of apoptotic astrocytes in vitro. Cell Prolif 34:99–113PubMedCrossRefGoogle Scholar
  28. 28.
    Ogunwobi OO, Beales IL (2006) Glycine-extended gastrin stimulates proliferation and inhibits apoptosis in colon cancer cells via cyclo-oxygenase-independent pathways. Regul Pept 134:1–8PubMedCrossRefGoogle Scholar
  29. 29.
    Ostlund RE Jr, Yang JW, Klein S, Gingerich R (1996) Relation between plasma leptin concentration and body fat, gender, diet, age, and metabolic covariates. J Clin Endocrinol Metab 81:3909–3913PubMedCrossRefGoogle Scholar
  30. 30.
    Zimmet P, Hodge A, Nicolson M, Staten M, de Courten M, Moore J, Morawiecki A, Lubina J, Collier G, Alberti G et al (1996) Serum leptin concentration, obesity, and insulin resistance in Western Samoans: cross sectional study. BMJ 313:965–969PubMedGoogle Scholar
  31. 31.
    Datta SR, Dudek H, Tao X, Masters S, Fu H, Gotoh Y, Greenberg ME (1997) Akt phosphorylation of BAD couples survival signals to the cell-intrinsic death machinery. Cell 91:231–241PubMedCrossRefGoogle Scholar
  32. 32.
    Skov S, Klausen P, Claesson MH (1997) Ligation of major histocompatibility complex (MHC) class I molecules on human T cells induces cell death through PI-3 kinase-induced c-Jun NH2-terminal kinase activity: a novel apoptotic pathway distinct from Fas-induced apoptosis. J Cell Biol 139:1523–1531PubMedCrossRefGoogle Scholar
  33. 33.
    Bjorbaek C, Uotani S, da Silva B, Flier JS (1997) Divergent signaling capacities of the long and short isoforms of the leptin receptor. J Biol Chem 272:32686–32695PubMedCrossRefGoogle Scholar
  34. 34.
    Arico S, Pattingre S, Bauvy C, Gane P, Barbat A, Codogno P, Ogier-Denis E (2002) Celecoxib induces apoptosis by inhibiting 3-phosphoinositide-dependent protein kinase-1 activity in the human colon cancer HT-29 cell line. J Biol Chem 277:27613–27621PubMedCrossRefGoogle Scholar
  35. 35.
    Kumar NV, Bernstein LR (2001) Ten ERK-related proteins in three distinct classes associate with AP-1 proteins and/or AP-1 DNA. J Biol Chem 276:32362–32372PubMedCrossRefGoogle Scholar
  36. 36.
    Zhong SP, Ma WY, Quealy JA, Zhang Y, Dong Z (2001) Organ-specific distribution of AP-1 in AP-1 luciferase transgenic mice during the maturation process. Am J Physiol Regul Integr Comp Physiol 280:R376–R381PubMedGoogle Scholar
  37. 37.
    Shaulian E, Karin M (2001) AP-1 in cell proliferation and survival. Oncogene 20:2390–2400PubMedCrossRefGoogle Scholar
  38. 38.
    Ashida R, Tominaga K, Sasaki E, Watanabe T, Fujiwara Y, Oshitani N, Higuchi K, Mitsuyama S, Iwao H, Arakawa T (2005) AP-1 and colorectal cancer. Inflammopharmacology 13:113–125PubMedCrossRefGoogle Scholar
  39. 39.
    Rivat C, Rodrigues S, Bruyneel E, Pietu G, Robert A, Redeuilh G, Bracke M, Gespach C, Attoub S (2005) Implication of STAT3 signaling in human colonic cancer cells during intestinal trefoil factor 3 (TFF3)—and vascular endothelial growth factor-mediated cellular invasion and tumor growth. Cancer Res 65:195–202PubMedGoogle Scholar
  40. 40.
    Bromberg J (2002) Stat proteins and oncogenesis. J Clin Invest 109:1139–1142PubMedCrossRefGoogle Scholar
  41. 41.
    Rouet-Benzineb P, Aparicio T, Guilmeau S, Pouzet C, Descatoire V, Buyse M, Bado A (2004) Leptin counteracts sodium butyrate_induced apoptosis in human colon cancer HT-29 cells via NF-kappaB signaling. J Biol Chem 279:16495–16502PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2006

Authors and Affiliations

  1. 1.Gastroenterology DepartmentNorfolk and Norwich University HospitalNorwichUK
  2. 2.Gastroenterology Research Unit, School of Medicine, Health Policy and PracticeUniversity of East AngliaNorwichUK

Personalised recommendations