Hepatocellular alterations and dysregulation of oncogenic pathways in the liver of transgenic mice overexpressing growth hormone

Cell Cycle

Volumn 12, Issue 7, 2013, Pages 1042-1057

  Miquet, Johanna G ^a   Freund, Thomas ^a   Martinez, Carolina S ^a   González, Lorena ^a   Díaz, María E ^a   Micucci, Giannina P ^a   Zotta, Elsa ^a   Boparai, Ravneet K ^b   Bartke, Andrzej ^b   Turyn, Daniel ^a   Sotelo, Ana I ^a  

^a UNIVERSIDAD DE BUENOS AIRES   (Argentina)

^b SOUTHERN ILLINOIS UNIVERSITY SCHOOL OF MEDICINE   (United States)

Author keywords

C fos; C jun; C myc; Cyclin D1; Cyclin E; Growth hormone; GSK3 ; Liver; NF B; catenin

Indexed keywords

BETA CATENIN; CYCLIN D1; CYCLIN E; GROWTH HORMONE; IMMUNOGLOBULIN ENHANCER BINDING PROTEIN; PROTEIN C JUN; PROTEIN KINASE B BETA;

ANIMAL CELL; ARTICLE; CELL GROWTH; CELL PROLIFERATION; CELL SURVIVAL; CONTROLLED STUDY; ENZYME ACTIVATION; FEMALE; GENE OVEREXPRESSION; HYPERTROPHY; LIVER CELL CARCINOMA; LONG TERM EXPOSURE; MALE; MOUSE; NONHUMAN; NUCLEAR SIZE; ONCOGENE C FOS; ONCOGENE C MYC; SEX DIFFERENCE; SIGNAL TRANSDUCTION; TRANSGENIC MOUSE;

EID: 84875906278     PISSN: 15384101     EISSN: 15514005     Source Type: Journal    
DOI: 10.4161/cc.24026     Document Type: Article

References (80)

1. Brooks AJ, Waters MJ. The growth hormone receptor: mechanism of activation and clinical implications. Nat Rev Endocrinol 2010; 6:515-25; PMID:20664532; http://dx.doi.org/10.1038/nrendo.2010.123
2. Lichanska AM, Waters MJ. How growth hormone controls growth, obesity and sexual dimorphism. Trends Genet 2008; 24:41-7; PMID:18063438; http://dx.doi. org/10.1016/j.tig.2007.10.006
3. Xu J, Messina JL. Crosstalk between growth hormone and insulin signaling. Vitam Horm 2009; 80:125-53; PMID:19251037; http://dx.doi.org/10.1016/S0083- 6729(08)00606-7
4. Jansson JO, Ekberg S, Hoath SB, Beamer WG, Frohman LA. Growth hormone enhances hepatic epidermal growth factor receptor concentration in mice. J Clin Invest 1988; 82:1871-6; PMID:3264288; http://dx.doi.org/10.1172/JCI113804
5. Ekberg S, Carlsson L, Carlsson B, Billig H, Jansson JO. Plasma growth hormone pattern regulates epidermal growth factor (EGF) receptor messenger ribonucleic acid levels and EGF binding in the rat liver. Endocrinology 1989; 125:2158-66; PMID:2791983; http://dx.doi.org/10.1210/endo-125-4-2158
6. Miquet JG, González L, Matos MN, Hansen CE, Louis A, Bartke A, et al. Transgenic mice overexpressing GH exhibit hepatic upregulation of GH-signaling mediators involved in cell proliferation. J Endocrinol 2008; 198:317-30; PMID:18480380; http://dx.doi. org/10.1677/JOE-08-0002
7. Jenkins PJ, Mukherjee A, Shalet SM. Does growth hormone cause cancer? Clin Endocrinol (Oxf ) 2006; 64:115-21; PMID:16430706; http://dx.doi. org/10.1111/j.1365-2265.2005.02404.x
8. Perry JK, Emerald BS, Mertani HC, Lobie PE. The oncogenic potential of growth hormone. Growth Horm IGF Res 2006; 16:277-89; PMID:17101287; http://dx.doi.org/10.1016/j.ghir.2006.09.006
9. McGrane MM, Yun JS, Moorman AFM, Lamers WH, Hendrick GK, Arafah BM, et al. Metabolic effects of developmental, tissue-, and cell-specific expression of a chimeric phosphoenolpyruvate carboxykinase (GTP)/bovine growth hormone gene in transgenic mice. J Biol Chem 1990; 265:22371-9; PMID:1702419
10. Kopchick JJ, Bellush LL, Coschigano KT. Transgenic models of growth hormone action. Annu Rev Nutr 1999; 19:437-61; PMID:10448532; http://dx.doi. org/10.1146/annurev.nutr.19.1.437
11. Bartke A. Can growth hormone (GH) accelerate aging? Evidence from GH-transgenic mice. Neuroendocrinology 2003; 78:210-6; PMID:14583653; http://dx.doi.org/10.1159/000073704
12. Wanke R, Hermanns W, Folger S, Wolf E, Brem G. Accelerated growth and visceral lesions in transgenic mice expressing foreign genes of the growth hormone family: an overview. Pediatr Nephrol 1991; 5:513-21; PMID:1911131; http://dx.doi.org/10.1007/BF01453693
13. Orian JM, Lee CS, Weiss LM, Brandon MR. The expression of a metallothionein-ovine growth hormone fusion gene in transgenic mice does not impair fertility but results in pathological lesions in the liver. Endocrinology 1989; 124:455-63; PMID:2642419; http://dx.doi.org/10.1210/endo-124-1-455
14. Orian JM, Tamakoshi K, Mackay IR, Brandon MR. New murine model for hepatocellular carcinoma: transgenic mice expressing metallothionein-ovine growth hormone fusion gene. J Natl Cancer Inst 1990; 82:393-8; PMID:2154583; http://dx.doi.org/10.1093/jnci/82.5.393
15. Quaife CJ, Mathews LS, Pinkert CA, Hammer RE, Brinster RL, Palmiter RD. Histopathology associated with elevated levels of growth hormone and insulinlike growth factor I in transgenic mice. Endocrinology 1989; 124:40-8; PMID:2642417; http://dx.doi. org/10.1210/endo-124-1-40
16. Snibson KJ, Bhathal PS, Hardy CL, Brandon MR, Adams TE. High, persistent hepatocellular proliferation and apoptosis precede hepatocarcinogenesis in growth hormone transgenic mice. Liver 1999; 19:242-52; PMID:10395045; http://dx.doi. org/10.1111/j.1478-3231.1999.tb00042.x
17. Snibson KJ. Hepatocellular kinetics and the expression of growth hormone (GH) in the livers and liver tumours of GH-transgenic mice. Tissue Cell 2002; 34:88-97; PMID:12165243; http://dx.doi. org/10.1016/S0040-8166(02)00012-5
18. Hoeflich A, Nedbal S, Blum WF, Erhard M, Lahm H, Brem G, et al. Growth inhibition in giant growth hormone transgenic mice by overexpression of insulinlike growth factor-binding protein-2. Endocrinology 2001; 142:1889-98; PMID:11316754; http://dx.doi. org/10.1210/en.142.5.1889
19. Coleman WB. Mechanisms of human hepatocarcinogenesis. Curr Mol Med 2003; 3:573-88; PMID:14527088; http://dx.doi.org/10.2174/1566524033479546
20. Teoh NC. Proliferative drive and liver carcinogenesis: too much of a good thing? J Gastroenterol Hepatol 2009; 24:1817-25; PMID:20002941; http://dx.doi. org/10.1111/j.1440-1746.2009.06121.x
21. Snibson KJ, Bhathal PS, Adams TE. Overexpressed growth hormone (GH) synergistically promotes carcinogen-initiated liver tumour growth by promoting cellular proliferation in emerging hepatocellular neoplasms in female and male GH-transgenic mice. Liver 2001; 21:149-58; PMID:11318985; http://dx.doi. org/10.1034/j.1600-0676.2001.021002149.x
22. Vergani G, Mayerhofer A, Bartke A. Acute effects of human growth hormone on liver cells in vitro: a comparison with livers of mice transgenic for human growth hormone. Tissue Cell 1991; 23:607-12; PMID:1776152; http://dx.doi.org/10. 1016/0040- 8166(91)90017-N
23. Llovet JM, Bruix J. Molecular targeted therapies in hepatocellular carcinoma. Hepatology 2008; 48:1312- 27; PMID:18821591; http://dx.doi.org/10. 1002/hep.22506
24. Spangenberg HC, Thimme R, Blum HE. Targeted therapy for hepatocellular carcinoma. Nat Rev Gastroenterol Hepatol 2009; 6:423-32; PMID:19488072; http://dx.doi.org/10.1038/nrgastro.2009.86
25. Zender L, Villanueva A, Tovar V, Sia D, Chiang DY, Llovet JM. Cancer gene discovery in hepatocellular carcinoma. J Hepatol 2010; 52:921-9; PMID:20385424; http://dx.doi.org/10.1016/j.jhep.2009.12.034
26. Rogers AB, Theve EJ, Feng Y, Fry RC, Taghizadeh K, Clapp KM, et al. Hepatocellular carcinoma associated with liver-gender disruption in male mice. Cancer Res 2007; 67:11536-46; PMID:18089782; http://dx.doi. org/10.1158/0008- 5472.CAN-07-1479
27. Ruggieri A, Barbati C, Malorni W. Cellular and molecular mechanisms involved in hepatocellular carcinoma gender disparity. Int J Cancer 2010; 127:499- 504; PMID:20201099; http://dx.doi.org/10.1002/ijc.25298
28. Popow A, Nowak D, Malicka-Błaszkiewicz M. Actin cytoskeleton and beta-actin expression in correlation with higher invasiveness of selected hepatoma Morris 5123 cells. J Physiol Pharmacol 2006; 57(Suppl 7):111-23; PMID:17228099
29. Miller LM, Menthena A, Chatterjee C, Verdier-Pinard P, Novikoff PM, Horwitz SB, et al. Increased levels of a unique post-translationally modified betaIVb-tubulin isotype in liver cancer. Biochemistry 2008; 47:7572- 82; PMID:18570381; http://dx.doi.org/10.1021/bi8005225
30. Miquet JG, Sotelo AI, Bartke A, Turyn D. Suppression of growth hormone (GH) Janus tyrosine kinase 2/signal transducer and activator of transcription 5 signaling pathway in transgenic mice overexpressing bovine GH. Endocrinology 2004; 145:2824-32; PMID:15016718; http://dx.doi.org/10.1210/en.2003-1498
31. Sotelo AI, Miquet JG, González L, Bartke A, Turyn D. Vitamin D3 cannot revert desensitization of growth hormone (GH)-induced STAT5-signaling in GH-overexpressing mice non-calcemic tissues. Growth Horm IGF Res 2008; 18:148-56; PMID:17881271; http://dx.doi.org/10.1016/j.ghir.2007.08.001
32. Jope RS, Yuskaitis CJ, Beurel E. Glycogen synthase kinase-3 (GSK3): inflammation, diseases, and therapeutics. Neurochem Res 2007; 32:577-95; PMID:16944320; http://dx.doi.org/10.1007/s11064- 006-9128-5
33. Jacobs KM, Bhave SR, Ferraro DJ, Jaboin JJ, Hallahan DE, Thotala D. GSK-3β: A Bifunctional Role in Cell Death Pathways. Int J Cell Biol 2012; 2012:930710; PMID:22675363; http://dx.doi. org/10.1155/2012/930710
34. Nicholson KM, Anderson NG. The protein kinase B/Akt signalling pathway in human malignancy. Cell Signal 2002; 14:381-95; PMID:11882383; http://dx.doi.org/10.1016/S0898-6568(01)00271-6
35. Takahashi-Yanaga F, Sasaguri T. GSK-3beta regulates cyclin D1 expression: a new target for chemotherapy. Cell Signal 2008; 20:581-9; PMID:18023328; http://dx.doi.org/10.1016/j.cellsig.2007.10.018
36. Luedde T, Schwabe RFNF. NF-κB in the liver- -linking injury, fibrosis and hepatocellular carcinoma. Nat Rev Gastroenterol Hepatol 2011; 8:108-18; PMID:21293511; http://dx.doi.org/10.1038/nrgastro. 2010.213
37. Schmid JA, Birbach A. IkappaB kinase beta (IKKbeta/IKK2/IKBKB)-a key molecule in signaling to the transcription factor NF-kappaB. Cytokine Growth Factor Rev 2008; 19:157-65; PMID:18308615; http://dx.doi. org/10.1016/j.cytogfr. 2008.01.006
38. Song G, Ouyang G, Bao S. The activation of Akt/PKB signaling pathway and cell survival. J Cell Mol Med 2005; 9:59-71; PMID:15784165; http://dx.doi. org/10.1111/j.1582-4934.2005.tb00337.x
39. Feitelson MA, Sun B, Satiroglu Tufan NL, Liu J, Pan J, Lian Z. Genetic mechanisms of hepatocarcinogenesis. Oncogene 2002; 21:2593-604; PMID:11971194; http://dx.doi.org/10.1038/sj.onc.1205434
40. Yuen MF, Wu PC, Lai VC, Lau JY, Lai CL. Expression of c-Myc, c-Fos, and c-jun in hepatocellular carcinoma. Cancer 2001; 91:106-12; PMID:11148566; http://dx.doi. org/10.1002/1097-0142(20010101)91:1<106::AIDCNCR14> 3.0.CO;2-2
41. Stamatakos M, Palla V, Karaiskos I, Xiromeritis K, Alexiou I, Pateras I, et al. Cell cyclins: triggering elements of cancer or not? World J Surg Oncol 2010; 8:111; PMID:21176227; http://dx.doi. org/10.1186/1477-7819-8-111
42. Baik M, Yu JH, Hennighausen L. Growth hormone- STAT5 regulation of growth, hepatocellular carcinoma, and liver metabolism. Ann N Y Acad Sci 2011; 1229:29-37; PMID:21793836; http://dx.doi. org/10.1111/j.1749-6632.2011.06100.x
43. Waxman DJ, O'Connor C. Growth hormone regulation of sex-dependent liver gene expression. Mol Endocrinol 2006; 20:2613-29; PMID:16543404; http://dx.doi.org/10.1210/me.2006-0007
44. Zhang Y, Laz EV, Waxman DJ. Dynamic, sex-differential STAT5 and BCL6 binding to sex-biased, growth hormone-regulated genes in adult mouse liver. Mol Cell Biol 2012; 32:880-96; PMID:22158971; http://dx.doi.org/10.1128/MCB.06312-11
45. Tannenbaum GS, Choi HK, Gurd W, Waxman DJ. Temporal relationship between the sexually dimorphic spontaneous GH secretory profiles and hepatic STAT5 activity. Endocrinology 2001; 142:4599- 606; PMID:11606424; http://dx.doi.org/ 10.1210/en.142.11.4599
46. MacLeod JN, Pampori NA, Shapiro BH. Sex differences in the ultradian pattern of plasma growth hormone concentrations in mice. J Endocrinol 1991; 131:395- 9; PMID:1783886; http://dx.doi.org/10.1677/joe.0.1310395
47. Sueyoshi T, Yokomori N, Korach KS, Negishi M. Developmental action of estrogen receptor-alpha feminizes the growth hormone-Stat5b pathway and expression of Cyp2a4 and Cyp2d9 genes in mouse liver. Mol Pharmacol 1999; 56:473-7; PMID:10462534
48. Martinez CS, Piazza VG, Ratner LD, Matos MN, González L, Rulli SB, et al. Growth hormone STAT5- mediated signaling and its modulation in mice liver during the growth period. Growth Horm IGF Res 2013;23:19-28; PMID:23245546; http://dx.doi. org/10.1016/j.ghir.2012.11.002
49. Miquet JG, Muñoz MC, Giani JF, González L, Dominici FP, Bartke A, et al. Ames dwarf (Prop1(df )/Prop1(df )) mice display increased sensitivity of the major GH-signaling pathways in liver and skeletal muscle. Growth Horm IGF Res 2010; 20:118- 26; PMID:20022531; http://dx.doi.org/10.1016/ j. ghir.2009.11.003
50. Haga S, Ozaki M, Inoue H, Okamoto Y, Ogawa W, Takeda K, et al. The survival pathways phosphatidylinositol- 3 kinase (PI3-K)/phosphoinositide- dependent protein kinase 1 (PDK1)/Akt modulate liver regeneration through hepatocyte size rather than proliferation. Hepatology 2009; 49:204-14; PMID:19065678; http://dx.doi.org/10.1002/hep.22583
51. Mullany LK, Nelsen CJ, Hanse EA, Goggin MM, Anttila CK, Peterson M, et al. Akt-mediated liver growth promotes induction of cyclin E through a novel translational mechanism and a p21-mediated cell cycle arrest. J Biol Chem 2007; 282:21244- 52; PMID:17517888; http://dx.doi.org/10.1074/jbc. M702110200
52. Friedbichler K, Themanns M, Mueller KM, Schlederer M, Kornfeld JW, Terracciano LM, et al. Growthhormone-induced signal transducer and activator of transcription 5 signaling causes gigantism, inflammation, and premature death but protects mice from aggressive liver cancer. Hepatology 2012; 55:941- 52; PMID:22031092; http://dx.doi.org/10.1002/hep.24765
53. Fabregat I. Dysregulation of apoptosis in hepatocellular carcinoma cells. World J Gastroenterol 2009; 15:513- 20; PMID:19195051; http://dx.doi.org/10. 3748/wjg.15.513
54. Normanno N, De Luca A, Bianco C, Strizzi L, Mancino M, Maiello MR, et al. Epidermal growth factor receptor (EGFR) signaling in cancer. Gene 2006; 366:2-16; PMID:16377102; http://dx.doi. org/10.1016/j.gene.2005.10.018
55. Xu X, Sakon M, Nagano H, Hiraoka N, Yamamoto H, Hayashi N, et al. Akt2 expression correlates with prognosis of human hepatocellular carcinoma. Oncol Rep 2004; 11:25-32; PMID:14654898
56. Dahmani R, Just PA, Perret C. The Wnt/β-catenin pathway as a therapeutic target in human hepatocellular carcinoma. Clin Res Hepatol Gastroenterol 2011; 35:709-13; PMID:21778132; http://dx.doi. org/10.1016/j.clinre.2011.05.010
57. Monga SP. Role of Wnt/β-catenin signaling in liver metabolism and cancer. Int J Biochem Cell Biol 2011; 43:1021-9; PMID:19747566; http://dx.doi. org/10.1016/j.biocel.2009.09.001
58. Gotoh J, Obata M, Yoshie M, Kasai S, Ogawa K. Cyclin D1 over-expression correlates with beta-catenin activation, but not with H-ras mutations, and phosphorylation of Akt, GSK3 beta and ERK1/2 in mouse hepatic carcinogenesis. Carcinogenesis 2003; 24:435- 42; PMID:12663502; http://dx.doi.org/10.1093/ carcin/24.3.435
59. Ban KC, Singh H, Krishnan R, Seow HF. GSK-3beta phosphorylation and alteration of beta-catenin in hepatocellular carcinoma. Cancer Lett 2003; 199:201-8; PMID:12969793; http://dx.doi.org/10.1016/S0304- 3835(03)00421-X
60. Joo M, Kang YK, Kim MR, Lee HK, Jang JJ. Cyclin D1 overexpression in hepatocellular carcinoma. Liver 2001; 21:89-95; PMID:11318977; http://dx.doi. org/10.1034/j.1600-0676.2001.021002089.x
61. Nelsen CJ, Rickheim DG, Timchenko NA, Stanley MW, Albrecht JH. Transient expression of cyclin D1 is sufficient to promote hepatocyte replication and liver growth in vivo. Cancer Res 2001; 61:8564-8; PMID:11731443
62. Nelsen CJ, Rickheim DG, Tucker MM, Hansen LK, Albrecht JH. Evidence that cyclin D1 mediates both growth and proliferation downstream of TOR in hepatocytes. J Biol Chem 2003; 278:3656- 63; PMID:12446670; http://dx.doi.org/ 10.1074/jbc. M209374200
63. Ahn KS, Aggarwal BB. Transcription factor NF-kappaB: a sensor for smoke and stress signals. Ann NY Acad Sci 2005; 1056:218-33; PMID:16387690; http://dx.doi. org/10.1196/annals.1352.026
64. Lin CP, Liu CR, Lee CN, Chan TS, Liu HE. Targeting c-Myc as a novel approach for hepatocellular carcinoma. World J Hepatol 2010; 2:16-20; PMID:21160952
65. Beer S, Zetterberg A, Ihrie RA, McTaggart RA, Yang Q, Bradon N, et al. Developmental context determines latency of MYC-induced tumorigenesis. PLoS Biol 2004; 2:e332; PMID:15455033; http://dx.doi. org/10.1371/journal.pbio.0020332
66. Hartl M, Bader AG, Bister K. Molecular targets of the oncogenic transcription factor jun. Curr Cancer Drug Targets 2003; 3:41-55; PMID:12570660; http://dx.doi.org/10.2174/1568009033333781
67. Behrens A, Sibilia M, David JP, Möhle-Steinlein U, Tronche F, Schütz G, et al. Impaired postnatal hepatocyte proliferation and liver regeneration in mice lacking c-jun in the liver. EMBO J 2002; 21:1782- 90; PMID:11927562; http://dx.doi.org/10.1093/emboj/21.7.1782
68. Milde-Langosch K. The Fos family of transcription factors and their role in tumourigenesis. Eur J Cancer 2005; 41:2449-61; PMID:16199154; http://dx.doi. org/10.1016/j.ejca.2005.08.008
69. Okamoto T, Sanda T, Asamitsu K. NF-kappa B signaling and carcinogenesis. Curr Pharm Des 2007; 13:447-62; PMID:17348842; http://dx.doi. org/10.2174/138161207780162944
70. Perkins ND. The diverse and complex roles of NF-κB subunits in cancer. Nat Rev Cancer 2012; 12:121-32; PMID:22257950
71. Wullaert A, van Loo G, Heyninck K, Beyaert R. Hepatic tumor necrosis factor signaling and nuclear factor-kappaB: effects on liver homeostasis and beyond. Endocr Rev 2007; 28:365-86; PMID:17431229; http://dx.doi.org/10.1210/er. 2006-0031
72. Jeay S, Sonenshein GE, Postel-Vinay MC, Kelly PA, Baixeras E. Growth hormone can act as a cytokine controlling survival and proliferation of immune cells: new insights into signaling pathways. Mol Cell Endocrinol 2002; 188:1-7; PMID:11911939; http://dx.doi.org/10.1016/S0303-7207(02)00014-X
73. He G, Karin MNF. NF-κB and STAT3 - key players in liver inflammation and cancer. Cell Res 2011; 21:159- 68; PMID:21187858; http://dx.doi.org/10.1038/cr.2010.183
74. Abramovitch R, Tavor E, Jacob-Hirsch J, Zeira E, Amariglio N, Pappo O, et al. A pivotal role of cyclic AMP-responsive element binding protein in tumor progression. Cancer Res 2004; 64:1338-46; PMID:14973073; http://dx.doi.org/10. 1158/0008- 5472.CAN-03-2089
75. Chin KT, Zhou HJ, Wong CM, Lee JM, Chan CP, Qiang BQ, et al. The liver-enriched transcription factor CREB-H is a growth suppressor protein underexpressed in hepatocellular carcinoma. Nucleic Acids Res 2005; 33:1859-73; PMID:15800215; http://dx.doi. org/10.1093/nar/gki332
76. Mayo KE, Hammer RE, Swanson LW, Brinster RL, Rosenfeld MG, Evans RM. Dramatic pituitary hyperplasia in transgenic mice expressing a human growth hormone-releasing factor gene. Mol Endocrinol 1988; 2:606-12; PMID:3137455; http://dx.doi.org/10.1210/mend-2-7-606
77. Welinder C, Ekblad L. Coomassie staining as loading control in Western blot analysis. J Proteome Res 2011; 10:1416-9; PMID:21186791; http://dx.doi. org/10.1021/pr1011476
78. Chomczynski P, Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenolchloroform extraction. Anal Biochem 1987; 162:156- 9; PMID:2440339; http://dx.doi.org/10.1016/0003- 2697(87)90021-2
79. Masternak MM, Al-Regaiey KA, Del Rosario Lim MM, Bonkowski MS, Panici JA, Przybylski GK, et al. Caloric restriction results in decreased expression of peroxisome proliferators-activated receptors (PPARs) superfamily in muscle of normal and longlived GHR-KO mice. J Gerontol A Biol Sci Med Sci 2005; 60:1238-45; PMID:16282554; http://dx.doi. org/10.1093/gerona/60.10.1238
80. Wang Z, Masternak MM, Al-Regaiey KA, Bartke A. Adipocytokines and the regulation of lipid metabolism in growth hormone transgenic and calorierestricted mice. Endocrinology 2007; 148:2845-53; PMID:17347312; http://dx.doi.org/10.1210/en.2006- 1313