Insulin-like Growth Factor I (IGF-I) Stimulates Proliferation but also Increases Caspase-3 Activity, Annexin-V Binding, and DNA-fragmentation in Human MG63 Osteosarcoma Cells: Co-activation of Pro- and Anti-apoptotic Pathways by IGF-I

K. Raile; R. Hille; S. Laue; A. Schulz; G. Pfeifer; F. Horn; W. Kiess

doi:10.1055/s-2004-814140

Hormone and Metabolic Research, Inhaltsverzeichnis

Horm Metab Res 2003; 35(11/12): 786-793
DOI: 10.1055/s-2004-814140

Original

Insulin-like Growth Factor I (IGF-I) Stimulates Proliferation but also Increases Caspase-3 Activity, Annexin-V Binding, and DNA-fragmentation in Human MG63 Osteosarcoma Cells: Co-activation of Pro- and Anti-apoptotic Pathways by IGF-I

K. Raile¹ , R. Hille¹ , S. Laue¹ , A. Schulz¹ , G. Pfeifer² , F. Horn² , W. Kiess¹

¹Research Laboratory, Hospital for Children and Adolescents, Faculty of Medicine, University of Leipzig, Leipzig, Germany
²Dept. of Immunology, Faculty of Medicine, University of Leipzig, Leipzig, Germany

Abstract

Insulin-like growth factor-I (IGF-I) was found to promote proliferation, cell survival, and inhibition of apoptosis. But in some instances, IGF-I was found to mildly induce apoptosis, i. e. Fas-mediated apoptosis in human MG63 osteosarcoma cells. In the present study, we intended to further investigate IGF-I dependent pathways leading either to proliferation and cell survival or to cell death. MG63 osteosarcoma cells were treated with serum free medium alone or in combination with IGF-I, a neutralizing antibody against the human IGF-I receptor (αIR-3) or non-immune control IgG₁ for two to six days. We investigated cell survival (cell count), proliferation (CD71-FACS), apoptosis (Annexin-V-FACS, Caspase-3 activity, PCD) and anti-apoptosis (112-Ser Bad phosphorylation), and regulation of IGF-I receptor surface expression (IGF-I receptor-FACS).

We found that IGF-I treatment (48 h) stimulated cell growth and proliferation, but also mildly induced apoptosis. IGF-I activated specific apoptotic pathways (Caspase-3 activation, Annexin-V binding and DNA degradation), as well as anti-apoptotic signals (Bad phosphorylation at serine 112). αIR-3 blocked cell proliferation, strongly induced apoptosis, and inhibited Bad-phosphorylation. Thus, IGF-I treatment overall resulted in increased tumour cell mass, despite a detectable stimulation of apoptosis; in other words proliferation exceeded cell death. If IGF-I was first added on day 0, 2, or 4 of serum free culture, we found decreasing IGF-I specific effects on proliferation and apoptosis. In parallel, we found a down-regulation of IGF-I receptors (FACS) by serum withdrawal, which was partly reversed if IGF-I was added. Therefore receptor number might have an impact on IGF-I function in MG63 cells. In conclusion, co-activation of apoptosis and proliferation by IGF-I might result in higher cell turnover in MG63 osteosarcoma cells. Furthermore, in sarcomas or carcinomas showing clinical association to IGF-I levels and malignancy, IGF-I dependent apoptosis and proliferation could be a significant mechanism of malignant tumour growth.

Key words

Osteosarcoma - Insulin-like growth factor-I - Proliferation - Apoptosis - Caspase-3 - Insulin-like growth factor-I receptor - Bad

Volltext

Referenzen

References

1 LeRoith D, Roberts C T. The insulin-like growth factor system and cancer. Cancer Lett. 2003; 195 127-137
2 Gallaher B W, Hille R, Raile K, Kiess W. Apoptosis: live or die - hard work either way!. Horm Metab Res. 2001; 33 511-519
3 Herzlieb N, Gallaher B W, Berthold A, Hille R, Kiess W. Insulin-like growth factor-I inhibits the progression of human U-2 OS osteosarcoma cells towards programmed cell death through interaction with the IGF-I receptor. Cell Mol Biol (Noisy-le-grand). 2000; 46 71-77
4 Gooch J L, Van Den Berg C L, Yee D. Insulin-like growth factor (IGF)-I rescues breast cancer cells from chemotherapy-induced cell death - proliferative and anti-apoptotic effects. Breast Cancer Res Treat. 1999; 56 1-10
5 Holly J M, Gunnell D J, Davey Smith G. Growth hormone, IGF-I and cancer. Less intervention to avoid cancer? More intervention to prevent cancer?. J Endocrinol. 1999; 162 321-330
6 Khatib A M, Siegfried G, Prat A, Luis J, Chretien M, Metrakos P, Seidah N G. Inhibition of proprotein convertases is associated with loss of growth and tumourigenicity of HT-29 human colon carcinoma cells: importance of insulin-like growth factor-1 (IGF-1) receptor processing in IGF-1-mediated functions. J Biol Chem. 2001; 276 30686-30693
7 Ngo T H, Barnard R J, Leung P S, Cohen P, Aronson W J. Insulin-like growth factor I (IGF-I) and IGF binding protein-1 modulate prostate cancer cell growth and apoptosis: possible mediators for the effects of diet and exercise on cancer cell survival. Endocrinology. 2003; 144 2319-2324
8 Firth S M, Baxter R C. Cellular actions of the insulin-like growth factor binding proteins. Endocr Rev. 2002; 23 824-854
9 Raile K, Hoflich A, Kessler U, Yang Y, Pfuender M, Blum W F, Kolb H, Schwarz H P, Kiess W. Human osteosarcoma (U-2 OS) cells express both insulin-like growth factor-I (IGF-I) receptors and insulin-like growth factor-II/mannose-6-phosphate (IGF-II/M6P) receptors and synthesize IGF-II: autocrine growth stimulation by IGF-II via the IGF-I receptor. J Cell Physiol. 1994; 159 531-541
10 Hellawell G O, Turner G D, Davies D R, Poulsom R, Brewster S F, Macaulay V M. Expression of the type 1 insulin-like growth factor receptor is up-regulated in primary prostate cancer and commonly persists in metastatic disease. Cancer Res. 2002; 62 2942-2950
11 Scotlandi K, Avnet S, Benini S, Manara M C, Serra M, Cerisano V, Perdichizzi S, Lollini P L, de Giovanni C, Landuzzi L, Picci P. Expression of an IGF-I receptor dominant negative mutant induces apoptosis, inhibits tumorigenesis and enhances chemosensitivity in Ewing’s sarcoma cells. Int J Cancer. 2002; 101 11-16
12 Papa V, Gliozzo B, Clark G M. et al . Insulin-like growth factor-I receptors are overexpressed and predict a low risk in human breast cancer. Cancer Res. 1993; 53 3736-3740
13 Pennisi P A, Barr V, Nunez N P, Stannard B, LeRoith D. Reduced expression of insulin-like growth factor I receptors in MCF-7 breast cancer cells leads to a more metastatic phenotype. Cancer Res. 2002; 62 6529-6537
14 Guvakova M A, Surmacz E. Overexpressed IGF-I receptors reduce estrogen growth requirements, enhance survival, and promote E-cadherin-mediated cell-cell adhesion in human breast cancer cells. Exp Cell Res. 1997; 231 149-162
15 Hayden J M, Mohan S, Baylink D J. The insulin-like growth factor system and the coupling of formation to resorption. Bone. 1995; 17 93S-98S
16 Johansson A G, Lindh E, Ljunghall S. Insulin-like growth factor I stimulates bone turnover in osteoporosis. Lancet. 1992; 339 1619
17 Ebeling P R, Jones J D, O’Fallon W M, Janes C H, Riggs B L. Short-term effects of recombinant human insulin-like growth factor I on bone turnover in normal women. J Clin Endocrinol Metab. 1993; 77 1384-1387
18 Ferrier J, Xia S L, Lagan E, Aubin J E, Heersche J N. Displacement and translocation of osteoblast-like cells by osteoclasts. J Bone Miner Res. 1994; 9 1397-1405
19 Kawakami A, Nakashima T, Tsuboi M. et al . Insulin-like growth factor I stimulates proliferation and Fas-mediated apoptosis of human osteoblasts. Biochem Biophys Res Commun. 1998; 247 46-51
20 Baserga R. The insulin-like growth factor I receptor: a key to tumor growth?. Cancer Res. 1995; 55 249-252
21 Werner H, Le Roith D. The insulin-like growth factor-I receptor signalling pathways are important for tumorigenesis and inhibition of apoptosis. Crit Rev Oncog. 1997; 8 71-92
22 Yang B C, Chang H M, Wang Y S, Chen R F, Lin S J. Transient induction of apoptosis in serum-starved glioma cells by insulin and IGF-1. Biochim Biophys Acta. 1996; 1314 83-92
23 Parrizas M, LeRoith D. Insulin-like growth factor-1 inhibition of apoptosis is associated with increased expression of the bcl-xL gene product. Endocrinology. 1997; 138 1355-1358
24 Gilmore A P, Valentijn A J, Wang P. et al . Activation of Bad by therapeutic inhibition of epidermal growth factor receptor and transactivation by insulin-like growth factor receptor. J Biol Chem. 2002; 277 27643-27650
25 Fang X, Yu S, Eder A. et al . Regulation of Bad phosphorylation at serine 112 by the Ras-mitogen-activated protein kinase pathway. Oncogene. 1999; 18 6635-6640
26 Hirai I, Wang H G. Survival-factor-induced phosphorylation of Bad results in its dissociation from Bcl-x(L) but not Bcl-2. Biochem J. 2001; 359 345-352
27 Tennant M K, Thrasher J B, Twomey P A, Drivdahl R H, Birnbaum R S, Plymate S R. Protein and messenger ribonucleic acid (mRNA) for the type 1 insulin-like growth factor (IGF) receptor is decreased and IGF-II mRNA is increased in human prostate carcinoma compared to benign prostate epithelium. J Clin Endocrinol Metab. 1996; 81 3774-3782
28 Happerfield L C, Miles D W, Barnes D M, Thomsen L L, Smith P, Hanby A. The localization of the insulin-like growth factor receptor 1 (IGFR-1) in benign and malignant breast tissue. J Pathol. 1997; 183 412-417
29 Schnarr B, Strunz K, Ohsam J, Benner A, Wacker J, Mayer D. Down-regulation of insulin-like growth factor-I receptor and insulin receptor substrate-1 expression in advanced human breast cancer. Int J Cancer. 2000; 89 506-513
30 Yang Y, Hoeflich A, Butenandt O, Kiess W. Opposite regulation of IGF-I and IGF-I receptor mRNA and concomitant changes of GH receptor and IGF-II/M6P receptor mRNA in human IM-9 lymphoblasts. Biochim Biophys Acta. 1996; 1310 317-324

K. Raile, M. D.

University Hospital for Children and Adolescents

Oststr. 21-25 · D-04317 Leipzig · Germany

Telefon: + 49 (341) 9726068

Fax: + 49 (341) 9726009

eMail: klemens.raile@medizin.uni-leipzig.de