Insulin-like Growth Factor-1 (IGF-1) regulates pheochromocytoma cellular proliferation in vitro and in vivo.

FERNANDEZ, MA. CELIA; VENARA MARCELA; NOWICKI SUSANA; ALVAREZ SEDO, CRISTIAN; CHEMES HECTOR; BARONTINI MARTA; PENNISI PATRICIA

Boston

Congreso; Reunion y Exposicion Anual, Sociedad de Endocrinologia (ENDO); 2011

We have previously shown that IGF-1R expression is increased in malignant compared to benign pheo suggesting a role of IGF/IGF-1R in this disease. We demonstrated that Mouse Pheochromocytoma Cells (MPC) expressed a functional IGF-1R thus we injected MPC in non immunodeficient mice to generate a pheo model. Aim: In order to understand the influence of IGF/IGF-1R on the biology of pheo, we evaluated the effects of IGF-1 in vitro and in vitro. Materials and Methods: MPC cells were grown in 0.5%FBS and 1%HS RPMI medium to assess proliferation and apoptosis with or without 50nM rhIGF-1. Cells were counted on days 1, 3, 5, 7 after seeding. On day 5 we performed BrdU incorporation and inmunofluorescence for cleaved caspase 3 and TUNEL. We used soft agar and Boyden chambers to study the effect of IGF-1 on cell’s colony formation and migration. For in vivo studies, 8 w/o liver IGF-1 deficient (LID) mice (n=25) and control (n=30) mice were injected sc (right flank, 106 MPC) and monitored for tumor development. A group of LID mice (n=7) were injected twice daily i.p. with 2 mg/kg of rhIGF-1 starting on the day of MPC injection. Results: In Vitro, MPC cells proliferation was blunted in basal condition and significantly increased upon rhIGF-1 stimulation. BrdU incorporation increased under IGF-1 stimulation and the percentage of apoptotic cells was significantly lower with rhIGF-1. MPC cells were able to form colonies in soft agar and rhIGF-1 stimuli increased the number and size of colonies. MPC cells migration was also stimulated by IGF-1. In Vivo, all control mice developed tumor between 5-6w post-cell inoculation. In contrast, 7/18 LID mice had tumor from 6-11w leaving a 61.1% of LID mice free of tumor by week 14. All LID mice treated with rhIGF-1 developed tumor between 5-6w. The latency period was shorter in control and LID+IGF-1 mice than in LID mice. The risk of developing tumor was 10 times lower for LID mice (HR 0.1 95%CI:0.02-0.27) and rhIGF-1 treatment increased this risk 8 times (HR 7.98 95%CI:1.60-67.06). Conclusions: We demonstrate that IGF-1 exacerbates in vitro MPC tumorigenic characteristics reducing apoptosis and increasing proliferation, migration and cell’s ability to grow unattached. In vivo, lower levels of IGF-1 diminish tumor incidence and lengthen the latency period. IGF-1 treatment of LID mice enhanced tumor development similarly to control mice. Therefore, experimental pheo development is regulated by the levels of circulating IGF-1 in mice.