The role of hypoxia-inducible factor-1 in three-dimensional tumor growth, apoptosis, and regulation by the insulin-signaling pathway

• Published in: Cancer research (Chicago, Ill.) Vol. 65; no. 10; pp. 4147 - 4152
• Main Authors: Leek, Russell D, Stratford, Ian, Harris, Adrian L
• Format: Journal Article
• Language: English
• Published: United States 15.05.2005
• Subjects: Animals; Apoptosis - drug effects; Apoptosis - physiology; Cell Growth Processes - physiology; Cell Line, Tumor; Chromones - pharmacology; DNA-Binding Proteins - drug effects; DNA-Binding Proteins - physiology; Enzyme Inhibitors - pharmacology; Hypoxia-Inducible Factor 1; Hypoxia-Inducible Factor 1, alpha Subunit; Insulin - pharmacology; Liver Neoplasms, Experimental - pathology; Mice; Morpholines - pharmacology; Nuclear Proteins - drug effects; Nuclear Proteins - physiology; Phosphatidylinositol 3-Kinases - antagonists & inhibitors; S Phase - physiology; Signal Transduction - drug effects; Signal Transduction - physiology; Spheroids, Cellular; Transcription Factors - drug effects; Transcription Factors - physiology
• ISSN: 0008-5472; 1538-7445
• DOI: 10.1158/0008-5472.can-04-2184

The purpose of this study was to establish the effect of hypoxia-inducible factor-1 (HIF-1) directly on tumor growth, independently of angiogenesis. This was done by growing wild-type mouse hepatoma cells (HEPA-1) and their HIF-1-deficient counterpart C4 as multicellular tumor spheroids and quantifying differences in growth rates and proliferative and apoptotic indices. Insulin and insulin-like growth factor-I are key growth factors, also able to regulate hypoxia-responsive genes via HIF-1; thus, the effects of insulin on this model were also investigated. Two-dimensional growth was serum dependent and no difference was seen between wild-type HEPA-1 and C4 cell growth profiles, but major differences were seen in three-dimensional growth. HIF-1 supported spheroid growth under hypoxia as the HEPA-1 spheroids grew faster than the C4. Surprisingly, the HIF-1-deficient cells had a higher proliferation rate in three-dimensional growth (C4 mean S-phase index, 13.6%; HEPA-1 mean S-phase index, 9%; P = 0.009) that was associated with an inhibition of apoptosis. However, the apoptosis rate was much greater in these spheroids (C4 mean apoptotic index, 6.4; HEPA-1 mean apoptotic index, 0.78%; P = 0.0006). Addition of insulin increased proliferation and apoptosis in both HEPA-1 and C4 spheroids, demonstrating an HIF-1-independent effect of insulin signaling in three-dimensional growth. These results indicate that the enhancing effect of HIF-1 in three-dimensional tumor growth is a balance of both reduced proliferation and enhanced survival, the latter being proportionally greater.