Elevated Interstitial Pressure Enhances Hypoxia-Induced VEGF Expression Via HIF-1 in Daoy Medulloblastoma Cells.
Hua Zhong, Ruoxiang Wang, Jonathan W Simons. New York University, NY; Emory University, Atlanta
Background: Tissue hypoxia, increased interstitial pressure and edema are common findings in solid tumor. Hypoxia activates transcription factor HIF-1 and regulates the expression of HIF-1-target genes including the gene that encodes VEGF, a critical factor underlying angiogenesis and interstitial edema. HIF-1α, the dominant subunit of HIF-1, is regulated by oxygen-dependent and –independent ways. We have developed a module to test our hypothesis that increased interstitial pressure enhances hypoxia-induced VEGF expression via HIF-1.
Design: As an alternative to an inflexible hypoxia chamber, we designed a pliable hypoxia chamber to compare cellular responses to three tissue culture environments including conventional CO2 culture, and hypoxia cultures (1% O2/5% CO2/balanced N2) with or without elevated interstitial pressure. The interior pressure within the chambers were monitored by water-mercury manometers. The pliable hypoxia chamber was 80% filled to mimic hypoxia setting without elevated interstitial pressure. The interior pressure remained at 0 mmHg within 24 hours at 37oC. The inflexible hypoxia chamber was fully filled so as to create an intermediate (40 mmHg) or a high (70 mmHg) interior pressure. After 24-hour at 37oC, the interior pressure was elevated from 40 mmHg to 60 mmHg (intermediate) and from 70 mmHg to 120 mmHg (high), respectively.
Results: The new pliable hypoxia chamber was functional based on our design. Due to transparent and flexible, it could be adjusted to a size which allowed us to periodically record, microscopiclly in real time, the nuclear translocation of GFP-HIF-1α and the endothelial tube formation following continuous exposure to hypoxia. In Daoy cells cultured at 37oC for 24 hours, the intermediate pressure enhanced the hypoxic induction of HIF-1α, HIF-1 transcriptional activity and VEGF production. However, high pressure did not affect HIF-1 but resulted in growth inhibition. Finally, the enhanced signal transduction activity upstream of HIF-1α synthesis was shown in cells under hypoxia with intermediate pressure as compared to those in conventional culture or regular hypoxia culture without elevated interstitial pressure.
Conclusions: Intermediate interstitial pressure augments hypoxia-induced VEGF expression through HIF-1. Increased HIF-1α expression likely involves increased protein synthesis in response to moderate pressure stress. Our results could explain the edematous change seen in pathological conditions such as solid tumors, especially those which are situated within a confined space.
Wednesday, March 2, 2011 9:30 AM
Poster Session V # 237, Wednesday Morning