(12c) Cellular Response to Gradients in Oxygen Concentration through 3D Hydrogel Scaffolds: a Correlation Between Oxygen Concentration and HIF-1alpha Expression in Tumor Cells | AIChE

(12c) Cellular Response to Gradients in Oxygen Concentration through 3D Hydrogel Scaffolds: a Correlation Between Oxygen Concentration and HIF-1alpha Expression in Tumor Cells

Authors 

Acosta, M. A. - Presenter, University of Maryland - Baltimore County
Ostrand-Rosenberg, S. - Presenter, University of Maryland - Baltimore County
Leach, J. B. - Presenter, University of Maryland Baltimore County


Oxygen diffusion through tissues is a critical factor in maintaining healthy engineered tissues in vitro. Therefore, understanding the cellular response to changes in soluble cues through their biomaterial microenvironment, such as oxygen concentration, is crucial for engineering materials that can more closely mimic conditions in vivo and that can potentially lead for improving methods for control over cell behavior in tissue engineering. Few methods exist for direct correlation of spatial changes in oxygen concentration through biomaterial scaffolds and the impact that they have on cellular function. For this purpose, we have developed oxygen-sensing microparticles that are suspendable through the volume of any transparent biomaterial scaffold used in cell culture and tissue engineering. Additionally, ratiometric measurements of oxygen concentration can be obtained in a non-invasive manner, as operation of the microparticles is based on dynamic fluorescence quenching of a ruthenium based organic-metal complex. Cell response to hypoxia is primarily controlled by the hypoxia inducible factor-1 (HIF-1). The degradation process of its alpha subunit, the hypoxia inducible factor-1alpha (HIF-1alpha), is directly regulated by oxygen concentration. Thus, HIF-1alpha is an ideal marker for correlating cellular response to oxygen concentration in engineered tissues. We have created two cell lines transfected to express an EGFP-HIF-1alpha fusion: 4T1 murine mammary carcinoma and C8161 human subcutaneous melanoma cells. Using microscopy, expression of the fusion protein and oxygen concentration can be monitored simultaneously, allowing us to construct a direct correlation between the two. In this study, we present the characterization of the two transfected cell lines by demonstrating that differential expression of measurable quantities of the EGFP-HIF-1alpha fusion is achieved with prolonged exposure to hypoxia. We also present the construction of a correlation between changes in the expression of the EGFP-HIF-1alpha fusion protein and gradients in oxygen concentration through the volume of hydrogel scaffold.

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