Control and characterization of oxygen concentration in tissue culture

Authors

• MH Ronzetti
• TW Sanchez
• B Baljinnyam
• AE Owens
• M Antony
• T Voss
• E Wallgren
• D Talley
• K Balakrishnan
• SE Leyes Porello
• G Rai
• JJ Marugan
• SG Michael
• B Baljinnyam
• N Southall
• C Klumpp-Thomas
• A Simeonov
• MJ Henderson

Abstract

The oxygen concentration experienced by cells plays a crucial role in controlling metabolic pathways and determining culture health, yet delivery of oxygen to cells in culture is often poorly characterized. This can have significant impacts on the relevance of culture models to physiological and pathophysiological environments in vivo; for example, hypoxia in cancer is linked to drug resistance and metastasis. Although the ambient atmosphere in which most tissue culture is performed has an oxygen concentration much higher than physiological levels, the microenvironment of the cells typically experiences substantial depletion due to limits of oxygen transport, primarily diffusive, through the media and, in the case of 3D culture, through the surrounding matrix. The balance between source concentration, transport to the cells, and consumption is what determines the intracellular oxygen concentration, which can therefore depend heavily on the density and location of the cultured cells. We will present work on several aspects of this problem: 1) development of a scalable bioreactor system which controls oxygen delivery to 3D culture in a multiwell plate format, using an oxygen permeable membrane with patterned micropillars to mimic a capillary bed; 2) measurement of oxygen concentrations in vitro in cells transfected to express a novel FLIM-FRET probe, which uses the change in myoglobin absorption spectrum induced by oxygen binding to sense intracellular oxygen levels; 3) finite element modeling aimed at understanding the interplay of delivery, transport, and consumption in determining in vitro oxygen concentrations.

Scientific Focus Area: Biomedical Engineering and Biophysics

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