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Measurement of oxygen diffusion distance in tumor cubes using a fluorescent hypoxia probe
P L Olive1, C Vikse, M J Trotter
1Medical Biophysics Unit, British Columbia Cancer Research Centre, Vancouver, Canada.
Abstract:
Oxygen diffusion distance was measured in solid tumor "cubes" prepared by excising the tumor from the mouse and incubating 1-2 mm sided tumor cubes in spinner culture flasks with fluorescent drugs (AF-2 or DM113) which bind to hypoxic cells. After incubation, frozen sections were prepared and examined for AF-2 or DM113 binding using a fluorescence image processing system. Alternatively, cubes were stained with a slowly penetrating fluorescent dye, Hoechst 33342, prior to disaggregation and measurement of AF-2 binding using flow cytometry. The distance from the cube surface to the AF-2 stained region (i.e., the thickness of the unstained rim) was used as an indication of oxygen diffusion distance, which depends on the square root of the tumor oxygen consumption rate. Oxygen diffusion distance was dependent on tumor type, external oxygen concentration, and temperature during incubation with AF-2, but was unaffected by tumor size up to 1.2 gm or position of the cube within the tumor. Oxygen diffusion distances (in planar geometry) measured for cubes prepared from SCCVII, RIF-1, Lewis lung or WiDr tumors were 107, 123, 153 and 193 microns, respectively. For SCCVII and WiDr tumors, the percentage of blood vessels separated by a distance greater than double the mean oxygen diffusion distance was used as an indication of the hypoxic fraction.
Insights
This study measured oxygen diffusion distance in solid tumors using fluorescent drugs. Results show diffusion distance varies by tumor type and incubation conditions, impacting tumor hypoxia.
Area of Science:
- Biomedical Engineering
- Oncology
- Cancer Research
Background:
- Solid tumors often contain hypoxic regions due to inadequate oxygen supply.
- Tumor hypoxia is linked to treatment resistance and poor patient outcomes.
- Accurate measurement of oxygen diffusion distance is crucial for understanding tumor microenvironments.
Purpose of the Study:
- To quantify oxygen diffusion distance in solid tumor models.
- To investigate factors influencing oxygen diffusion, such as tumor type and incubation conditions.
- To establish a method for assessing tumor hypoxia based on oxygen diffusion.
Main Methods:
- Solid tumor cubes were incubated with fluorescent drugs (AF-2, DM113) that bind to hypoxic cells.
- Fluorescence imaging and flow cytometry were used to measure drug binding and determine oxygen diffusion distance.
- Hoechst 33342 staining was employed as an alternative method to assess hypoxia.
Main Results:
- Oxygen diffusion distance varied significantly between tumor types (e.g., SCCVII: 107 µm, WiDr: 193 µm).
- Diffusion distance was influenced by external oxygen concentration and incubation temperature.
- Tumor size (up to 1.2 gm) and position within the tumor did not affect diffusion distance.
Conclusions:
- The developed method accurately measures oxygen diffusion distance in solid tumors.
- Tumor type is a critical determinant of oxygen diffusion and potential hypoxia.
- These findings contribute to a better understanding of tumor oxygenation and hypoxia assessment.