OXYGEN DIFFUSION DISTANCE AND DEVELOPMENT OF NECROSIS IN MULTICELL SPHEROIDS

The multicell spheroid in vitro tumor model was used to study the effects of diffusion limitations of metabolites on the development of necrosis in a tumor-like environment. Chinese hamster V79-171b cells were grown in suspension as multicellular spheroids in different oxygen or glucose concentrations to diameters where they developed central necrosis surrounded by an outer rim of viable cells. The range of oxygen concentrations used in the gas phase was 1 to 20.3%, in equilibrium with complete nutrient medium. For each oxygen concentration the thickness of the rim of viable cells was determined from measurements of histological sections. The square of the thickness of the viable rims of spheroids grown at different oxygen concentrations between 5 and 20.3% increased linearly with the theoretically derived oxygen diffusion distance, showing that the oxygen diffusion distance is a main factor controlling, either directly or indirectly, the development of necrosis in this concentration range. The data were interpreted as meaning spheroids grown at more than 5% O2 contain a very small proportion of cells which were severely hypoxic or anoxic. The viable rim thickness was relatively less affected during growth at less than 5% oxygen, implying that an unknown mechanism controls the formation of necrosis when the oxygen diffusion distance is less than approximately 80 μm. Although alterations in the glucose concentration had some effect on viable rim thickness at these low oxygen concentrations, the effect was much less than expected if glucose alone were responsible for controlling viability in this outer region.