APOPTOSIS, ENERGY-METABOLISM, AND FRACTION OF RADIOBIOLOGICALLY HYPOXIC CELLS - A STUDY OF HUMAN-MELANOMA MULTICELLULAR SPHEROIDS

The magnitude of the fraction of radiobiologically hypoxic cells in tu mours is generally believed to reflect the efficiency of the vascular network. Theoretical studies have suggested that the hypoxic fraction might also be influenced by biological properties of the tumour cells. Quantitative experimental results of cell energy metabolism, hypoxia- induced apoptosis, and radiobiological hypoxia are reported here. Huma n melanoma multicellular spheroids (BEX-c and WIX-c) were used as tumo ur models to avoid confounding effects of the vascular- network. Radio biological studies showed that the fractions of hypoxic cells in 1000- mu m spheroids were 32 +/- 12% (BEX-c) and 2.5 +/- 1.1% (WIX-c). The s pheroid hypoxic volume fractions (28 +/- 6% (BEX-c) and 1.4 +/- 7% (WI X-c)), calculated from the rate of oxygen consumption per cell, the ce ll packing density, and the thickness of the viable rim, were similar to the fractions of radiobiologically hypoxic cells. Large differences between tumours in fraction of hypoxic cells are therefore not necess arily a result of differences in the efficiency of the vascular networ k. Studies of monolayer cell cultures, per-formed to identify the biol ogical properties of the BEX-c and WIX-c cells leading to this large d ifference in fraction of hypoxic cells, gave the following results: (1 ) WIX-c showed lower cell surviving fractions after exposure to hypoxi a than BEX-c, (2) WIX-c showed higher glucose uptake and lactate relea se rates than BEX-c both under aerobic and hypoxic conditions, and (3) hypoxia induced apoptosis in WIX-c but not in BEX-c. These observatio ns suggested that the difference between BEX-c and WIX-c spheroids in fraction of hypoxic cells resulted partly from differences in cell ene rgy metabolism and partly from a difference in capacity to retain viab ility under hypoxic stress. The induction of apoptosis by hypoxia was identified as a phenomenon which has an important influence on the mag nitude of the fraction of radiobiologically hypoxic cells in multicell ular spheroids.