Development of multicellular spheroids of HeLa cells cocultured with fibroblasts and their response to X-irradiation.

HeLa cells cocultured with human diploid fibroblasts developed a more stable and tightly packed spheroid than did HeLa cells alone. Mature spheroids with central necrosis resembled the cord structure of solid tumors in histological appearance and distribution of cells labeled by [3H]thymidine. Electron microscopy showed that HeLa cells in the spheroid are in close contact with each other and have abundant intercellular junctions. The rate of cell loss from spheroid surface, expressed as cells lost per sq mm of spheroid surface per hr, was twice as high for spheroids of HeLa cells alone as for cocultured spheroids. The radiation dose-survival curve for cells in small spheroids without central necrosis was characterized by a wide shoulder than that for cells grown in monolayer or that for single-cell suspension from the same batch of the spheroids, but the terminal slope was similar for all survival curves. Development of central necrosis in larger spheroids coincided with a further increase in radioresistance, and the terminal D0 in the survival curve was 3 times that of single-cell suspension from the spheroids of the same batch. The radioresistant tail in the survival curve, however, disappeared when the spheroids were irradiated in medium containing a hypoxic cell radiosensitizer, misonidazole. These findings suggest that spheroids of HeLa cells cocultured with fibroblasts simulate the cord structure of solid tumors not only in structure but also in response to radiation.