Products of cells cultured from gliomas. V. Cytology and morphometry of two cell types cultured from glioma

Explants of cells of a human glioma were evaluated with the nuclear fluorochrome 4',6-diamidino-2-phenylindole, by phase-contrast illumination, and by Giemsa staining correlated with double immunofluorescence for glial fibrillary acidic protein (GFAP) and fibronectin (FN). FN-postive (FN±) cells lacked GFAP detectable by immunofluorescence. Their mean nuclear-to-cytoplasmic ratio was large (0.192). Actual mean areas of nuclei (1,252 μm²) and cytoplasm (8.376 μm²) of FN⁺ cells compared with mean areas of fibroblasts suggested that the high nuclear-to-cytoplasmic ratio of FN⁺ cells was due to their microscopically evident reduced cytoplasmic spreading rather than to larger nuclei. Some FN⁺ cells showed marked variation in nuclear and nucleolar size and shape. Others had abnormal mitoses or hyperchromatic nuclei. GFAP-positive (GFAP⁺) cells lacked FN detectable by immunofluorescence. GFAP⁺ cells were smaller and less round than FN⁺ cells. Their usual location was growing on a layer of FN⁺ cells. The mean nuclear-to-cytoplasmic ratio (0.245) of GFAP⁺ cells was the highest in the study, surpassing the ratio of the continuous glioma line LM (0.176). Mean areas of nuclei (289 μm²) and of cytoplasm (1,350 μm²) of GFAP⁺ cells suggested that their high nuclear-to-cytoplasmic ratio was due to their microscopically evident reduced cytoplasmic spreading. Reduced spreading was associated with extension of long, thin cytoplasmic processes. The majority of GFAP⁺ cells showed marked cytoplasmic basophilia, nuclear hyperchromasia, and clumped chromatin. Features observed in both FN⁺ and GFAP⁺ cells from this high-grade astrocytoma are features associated with malignant transformation in more thoroughly studied tumor systems.