Increased Rhodamine 123 Uptake by Carcinoma Cells1

The total cellular content of the fluorescent mitochondrialspecific dye rhodamine 123 (Rh-123) was quantified by butanol extraction as a function of time of exposure and dose for a variety of cell lines. These results were compared with observa tions made by fluorescence microscopy on dye localization and mitochondrial morphology. There appeared to be two categories of cell types based on Rh-123 uptake: those which progressively accumulate the dye, such as Ehrlich ascites tumor cells, carci noma-derived lines MCF-7, PaCa-2, EJ, HeLa, and normal fibroblast line CCL 64; and those which appear to equilibrate with the extracellular dye within 1 h of incubation in Rh-123 (1 M9/ml) with a minimal level of uptake, such as the normal epithelialderived lines CV-1 and MDCK and the transformed fibroblast line 64F3. Within the first category, the absolute value of uptake per cell correlated with the concentration of Rh-123 in the medium and with the period of exposure to the dye up to a point of apparent cellular saturation. The length of time required for apparent saturation depended on the cell type. In the second category equilibration was very early, and the total uptake was a function of the extracellular concentration of Rh-123. This probably does not represent a saturation level of dye content in the non-accumulating, low uptake cell lines. Fluorescence mi croscopy revealed that Rh-123 localization was initially mitochondrial-specific for all of the cell lines examined. Over time, alterations in mitochondrial morphology and cytoplasmic fluores cence were observed in the high uptake cell lines but not in the minimal uptake cell lines. Incubation of the high uptake HeLa cell line with the mitochondrial membrane potential inhibitor ptrifluoromethoxyphenylhydrazone substantially decreased Rh123 uptake. These observations may indicate a transformationrelated characteristic of carcinoma cell mitochondria. It may be possible to exploit the mechanism responsible for the progres sive accumulation of Rh-123 by carcinoma-derived cell types for chemotherapeutic approaches to certain types of carcinomas.