Mitochondrial Defects in cw-Diamminedichloroplatinum(II)-resistant Human Ovarian Carcinoma Cells1

We have been studying the membranes of cisplatin (DDP)-resistant 2008 human ovarian carcinoma cells (C13* cells) for alterations that may account for their decreased DDP accumulation. We now report that C13* cells have significant changes in their mitochondrial and plasma mem brane potentials and in their mitochondrial morphology. C13* cells accumulated 2.0 ±0.1-fold more of the membrane potential marker |'l I] tetraphenylphosphonium cation (TPP*) than sensitive cells. In high K+ medium, which depolarizes the plasma membrane but not the mitochon drial membrane, |'lI] I l'I" accumulation was still 2.3 ±0.1fold greater in resistant cells, indicating that the mitochondrial membrane potential was higher. In the presence of car bonyIcyanide /Mrifluoromethoxyphenyl hydrazone, which depolarizes the mitochondrial membrane but not the plasma membrane, | '11]1PI>+accumulation demonstrated that the plasma membrane potential in C13* cells was elevated as well. These elevations were also present in C8 cells with low-level DDP resistance. After ouabain treatment, exposure to nigericin stimulated | 'I l|TIT+ accumulation 3fold in sensitive cells but had no effect in C13* cells, indicating either that: (a) the mitochondrial pH gradient was minimal; or (b) the mito chondrial electric potential was already at a maximal level in C13* cells. Fluorescence microscopy of living cells stained with the mitochondriaspecific dye rhodamine 123 revealed that resistant cells had significant changes in their mitochondrial morphology. Electron microscopy also revealed major alterations in the cristae structure. The C13* cells, which were approximately 15-fold resistant to DDP, were S-fold hypersensitive to the mitochondria! poison rhodamine 123. We conclude that these DDP-resistant 2008 cells have an elevated plasma membrane potential and alterations in their mitochondria as indicated by their membrane potential, morphology, and sensitivity to mitochondrial poisons. These results imply that mitochondria play an important role in the cellular pharmacology of DDP.