Fluorescence imaging of mitochondrial responses to glucose challenge and probes in mitochondrial DNA-deficient osteosarcoma cells

E Kohen 1, JG Hirschberg 2, C Ornek 1, M Monti 2, JP Berry 3

Departments of Biology 1, Physics 2, and Chemistry3, University of Miami, Coral Gables, FL 33124, USA

Wild-type osteosarcoma 143B and mutant DNA-deficient osteosarcoma 1436rho cells were studied by fluorescence imaging at 360nm and 436nm excitation. With the vital probe Mitotracker Green, real-time visualization was possible in the wild-type, while in the DNA-deficient cells, the accumulation of 30 or more scans was required. In the rho-type cells, broad tentacle-like processes appeared which contained mitochondria-shaped granules loaded with Mitotracker Green. Generally, two mitochondrial morphologies were seen: filamentous or granular. NADH response was triggered by a glucose challenge. Within seconds, the mitochondria responded by increased NADH fluorescence which appeared first in a few scattered clusters, and very soon spread to the rest of the mitochondria with increased intensity. Despite the low quantum yield of the NADH fluorescence and the rapidity of the response, the ability to image the fluorescence increase and its spread pattern at the limit of sensitivity is noteworthy. It indentifies the osteosarcoma cells as particularly suitable for the study of mitochondrial metabolic activation. The method's further development by application of Fourier interferometry for fluorescence excitation imaging will be discussed. These osteosarcoma cells are a good model for the extension of the method in diagnostics of other malignant cells, and potentially for therapy.

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Paper presented at the International Symposium on Predictive Oncology and Intervention Strategies; Paris, France; February 9 - 12, 2002; in the section on Diagnostic Markers.