Contact effects by mitochondria: biological destruction of cultivated B16 - F10 melanoma cells

In recent years therapeutic effects produced by mitochondria, transplanted to pathogenic regions in an organism, which demonstrate their high migration activity and tropicity with respect to filling of energetic vacuum, have been inspiring a renewed interest in the scientific community. In this context, this raises the question of the source of metabolically active mitochondria and their histological compatibility required for the mitochondrial transplantation. Aim. The aim of our research work is to study characteristics/ quality of mitochondria from cells of the liver and the heart in rats, which in the in vitro system have produced their impact on different biological features of the B16-F10 murine melanoma cell culture. Materials and methods. In our research work we have used cells of the B16-F10 murine melanoma cell line culture. In the framework of the study, an experiment with mitochondria harvested from the liver and the heart of a rat has been conducted. Mitochondria have been isolated using differential centrifugation with a high-speed refrigerated centrifuge. With the B16-F10 culture cells, we have designed the following variants of our experiments: 1) use of cardiac mitochondria (1 mg/mL, in terms of total protein); 2) use of cardiac mitochondria (1 mg/mL) + succinic acid (10-4%); 3) mitochondria of the liver (1 mg/mL, in terms of total protein); 4) use of mitochondria of the liver (1 mg/mL) + succinic acid (10-4%); 5) use of succinic acid (10-4%) solely; 6) the reference specimen with no use of mitochondria and the above agents. An assessment of the impact made by mitochondria on the migration of the B16-F10 cells has been performed with the scratch wound healing test. For the purpose of an analysis of the effect produced by mitochondria on the energetic metabolism of the B16-F10 cell culture we have measured main parameters of the cell respiration and glycolysis in stress tests with adding some toxic chemicals. The rate of the cellular respiration has been assessed by measuring the amount of oxygen taken in (oxygen consumption rate, OCR), and the glycolysis level has been evaluated by the extracellular acidification rate (ECAR). Results. Adding cardiac and hepatic mitochondria to the cultivated B16-F10 cells has produced a pronounced cytopathic effect, which has become more remarkable upon expiration of two days of the cell cultivation and which has consisted in cytoplasm granulation and partial detaching of the cells. Introducing mitochondria of the heart to the cultivated B16-F10 cells has induced a considerable decrease both in the background-related and the maximum level of oxygen consumption by the B16-F10 cells as against the reference samples without adding of mitochondria. Adding the cardiac mitochondria has led to a statistically significant decrease in the base level of ECAR by 14,36 mpH/min (t = 3,12, df = 10) as compared with the reference values. Introducing hepatic mitochondria has also resulted in a reduction of the average value of ECAR as against the background by 4,8 mpH/min. Conclusion. Metabolically active mitochondria are capable of reformatting energetic fluxes in tumor cells and change their cellular respiration that leads to the most effectively realized death of the cultivated B16-F10 tumor cells.