Aim: The main goal of this study was to evaluate the difference in energy status of MRC-5 cell spheroids of different sizes and with different availability of nutrients. Study design: In vitro study - controlled experiment Materials and methods: During the formation of cell spheroids, the selected, commercially available cell line MRC-5 (Medical Research Council cell strain 5; CCL-171™) was used. Cells were cultured in wells with faintly adhering base to form cell spheroids. One set of spheroids was cultured with a regularly change of culture medium, while the other set of cell spheres was cultured without medium change. Spheres were collected after the 8th and 13th days of cultivation, followed by the determination of LDH enzyme proportion, mitochondrial activity and colorimetric determination of ATP concentration. Results: No significant difference was recorded in the proportion of LDH between the spheres grown for 8 and 13 days, with and without medium change. The proportion of LDH of about 0,4 is visible in spheres with regular medium changes, regardless of incubation time. Spheres cultured under nutrient-reduced conditions reflect greater sensitivity with an increased proportion of cells with compromised cellular integrity. Mitochondrial activity and ATP concentration were recorded to be equal between groups of spheres without statistical significance. Conclusion: The availability of nutrients during the cultivation and maintenance of the culture of MRC-5 cell spheroids leads to a change of the released LDH into the cell medium without changes in the mitochondrial potential and the ATP concentration between the set of spheres.