Biomedical subjects
Publications and source records attributed to T E Novikova.
Synchronous changes were found in the intracellular ATP content and in the ratio of "dark" and "light" cells in the monolayer culture of hepatocytes. An attempt has been made to change these ratios by the effect of hormones. The rhythmical processes in the culture are interpreted as a universal cell property.
Incorporation of 3H-leucine in proteins and radioactivity of free leucine have been studied in hepatocyte culture on collagen-coated glass. A high density culture obtained from a suspension of isolated rat hepatocytes at 5 x 10(6) cells/ml was compared with low density cultures diluted from the same suspension 10- to 20-fold. The incorporation pool ratio showing the efficiency of the precursor utilization was higher in the low density culture as compared with the high density monolayer. The effect was due to disproportionate low pool in the diluted cultures. The total radioactivity of proteins was lower in the diluted cultures as compared with the monolayer, although the incorporation was not in proportion with the cell density. Specific rate of protein synthesis (per cell) was higher in the diluted cultures than in the monolayer.
In a culture of rat hepatocytes, kept in a serum-free medium 199, the number of attached cells was decreased and a monolayer was not formed within 24 h. Soon after the cells were attached to collagen, the intensity of protein synthesis in the serum-free medium was decreased due to the smaller number of cells in the culture and was calculated per cell. The properties of cultures, i.e., the number and shape of cells, external appearance of the culture, and intensity of protein synthesis, were similar in a serum-containing medium and in the serum-free medium 199 complemented with 0.2 mg/ml albumin and 5 micrograms/ml insulin. A monolayer typical for the medium with serum was formed in the medium with albumin and insulin within approximately 20 h after attachment of the hepatocytes to the substrate.
The kinetics of protein synthesis was studied in the cultures of rat hepatocytes on slides covered with collagen or fibronectin and kept in a serum-free medium 199 complemented by albumin and insulin. No protein synthesis rhythm was found in the cultures with markedly separated cells obtained from a diluted suspension of hepatocytes ("sparse cultures"). After 0.3-0.4 microM total gangliosides were added to the medium with such cultures, the rhythm was found in all experiments. Such an effect was earlier found under the influence of a medium conditioned by a monolayer on the sparse cultures. The results obtained suggest the involvement of gangliosides in synchronization of oscillations of the protein synthesis intensity and call for studies of the conditioned medium properties.
We studied the kinetics of proteins synthesis in the cultures of hepatocytes on collagen-coated slides in medium 199 enriched with 0.2 mg/ml albumin and 0.5 microgram/ml insulin and devoid of bovine serum. Circahoral fluctuations of proteins synthesis intensity were found in the monolayer cultures and sparse cultures in a conditioned medium. No protein synthesis rhythm was expressed in a fresh medium after repeated washing of the cultures. Addition of micromolar concentrations of gangliosides GM1 or GD1a to the medium revealed the rhythm in washed cultures. Addition of GT1b or GM3 to the medium did not synchronize the oscillations: the kinetics of protein synthesis did not differ from that in the control space cultures in a fresh medium without exogenous gangliosides. Accumulation of gangliosides GM1 and GM3 in the hepatocytes in vitro upon conditioning of a serum-free culture medium was shown using the indirect immunocytochemical method. The effect was found in dense monolayer cultures and in cultures with separated cells in a conditioned medium. The protein synthesis rhythm was found in such cultures. Gangliosides are weakly expressed in most cells of a repeatedly washed 24-hour monolayer and washed sparse cultures. No protein synthesis rhythm was found in washed cultures. Similar changes in the dynamics of the culture medium conditioning, accumulation of gangliosides in cells and rhythmic activity of cells population confirm the concept of the synchronizing role of gangliosides.