Fluorimetric Evaluation of the Effect of Malic, Succinic and Ascorbic Acids on the Growth Properties of A549 Cells in Culture

Abstract

Relevance: A number of researchers consider the study of the radioprotective properties of non-toxic or low-toxic natural substances to be a promising direction. A special place among them is occupied by antioxidants and participants in the basic reactions of metabolism. In order to avoid methodological errors when performing these studies, it is necessary to conduct a number of additional experiments. For example, in order to study the properties of various substances on cell cultures using tablet readers, it is first necessary to make sure that these substances do not affect the ability of cells to adsorb to the bottom of the wells of the tablet and do not interfere with cell proliferation. And if such an influence is detected, further experiments with these substances should be planned taking into account the information received. Purpose: To search the effect of ascorbic, malic and succinic acids on the ability of lung adenocarcinoma cells (A549) to adhere in a 96-well plate, followed by the onset of proliferation by fluorescence registration method using Hoechst-33342 fluorophore. Methodology: The experiment was carried out in a 96-well tablet. The working concentration of Hoechst-33342 was 1 μg/ml (1.62 μM). Fluorescence was recorded at a wavelength of 460 nm when the samples were excited by light with a wavelength of 355 nm. In an experiment to study the effect of ascorbate, malate and succinate on cell adhesion and proliferation, 20,000 cells and a solution of one of these substances in a working concentration of 2 mM were introduced into the cells of the tablet. The number of cells in the wells was estimated based on the fluorescence of Hoechst-33342 after a day of incubation. Result: In samples containing 2 mM succinic acid and ascorbic acid, a statistically significant decrease in the intensity of fluorescence was observed compared with a sample that did not contain the drug. This suggests that these compounds negatively affect the growth properties of the A549 culture: they inhibit cell adhesion or slow down their proliferation. Scope of the results and conclusions:The results obtained are necessary for the methodologically correct planning of the most detailed studies on the A549 cell line model using fluorescent methods, including studies on the radioprotective properties of ascorbate, malate and succinate under the influence of rare ionizing and neutron radiation.