Oxidative stress modeling in the blood lymphocytes in vitro to examine the Jurkat line tumor cells apoptosis

Abstract

Aim. To experimentally select the optimum concentration of hydrogen peroxide capable of efficiently induce oxidative stress and launch the programmed death of the maximum number of lymphocytes, but not induce the necrosis. Methods. Jurkat tumor cell line (human T-lymphoblastic leukemia) lymphocytes isolated from the blood of healthy donors (15 males, 18 females) aged 18 to 25 years were the objects of the study. To confirm the object of study, blood cells typing for CD5 using flow cytometry was performed. To model the oxidative stress in vitro, blood lymphocytes were incubated in the presence of hydrogen peroxide at a final concentration of 0.3, 0.5, 1.0 and 2.0 mM. Reduced and oxidized glutathione levels estimation, the ratio between the fractions, and the level of reactive oxygen forms in lymphocytes for a relative assessment of the oxidative stress degree in cancer cells, were used. Results. An optimal final concentration of hydrogen peroxide - 0.5 mM - was established, causing an increase of active oxygen forms concentration in cells, comparable to levels in tumor cells, the formation of a maximum number of annexin positive cells and minimum number propidium-positive cells and the comparable ratio of the reduced and oxidized glutatione levels. Conclusion. The optimum concentration of hydrogen peroxide (0.5 mM) was selected for the oxidative stress formation in the peripheral blood lymphocytes to study the apoptosis dysregulation in oxidative stress in Jurkat line tumor cells (human T-lymphoblastic leukemia).