Reversal of Methotrexate-Induced Folate Pool Depletion by Thymidine in a Human Leukemia Cell Line in Vitro

Abstract

In an In vitro study using a human monocytic leukemia cell line, U-937, the effects of interferon-γ (IFN-γ) in combination with the antifolate methotrexate and the role of thymidine introduced as a biochemical modulator were investigated. Methotrexate alone or in combination with INF-γ was found to enhance the induction of morphologic and functional monocytic differentiation in the U-937 cell line. Various cellular effects with the addition of thymidine to the medium with methotrexate and IFN-γ were studied. Enhanced inhibition of cell growth and perturbation of the cell cycle were noted when methotrexate and IFN-γ were used in combination, but not when methotrexate was used alone. The reduction of cellular folate by methotrexate was also enhanced in combination with IFN-γ. Cell cycle delay, resulting in cell growth inhibition of folate depletion, caused the induction of differentiation in U-937 cells, which was found to be greater with methotrexate + IFN-γ than with methotrexate alone. Cellular differentiation, as assessed by nitroblue tetrazolium reduction assay, indirect immunofluorescence and morphology, showed better effects towards the differentiation of U-937 cells when the agents were used in combination. However, addition of thymidine to the medium was found to cancel all the aforementioned effects. The addition of thymidine to the medium also caused reversal of the inhibitory effect of methotrexate and IFN-γ on cell growth and repletion of the endogenous folate level. Repletion of the folate level by exogenous thymidine is a new possibility for the role of the thymidine in cellular growth.