engleski

Nucleosides prevent 5-aminoimidazole-4-carboxamide ribonucleoside-mediated effects on cell cycle progression and differentiation

Introduction: Our previous studies demonstrated that 5- aminoimidazole-4-carboxamide ribonucleoside (AICAr) reduced proliferation, stimulated autophagy and increased the expression of differentiation markers in U937 leukemia cells. As a pharmacological modulator of AMP-activated kinase (AMPK), AICAr is known to have profound effects on cell metabolism. Aim: The aim of this study is to test for the possible role of major metabolic pathways in AICAr-mediated effects on cell cycle progression, autophagy and differentiation. Materials and methods: HL60 and U937 cells were incubated in the presence of AICAr, all-trans retinoic acid (ATRA) and metformin. The expression of differentiation markers CD11b and CD64 and DNA content for cell cycle analysis were determined by flow cytometry. Glucose, lactate and ammonia levels were measured using commercially available kits. Total cell lysates were analyzed for the level of LC3B and actin by Western blot. Results: The results show that AICAr has no significant effects on glucose consumption and lactate production, but significantly increases the production of ammonia and that increase depends on the presence of glutamine in medium. Although ammonia has been reported to induce autophagy flux in some cell types, AICAr-mediated increase in autophagy flux does not depend on glutamine. However, AICAR-mediated effects on the expression of differentiation markers are significantly inhibited by glutamine deprivation and treatment with a pharmacological inhibitor of glutaminase-1. Within the cell, glutamine has important roles as a carbon donor for intermediates of Krebs cycle and as a nitrogen donor for nucleotide synthesis. Cell cycle analysis of propidium iodide-stained U937 cells reveals that AICAr induces an arrest in S-phase of the cell cycle. Addition of nucleosides completely prevents AICAr-mediated S-phase arrest and significantly inhibits the expression of differentiation markers. Conclusion: The results of this study suggest that nucleosides and glutamine metabolism have important roles in AICAr-mediated effects on U937 cells.

nucelosides ; AICAr ; cell cycle ; differentiation

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