Effect of Methotrexate on Cell Growth of Human Monocyte-like Cell Line U937 and Its Induction of Apoptosis

Ye Hui, Gu Longjun, Chen Hongxin, Chen Jing, Ye Yuchen, Xue Huiliang

Shanghai Children’s Medical Center, Shanghai Second Medical University,

Shanghai, China

 

Objective: To explore the novel approach of treatment in the patients with acute monocytic leukemia (AML-M5),

Methods: The U937 cell line, which has the monocytic characters, was used. Cell growth inhibition was measured by XTT assay after 24 and 48 hours in the continuous presence of various concentrations ranging from 1nmol/L to 100 mmol/L. To understand the mechanism of MTX cytotoxicity, a variety of assay, including trypan blue exclusion, flow cytometry, light microscopy (Wright’s staining) and DNA fragment electrophoresis, were performed.

Results: After 24 hours treatment, IC50 value for U937 cell was 0.04mmol/L. After 48 hours treatment, the IC50 was 0.037 mmol/L and IC90 was 0.39mmol/L. There were no significant apoptotic changes after shorter exposure of MTX (4 and 6 hours). After 8 hours at various concentrations of MTX treatment ranging from 5 nmol/L to 10 mmol/L, the percentage of the cells in the pre-G1 (apoptotic) was 3.2% at 0.1 mmol/L and it reached a peak of 18.2% at 5.0 mmol/L. The DNA synthesis in S-phase was inhibited from 41.2% (0.01 mmol/L) to 19.1% (10 mmol/L). DNA ladder band, a feature of apoptosis, was observed.

Conclusion: The arrest of cell growth and the apoptotic properties induced by MTX have lead to its evaluation as a potentially therapeutic agent in the treatment of AML-M5.

 
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