Hypoxia Mimics The Cocl <sub> 2 </ Sub> Ros And The Regulation Of Calcium And Its Significance In Human Acute Myeloid Leukemia Cells Hl60,

Objective.To induce differentiation and apoptosis of acute myeloid leukemic cells in treatment of leukemia has been an effective method recent years. Inherent cellular ROS and Ca²⁺ level of leukemia cells play a very important role in the proliferation,differentiation and apoptosis.Our research focus on the regulation of ROS and Ca²⁺ under a hypoxia condition which may explain the mechanisms of leukemia from an aspect of microenvironment.Methods.MTT assay was used to evaluate the cytotoxicity of hypoxia-mimicking agents CoCl₂ on two human acute myeloid leukemic cell lines,one human acute lymphoblastic leukemia cell line and one human chronic myeloid leukemia;RT-PCR,Western blot were used to observe the expression of HIF-1α.The level of ROS,Ca²⁺ under a hypoxia condition were detected by FACS and Laser Scanning Confocal MicroscopeResults:(1)The cytotoxicity of hypoxia-mimicking agents CoCl₂ was obviously detect in both human acute myeloid leukemic cell lines and human acute lymphoblastic leukemia cell lines,and the IC₅₀ values range from 71.35 to 160.25μM;chronic myeloid leukemia cell K562 seems not quite sensitive to hypoxia;(2)The expression of HIF-1αwas remarkably increased after 2 hours treatment of CoCl₂ on both mRNA and protein levels;(3)CoCl₂ could induce ROS production in a time-dependent manner.The peak of ROS was observed 8 hours after treatment.After then,the ROS level decreased gradually and fall to the base line;(4)CoCl₂ could induced an increase of cytosolic free Ca²⁺ concentration,and this process was also a time-dependent manner.Cytosolic free Ca²⁺ concentration climbed a peak after 8 hours treatment,and almost returned to the basic line after 10 hours treatment,but it didn't change a lot under an IC₂₀ CoCl₂ treatment.Conclusion.Different sensitivities of different leukemia cell lines to hypoxia may be closely related with bone marrow microenvironments and various differentiation grades.The similar concentration change of ROS and Ca²⁺ under hypoxia also suggested that as the key signal transduction factors,the signal transduction nets of each HIF-1α,ROS and Ca²⁺ could promote each other and restrict each other,integrating the vitro signals,regulating cell's life.