Effects of Protein Tyrosine Kinase, Protein Tyrosine Phosphatase and Protein Kinase C on the Apoptotic Process of Leukemia Cells and Human Cortex Neurons Managed by Arsenic Trioxide.

Objective Study on the effects of protein tyrosine kinase, protein tyrosine phosphatase and protein kinase C (PKC)on apoptotic process, observe the changes of cytosolic calcium ([Ca²⁺]i) of human leukemia cells and cortex neurons after clinical relevant dosage of arsenic trioxide (As₂O₃) exposure in vitro.

Methods Marked [Ca²⁺]I of human leukemia cells and cortex neurons with Fluo-3/AM, assayed the changes of [Ca²⁺]I with laser confocal microscopy in real-time after managed with As₂O₃ in different concentrations respectively, detected the effects of protein tyrosine kinase, the protein tyrosine phosphatase and the activation of protein kinase C on these changes with confocal microscopy and Phosphorus radioisotope assay. Assayed the DNA ladders of leukemia cells and cortex neurons after exposed in As₂O₃.

Results As₂O₃ 1μmol/L can increase the [Ca²⁺]I of NB4 cells remarkably but show no effects on neurons. Vanadate, a kind of protein tyrosine phosphatase inhibitor, can dose-dependently promote the increasing of [Ca²⁺]I caused by 2, 5, 10μmol/L As₂O_3.The mean value of total increased rate in 280 seconds after exposed in As₂O₃ in different concentrations were: 6.5±2.3%, 21.7±2.1 %,49.2±2.5% in NB4 cells, and 6.7±2.1%, 19.4±2.5 %, 52.3±2.7% in cortex neurons respectively. Genistein, a kind of protein tyrosine kinase inhibitor, can dose-dependently decrease the raising of [Ca²⁺]I caused by 2, 5, 10μmol/L As₂O_3.The mean value of total inhibited rate in 280 seconds after As₂O₃ management in different concentrations were 6.7±2.9%, 25.6±2.5 %, 52.2±3.5% in NB4 cells, and 7.8±3.1%, 18.1±2.8%, 51.3±3.3% in cortex neurons respectively. The activation of PKC began when exposed in As₂O₃1μmol/L for 3 h, and keep raising continuously in NB4 cells., DNA ladders appeared after exposed in As₂O₃1μmol/L for 24h. However, no appearances above were found in human cortex neurons. In 2μmol/L As₂O₃ exposedness, The activation of PKC and DNA ladders appeared in neurons.

Conclusions The phosphorylation and dephosphorylation of protein tyrosine kinase and protein tyrosine phosphatase participated in nonspecific apoptosis signal transduction pathway of As₂O₃, and accompanied with PKC activation. The [Ca²⁺] elevation was closely related to PKC increasing activation. The tolerances between leukemia cell and cortex neuron to As₂O₃ were differ in dosage.