JOURNAL ARTICLE
RESEARCH SUPPORT, NON-U.S. GOV'T
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Growth inhibitory effect and apoptosis induced by extracellular ATP and adenosine on human gastric carcinoma cells: involvement of intracellular uptake of adenosine.

AIM: To study the growth inhibitory and apoptotic effects of adenosine triphosphate (ATP) and adenosine (ADO) on human gastric carcinoma (HGC)-27 cells in vitro and the mechanisms related to the actions of ATP and ADO.

METHODS: MTT assay was used to determine the reduction of cell viability. The morphological changes of HGC-27 cells induced by ATP or ADO were observed under fluorescence light microscope by acridine orange/ethidium bromide double-stained cells. The internucleosomal fragmentation of genomic DNA was detected by agarose gel electrophoresis. The apoptotic rate and cell-cycle analysis after treatment with ATP or ADO was determined by flow cytometry.

RESULTS: ATP, ADO and the intermediate metabolites, ADP and AMP, and the agonist of purinergic receptors, reduced cell viability of HGC-27 cells at doses of 0.3 and 1.0 mmol/L. The distribution of cell cycle phase and proliferation index (PI) value of HGC-27 cells changed when exposed to ATP or ADO at the concentrations of 0.1, 0.3 and 1 mmol/L for 48 h. ATP and ADO both altered the distribution of cell cycle phase via G0/G1- phase arrest and significantly decreased PI value. Under light microscope, the tumor cells exposed to 0.3 mmol/L ATP or ADO displayed morphological changes of apoptosis; a ladder-like pattern of DNA fragmentation obtained from HGC-27 cells treated with 0.1-1 mmol/L ATP or ADO appeared in agarose gel electrophoresis; ATP and ADO induced the apoptosis of HGC-27 cells in a dose-dependent manner at concentrations between 0.03-1 mmol/L. The maximum apoptotic rate of HGC-27 cells exposed to ATP or ADO for 48 h was 13.53% or 15.9%, respectively. HGC-27 cell death induced by ATP or ADO was significantly inhibited by dipyridamole (10 mmol/L), an inhibitor of adenosine transporter, but was not affected by aminophylline, a broad inhibitor of P1 receptors and pyridoxal-phosphate-6-azophenyl-2, 4-disulphonic acid tetrasodium salt (30 micromol/L), a non-selective antagonist of P2 receptors.

CONCLUSION: Extracellular ATP and ADO reduced the cell viability, arrested cell cycle and induced apoptosis in HGC-27 cell line by intracellular uptake of ADO. One of the main routes of ATP-induced apoptosis in HGC-27 cells is through the breakdown to adenosine.

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