Background: Arsenic is an environmental pollutant particularly found in drinking water of developing countries in Asia and there has been broad concern that the exposure of arsenic elicits neurotoxicity in the area. As a putative mechanism underlying such neurotoxicity, we found previously that the exposure to arsenic induces neuronal cell death and suppresses neurite outgrowth. On the other hand, the effect of arsenic on astrocytes is not sufficiently clarified yet. In this study, we aimed to examine whether the exposure of sodium arsenite (NaAsO2) impairs the proliferation of primary astrocytes by using fluorescent ubiquitylation-based cell-cycle indicator (Fucci).
Method: We isolated and cultured astrocytes obtained from the cerebral cortex of the transgenic mice expressing monomeric Kusabira Orange 2 (mKO2) fused with the ubiquitylation domain of hCdt1, a maker for G1 phase, and monomeric Azami Green (mAG) fused with the ubiquitylation domain of hGem, a maker for G2, S and M phases. At 7 to 10 days in vitro, astrocytes were treated with serum-deprived DMEM for 72 h to synchronize the cell cycle. Subsequently, the cell cycle of astrocytes treated with NaAsO2 at the concentration of 0, 1, 2 or 4 μM was monitored by fluorescent time-lapse microscopy for 73 h in the DMEM containing 10 % FBS.
Results: We found that the length of the period from G1 to S phase was shortened by the exposure of 4 μM NaAsO2, although normal cell cycle was exhibited in astrocytes exposed to lower doses. We also found that a significant number of astrocytes treated with 4 μM NaAsO2 died after entry into S phase.
Discussion: These results suggest that the exposure of NaAsO2 at 4 μM could impair the cell cycle of astrocytes with accelerating entry into S phase. This adverse effect of NaAsO2 on cell cycle might be associated with the higher rate of cell death in astrocytes.