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《中国神经再生研究：英文版》
>Dose-dependent effects of lead on cell-cycle arrest, DNA damage, and cyclin D1 expression in primary cultured rat hippocampal neurons
Dose-dependent effects of lead on cell-cycle arrest, DNA damage, and cyclin D1 expression in primary cultured rat hippocampal neurons
BACKGROUND: Previous studies have suggested that the hippocampus is one of the neurotoxic target sites for lead. However, the molecular mechanisms of action, including the effect of lead on cell-cycle arrest, remain poorly understood. OBJECTIVE: To investigate the effects of different lead concentrations on cell-cycle arrest, DNA damage, and cyclin D1 expression in primary cultured rat hippocampal neurons. DESIGN, TIME AND SETTING: A randomized, controlled, in vitro experiment was performed at the China Medical University between July 2008 and May 2009. MATERIALS: Antibodies specific to cyclin D1 and actin were synthesized and purified by Santa Cruz Biotechnology, USA. FACStar flow cytometer was purchased from Becton Dickinson, San Jose, California, USA. METHODS: Wistar rat hippocampal neurons were primary cultured for 7 days. Neurons in the control group were treated with 0.01 mol/L phosphate buffered saline. Neurons in the 0.2, 1.0, and 10 μmol/L lead acetate groups were subjected to 0.2, 1.0, and 10 μmol/L lead acetate. Subsequently, hippocampal neurons in each group were cultured for 24 hours. MAIN OUTCOME MEASURES: The effects of lead on cell cycle were measured by flow cytometry, DNA damage was measured using the comet assay, and cyclin D1 expression was measured using Western blot analysis. RESULTS: Treatment of hippocampal neurons with 0.2 μmol/L lead acetate did not significantly alter cell cycle phase distribution, i.e., sub-G1, S, G0/G1, G2/M, whereas treatment with 1.0 and 10 μmol/L lead acetate significantly increased the percentage of S and sub-G1 phase cells (P 0.05). Following incubation with 0.2, 1.0, and 10 μmol/L lead acetate for 24 hours, cyclin D1 expression gradually decreased with exposure to increasing lead acetate concentrations (1.0-10 μmol/L). CONCLUSION: Lead exposure to primary cultured rat hippocampal neurons resulted in dose-dependently disturbed cellular homeostasis, including DNA damage, reduced cyclin D1 expression, and stagnation of cell-cycle progression.
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