Abstract:

Objective To discuss the expression of circs_EFCAB2 in epileptic cell model in vitro，and to clarify its possible mechanism. Methods The magnesium-free epilepsy cell model （model group） was constructed in the human neuroblastoma LA-N-5 cells， and the cells untreated with magnesium-free extracellular fluid were regarded as control group. Real-time fluorescence quantitative PCR （RT-qPCR） method was used to detect the expression level of circ_EFCAB2 mRNA in the cells in two groups.The cells were didived into ribonuclease（RNase） R digestion group and RNase R undigestion group. RNA enzyme digestion experiment and RT-qPCR method were used to detect the expression levels of circ_ EFCAB2 and linear EFCAB2 mRNA in the cells in two groups； nuclear cytoplasmic separation experiment was used to detect the expression levels of circle_EFCAB2 mRNA in the epileptic cells； CCK-8 method was used to detect the proliferation activities of the cells in two groups； flow cytometry was used to detect the apoptotic rate and percentages of the cells at different cell cycles in two groups. Results The cells in model group showed spontaneous high-frequency epileptiform discharges， indicating the epilepsy cell model was successfully established. The RT-qPCR results showed that compared with control group， the expression level of circ_ EFCAB2 mRNA in the cells in model group was increased （P<0.05）. The RNA enzyme digestion experiment and RT-qPCR results showed that compared with RNase R undigestion group， the expression level of linear EFCAB2 mRNA in the cells in RNase R digestion group was decreased （P<0.01）. The nuclear cytoplasmic separation experiment results showed that there was no statistically significant difference in the expression level of circ_ EFCAB2 mRNA in the epileptic cells between the cytoplasm and nucleus of the epileptic cells（P>0.05）. The CCK-8 results showed that at 72 h after transfection， compared with control group， the proliferation activity of the cells in model group was significantly decreased （P<0.01）. The flow cytometry analysis results showed that compared with control group， the apoptotic rate of the cells in model group was significantly increased （P<0.01）；compared with control group， the percentage of the cells at S phase in model group was significantly decreased （P<0.01）， and the percentage of the cells at G₁+G₂ phase was significantly increased （P<0.01）. Conclusion The upregulated circ_ EFCAB2 may play a role in the pathogenesis of epilepsy by inhibiting the cell proliferation， promoting the apoptosis， and blocking the cell cycles.

Key words: Epilepsy, Circ_EFCAB2, Epilepsy cell model, Apoptosis, Cell proliferation