A Study On If Arsenic Trioxide Affects MYCN MRNA Expression And Cell Invasion In LA-N-5 Cells

Background:Neuroblastam is the most common extracranial malignant solid tumor in infants and children.The expression of MYCN oncogene is associated with advanced-stage disease,rapid tumor progression,resistance to treatment and poor outcome.Though with intensive chemotherapy,the 5 year survival is still lower,thus, investigation on new strategy to improve the prognosis of NB becomes necessary.Arsenic trioxide(As₂O₃) is successfully used in the therapy of acute promyelocytic leukemia(APL) and excellent result was reached,while the mechanisms of As₂O₃ against the tumor cells is obscure,might be associated with apoptosis and drug resistant.Objective:To see if As₂O₃ down-regulate MYCN mRNA expression and inhibit invasiveness of LA-N-5 cells.Methods:Two parts:1.MYCN mRNA in LA-N-5 cells pre and post Co-culture with As₂O₃ was detected with semi-quantitative RT-PCR;2.The invasiveness of LA-N-5 cells after treated with As₂O₃ was detected with the Transwell chamber method.24h,48h and 72hs after co-culture with different concentration of As₂O₃,RNA was isolated from LA-N-5 cells,MYCN and Glyceraldehyde phosphate dehydrogenase(GAPDH) mRNA expression was measured using semi-quantitative RT-PCR.The product was detected by electrophoresis on agarose gel and scanner.The level of MYCN mRNA was calculated based on MYCN OD value/GAPDH OD value,The capability of invasion of LA-N-5 cells was evaluated by the transwell chamber assay.Results:1.MYCN mRNA level in LA-N-5 cells was significantly inhibited in 48h after treated with different concentrations of As₂O₃ at 0.75μmol/L,1.5μmol/L and 3.0μmol/L respectively.The effection of 3.0μmol/L is the most obvious. MYCN OD value/GAPDH OD value was 0.89±0.03,0.98±0.05, 0.52±0.03 respectively,compared to control of 1.21±0.02,p value＜0.01 in all groups;2.In same density(3.0μmol/L),the inhibition effect was detected after 48h,and this effect can be kept.MYCN OD value/GAPDH OD value was 1.18±0.04,0.64±0.03,0.86±0.01, compare to control with 1.15±0.02,P value was 0.081,0.000,0.000 respectively;3.After treated with different concentrations of As₂O₃ for 24h,The invasion of LA-N-5 cells was decreased,the number of cells was 28.00±4.00,19.33±4.16,6.33±1.53 respectively, compare to control with 46.33±6.11,P value was 0.013,0.003,0.000.Conclusion:As₂O₃ is capable of down-regulating MYCN mRNA expression and suppress the invasion ability of LA-N-5 cells, with dose dependent manner.