Inhibition Of HMGB1 Gene Expression In Hela Cervical Cancer Cells By RNA Interference

Objective To investigate the effects of RNA interference-induced gene silence of HMGB1 on the proliferation of human cervical cancer Hela cells line.Method The recombinant Plasmid PGCsi 3.0-1,PGCsi 3.0-3 and PGCsi3.0-Neg were constructed and transfected into Hela cell by transient transfection.The mRNA of HMGB1 was determined by fluorescence quantitative real-time polymerase chain reation(FRQ-PCR) assay;The protein expression level of HMGB1 was determined by cell immunohistochemistry and western-blot.The proliferation of Hela cell was detected by MTT method and trypan blue coloring method;the cell cycles were analyzed by flow cytometry.Result PGCsi 3.0-3and PGCsi 3.0-1 vectors for HMGB1 gene were constructed successfully.,The transcription and expression of HMGB1 gene were inhibited effectively by the constructed vectors in the cervical cancer cell line Hela and the latter was better.The disturbing effect arrive at the highest after transfection 48h. Westen-blot showed that the protein inhibition ratio of Plasmids PGCsi3.0-1and PGCsi3.0-3 were 56.52%/49.9%and 54.7%/40.89%after transfection 48h and 72h. The results of FRQ-PCR indicated that PGCsi 3.0-1and PGCsi 3.0-3 vectors could significantly knock down the transcription of HMGB1 gene(71.02%/56.11%,respecttively). On the contrast,the control plasmid did exhibit no inhibitory effect on the protein expression and mRNA transcription of HMGB1 gene in Hela cell,The proliferation of Hela cell were detected by MTT method both decrease distinctively. The cell cycles showed RNAi could inhibit cell cyceles into G2 phase.The number of cells in G2 phase was increased after PGCsi3.0-1 and PGCsi3.0-3 transfected into Hela cell at 48,72 hours(p＜0.05),S phase was declined(p＜0.01),respectively.Conclusion HMGB1 gene can be significantly downregulated by introduction of siRNA in Hela cells to result in declined proliferation of Hela cells.This paves way for the subsequent studies.RNAi should be a new strategy in gene silence therapy targeting surviving.