Construction of silencing glypican-3 gene plasmids and inhibition of human HepG2 cell proliferation

Objective: To investigate the silencing glypican-3(GPC-3) transcription on the effect of human HepG2 cell proliferation by constructing GPC-3 short hairpin RNA(shRNA).Methods: Few pairs of GPC-3 shRNA were designed,synthesized,and constructed according to GPC-3 sequence,inserted pGPU6/GFP/Neo vector,transformed into E.coli to screening and identify high efficiency plasmid.The effective sequence transfected into HepG2 cells for observing GPC-3 expression,respectively.Results: pGPU6/GFP/NeoGPC-3-shRNA1-4 plasmids were successfully constructed,and confirmed by enzymatic digestion with BamH Ⅰ or Pst Ⅰ.After HepG2 cells transfected with liposome mediated-shRNAs,the rate of silencing GPC-3 mRNA was up to 89.3 % by shRNA1.The proliferation rate of the transfected HepG2 cells was 71.1 % at 72 h in the GPC-3 shRNA1 group.The cell cycles after the cells transfected with shRNA were arrested in the G1 phase,and the apoptosis rate of HepG2 cells was increased to 65.6%.Conclusion: Specific GPC-3 shRNA might intervene effectively the gene transcription,inhibit HCC cell proliferation,and promote apoptosis.GPC-3 should be a potential target gene for HCC gene therapy.