title: In vitro application of RNA interference to silence livin gene expression to induce apoptosis in leukemia cells.
abstract: OBJECTIVE
  To search for new targets and novel methods of anti-leukemia treatment and to discuss the mechanism of silencing the livin gene using small RNA interference technology to induce apoptosis in the K562 leukemia cell line.
  
  
  METHODS
  We designed and synthesized livin-specific small interference RNA (siRNA). Transfected K562 cells were cultured. Reverse-transcription polymer chain reaction (RT-PCR) was used to detect livin mRNA expression. Protein expression for livin was detected using Western blotting. A non-transfected group was used as a control. Meanwhile, vectors carrying enhanced green fluorescent protein (EGFP) were transfected as a positive control and flow cytometry was used to determine the transfection efficiency by detecting green fluorescence. The rate of apoptosis was determined using the annexin V and propidium iodide double-staining method. ELISA was used to determine the activity of Caspase-3.
  
  
  RESULTS
  The transfection efficiency of electroporation was as high as 50%. The siRNA sequences could knockdown livin gene expression at both the mRNA and protein levels. Apoptosis rate of the cells was 27.41 ± 2.30% 48 h after transfection with specific siRNA. This was significantly higher than that of the control group (9.63 ± 0.89%, p < 0.05). The 48-h apoptosis rate of the combined effect group VP-16 (5 µmol/L) and transfection rate was 45.1 ± 4.40%, which was significantly elevated (p < 0.05) compared with the groups treated with only VP-16 or by transfection.
  
  
  CONCLUSIONS
  Caspase-3 activity in cells transfected with siRNA was significantly elevated compared to the cells in the non-transfected groups (p < 0.05).
authors.affiliation: LinYi Tumor Hospital, Linyi, China. wangzhen515_14@163.com.
authors.forename: J
authors.initials: J
authors.lastname: Lv
authors.forename: Z P
authors.initials: ZP
authors.lastname: Qin
authors.forename: M F
authors.initials: MF
authors.lastname: Zheng
authors.forename: Z C
authors.initials: ZC
authors.lastname: Chen
authors.forename: Z
authors.initials: Z
authors.lastname: Wang
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000235:genetics; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D053148:Caspase 3 / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000235:genetics; D020014:K562 Cells; D007938:Leukemia / Q000235:genetics / Q000378:metabolism*; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000235:genetics; D034622:RNA Interference / Q000502:physiology; D034741:RNA, Small Interfering / Q000008:administration & dosage* / Q000235:genetics
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C419866:BIRC7 protein, human; D051034:Inhibitor of Apoptosis Proteins; D009363:Neoplasm Proteins; D034741:RNA, Small Interfering; D053148:Caspase 3