title: [Influence of recombinant lentiviral vector encoding miR-15a/16-1 in biological features of human nasopharyngeal carcinoma CNE-2Z cells].
abstract: OBJECTIVE
  To study the influence of recombinant lentiviral vector encoding miR-15a/16-1 on biological features of human nasopharyngeal carcinoma CNE-2Z cells and underlying mechanisms.
  
  
  METHODS
  GFP-positive CNE-2Z cells transfected with recombinant lentiviral vector were selected. The experiment was divided into control group, transfected group, radiotherapy group, transfected-radiotherapy group. Cell proliferation was analyzed by MTT. Apoptosis was detected by flow cytometry. Radiotherapy sensitivity of the cells in control group and transfected group was evaluated by colony forming experiment. The expressions of miR-15a, miR-16-1 and bcl-2 mRNA were detected by real-time quantitative polymerase chain reaction (RT-qPCR). The expression of bcl-2 protein was detected by Western blot. The activation of Caspase-2 and Caspase-3 was evaluated by spectrophotometry.
  
  
  RESULTS
  Relative expression quantities of miR-15a and miR-16-1 in infected group were 524.80 ± 40.79 (t = 494.611, P = 0.000) and 466.11 ± 40.96 (t = 386.8, P = 0.000), respectively. The proliferation of the cells in transfected-radiotherapy group was the most obvious, followed by the cells in radiotherapy group and transfected group (F = 424.3, P = 0.000). The apoptosis rates of control group, transfected group, radiotherapy group and transfected-radiotherapy group were (2.2 ± 1.4)%, (9.6 ± 0.8)%, (2.9 ± 1.1)%, and (18.6 ± 0.7)% respectively(F = 158.5, P = 0.000). Clonogenic assay showed that the values of SF2, Do (1.473) and Dq (1.581) in transfected group were lower than those in control group. The relative expression levels of bcl-2 mRNA in transfected group, radiotherapy group, and transfected-radiotherapy group had no significant difference (P > 0.05). Decrease in the expression of bcl-2 protein in transfected-radiotherapy group was most significantly, followed by that in transfected group. The percentages of activated Caspase-2 in control group, radiotherapy group, transfected group and transfected -radiotherapy group were 0.12 ± 0.01, 0.24 ± 0.04, 0.35 ± 0.02, and 0.44 ± 0.04, respectively (F = 115.500, P = 0.000). The percentages of activated Caspase-3 in the groups were 0.13 ± 0.01, 0.27 ± 0.01, 0.43 ± 0.02, and 0.83 ± 0.06, respectively (F = 439.921, P = 0.000).
  
  
  CONCLUSIONS
  Recombinant lentiviral vector LV-miR15a/16-1 could improve the expression of miR-15a and miR-16-1 in CNE-2Z cells, inhibit the proliferation of CNE-2Z cells, promote apoptosis and enhance the sensitivity of the cells to radiotherapy probably by inhibiting bcl-2 expression, activating Caspase-2 and Caspase-3.
authors.affiliation: Department of Otorhinolaryngology, First Affiliated Hospital of Wenzhou Medical College, Wenzhou, China.
authors.forename: Chun-hong
authors.initials: CH
authors.lastname: Zhang
authors.forename: Xiao-bi
authors.initials: XB
authors.lastname: Fang
authors.forename: Wen-feng
authors.initials: WF
authors.lastname: Li
authors.forename: Qing-yuan
authors.initials: QY
authors.lastname: Shi
authors.forename: Li-ping
authors.initials: LP
authors.lastname: Wu
authors.forename: Xiao-yun
authors.initials: XY
authors.lastname: Chen
authors.forename: Zhen-xiao
authors.initials: ZX
authors.lastname: Huang
authors.forename: Peng
authors.initials: P
authors.lastname: Wu
authors.forename: Zhen-zhen
authors.initials: ZZ
authors.lastname: Wang
authors.forename: Zhi-su
authors.initials: ZS
authors.lastname: Liao
mesh_terms: D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D002219:Carbamates; D002277:Carcinoma; D053143:Caspase 2 / Q000378:metabolism; D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D003546:Cysteine Endopeptidases / Q000378:metabolism; D005822:Genetic Vectors; D006801:Humans; D035683:MicroRNAs / Q000378:metabolism*; D000077274:Nasopharyngeal Carcinoma; D009303:Nasopharyngeal Neoplasms / Q000378:metabolism*; D011720:Pyrazoles; D012333:RNA, Messenger; D000073739:Strobilurins; D014162:Transfection
chemical_list: D051017:Apoptosis Regulatory Proteins; D002219:Carbamates; C505361:MIRN15 microRNA, human; C505362:MIRN16 microRNA, human; D035683:MicroRNAs; D011720:Pyrazoles; D012333:RNA, Messenger; D000073739:Strobilurins; C513428:pyrachlostrobin; C490360:CASP2 protein, human; D053143:Caspase 2; D053148:Caspase 3; D003546:Cysteine Endopeptidases