title: [Effect of downregulation of Tiam1 by siRNA on esophageal squamous cell carcinoma EC9706 cells].
abstract: OBJECTIVE
  To explore the effect of downregulation of Tiam1 by siRNA on the esophageal squamous cell carcinoma (ESCC) EC9706 cells, and provide theoretical basis for gene therapy of ESCC using Tiam1 as a molecular target.
  
  
  METHODS
  Tiam1 siRNA was transfected into EC9706 cells, and expression changes of Tiam1 mRNA and protein after transfection were detected by quantitative real-time PCR and Western blotting. Cell proliferation was analyzed using CCK-8 kit. Cell cycle and apoptosis of the EC9706 cells were assessed by flow cytometry. Cell cycle-related proteins and cell apoptosis-associated proteins were analyzed by Western blotting.
  
  
  RESULTS
  Compared with the untreated group and control siRNA group, the relative expression levels of Tiam1 mRNA (1.00 and 0.11 ± 0.02) were not significantly different (P > 0.05). The relative expression levels of Tiam1 mRNA in the Tiam1 siRNA group at 24, 48 and 72 h after transfection were 0.30 ± 0.04, 0.09 ± 0.01 and 0.09 ± 0.006, respectively, significantly lower than that of the untreated group (P < 0.05 for all). The expression level of Tiam1 protein at 24 h after Tiam1 siRNA transfection in the EC9706 cells was 0.11 ± 0.02, significantly lower than that in the un-treated group (0.44 ± 0.05) and control siRNA group (0.44 ± 0.04, P < 0.05 for all). The percentages of G0/G1 cells in the Tiam1 siRNA group, untreated group and control siRNA group were (54.48 ± 2.14)%, (40.69 ± 1.85)% and (41.78 ± 1.31)%, respectively (P < 0.01). The percentages of S phase cells in the Tiam1 siRNA group, untreated group and control siRNA group were (27.18 ± 1.65)%, (32.32 ± 1.15)% and (30.35 ± 1.09)%, respectively (P < 0.01). The expression levels of cyclin D1 protein in the untreated group, control siRNA group and Tiam1 siRNA group were 0.43 ± 0.02, 0.41 ± 0.01 and 0.11 ± 0.02, respectively (P < 0.05). The expression levels of p27 protein in the untreated group, control siRNA group and Tiam1 siRNA group were 0.10 ± 0.01, 0.09 ± 0.02 and 0.20 ± 0.02, respectively (P < 0.05). The ratios of early apoptotic cells in the untreated group, control siRNA group and Tiam1 siRNA group were (10 ± 0.9)%, (10 ± 0.5)% and (27 ± 0.7)%, respectively (P < 0.01). The expression levels of Mcl-1 protein in EC9706 cells of untreated group, control siRNA group and Tiam1 siRNA group were 0.47 ± 0.12, 0.48 ± 0.13 and 0.16 ± 0.02, respectively (P < 0.05). The expression levels of Bcl-2 protein in EC9706 cells of the untreated group, control siRNA group and Tiam1 siRNA group were 0.49 ± 0.08, 0.50 ± 0.05 and 0.04 ± 0.03, respectively (P < 0.05). The caspase-3 activities in the untreated group, control siRNA group and Tiam1 siRNA group were 2.3 ± 0.09, 2.3 ± 0.10 and 16.0 ± 1.50, respectively; and that of caspase-9 were 2.3 ± 0.08, 2.3 ± 0.11 and 14.5 ± 0.9, respectively (P < 0.05 for all).
  
  
  CONCLUSIONS
  Tiam1 siRNA can significantly inhibit the proliferation of esophageal cancer EC9706 cells, induce cell cycle arrest and cell apoptosis. These effects are related to the regulation of the expressions of cell cycle-related genes (cyclin D1 and p27) and cell apoptosis-related genes (Mcl-1, Bcl-1, caspase-3 and caspase-9) by Tiam1 siRNA.
authors.affiliation: Department of Integrated Traditional and Western Medicine, the Affiliated Cancer Hospital of Zhengzhou University; Henan Cancer Hospital, Zhengzhou 450003, China.
authors.forename: Huaimin
authors.initials: H
authors.lastname: Liu
authors.affiliation: Department of Integrated Traditional and Western Medicine, the Affiliated Cancer Hospital of Zhengzhou University; Henan Cancer Hospital, Zhengzhou 450003, China. Email: jlf_2000@126.com.
authors.forename: Lifeng
authors.initials: L
authors.lastname: Jiang
authors.affiliation: Department of Integrated Traditional and Western Medicine, the Affiliated Cancer Hospital of Zhengzhou University; Henan Cancer Hospital, Zhengzhou 450003, China.
authors.forename: Xiaoli
authors.initials: X
authors.lastname: Liu
mesh_terms: D017209:Apoptosis; D002294:Carcinoma, Squamous Cell / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D053148:Caspase 3 / Q000378:metabolism; D053453:Caspase 9 / Q000378:metabolism; D059447:Cell Cycle Checkpoints; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D019938:Cyclin D1 / Q000378:metabolism; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000378:metabolism; D015536:Down-Regulation; D004938:Esophageal Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D000077277:Esophageal Squamous Cell Carcinoma; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D064549:Myeloid Cell Leukemia Sequence 1 Protein / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics*; D000075684:T-Lymphoma Invasion and Metastasis-inducing Protein 1; D014162:Transfection
chemical_list: D020662:Guanine Nucleotide Exchange Factors; C579095:MCL1 protein, human; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D000075684:T-Lymphoma Invasion and Metastasis-inducing Protein 1; C087334:TIAM1 protein, human; D019938:Cyclin D1; D050760:Cyclin-Dependent Kinase Inhibitor p27; C505407:CASP3 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053453:Caspase 9