title: [Effects of MIM-B gene on invasive and metastatic potentials of human hepatocellular carcinoma MHCC97H cells].
abstract: OBJECTIVE
  To investigate the effects of lentivirus mediated siRNA targeting human metastasis suppressor 1 (MTSS1, MIM-B gene) gene on the invasive and metastatic potentials of hepatocellular carcinoma (HCC) MHCC97H cells.
  
  
  METHODS
  The siRNA targeting MTSS1 was cloned into one lentivirus work vector. The work vector and three package plasmids were co-transfected into 293T cells with the help of lipefeetamine 2000. Lentivirus was collected in 72 hours and was added to the cultured MHCC97H cells. The total cell MIM-B mRNA and MIM-B protein were extracted and underwent real-time PCR and western-blot test respectively. Boden chamber assay was used to evaluate the invasive potential of MHCC97H cells. Gelatin zymography was used to detect matrix metalloproteinase-2 (MMP2) activity. Metastatic human HCC nude mice models were established by orthotopic implantation with a high metastatic potential human HCC cell line MHCC97H. Twenty-four nude mice bearing orthotopic xenografts were randomized into black control group, Lenti-GFP group and intervention group (Lenti-MTSS1 group) 14 days after orthotopic implantation (8 per group). The ultrasound-guided multi-point injection was performed on mice with borate buffered saline, Lenti-GFP and Lenti-MTSS1 respectively. Mice were sacrificed on day 35 for the examination of pulmonary metastasis. The SPSS 13.0 soft ware was applied to data analysis.
  
  
  RESULTS
  The small interfering RNA targeting MTSS1 was constructed successfully with a transfection efficiency of 97.0%, which produced a marked inhibition of invasive ability of MHCC97H cells through Matrigel, being 37.9+/-4.4, 37.4+/-5.3 and 26.6+/-4.6 in the black control group, Lenti-GFP group and Lenti-MTSS1 group (F = 26.695, P value is less than 0.01), respectively. MIM-B expression and MMP2 activity of intervention group were also significantly down-regulated as compared to the control group. The results of in vivo studies showed that the numbers of lung metastatic nodules were 6.5+/-2.6, 6.4+/-2.7 and 3.8+/-1.3 in the black control group, Lenti-GFP group and intervention group respectively with significant statistical difference (F = 3.637, P value is less than 0.05), accorded with tumor tissue MIM-B mRNA expression of 0.39+/-0.19, 0.38+/-0.10 and 0.16+/-0.11 respectively (F = 11.644, P value is less than 0.01) when comparison was made between control group and therapy group.
  
  
  CONCLUSIONS
  Small interfering RNA mediated by lentivirus inhibited MIM-B expression and resulted in inhibition of the invasive and metastatic potentials of MHCC97H cells, which may attributed, in part, the down regulation of MMP2 activity, and thus may provide a new molecular targeted therapy for HCC patients in the future.
authors.affiliation: Department of General Surgery, Shanghai Jiaotong University, Shanghai 200233, China.
authors.forename: Xiu-yan
authors.initials: XY
authors.lastname: Huang
authors.forename: Zi-li
authors.initials: ZL
authors.lastname: Huang
authors.forename: Yong-hua
authors.initials: YH
authors.lastname: Xu
authors.forename: Xin-yu
authors.initials: XY
authors.lastname: Huang
authors.forename: Jian
authors.initials: J
authors.lastname: Zhou
authors.forename: Sheng-long
authors.initials: SL
authors.lastname: Ye
authors.forename: Zhao-you
authors.initials: ZY
authors.lastname: Tang
authors.forename: Qi
authors.initials: Q
authors.lastname: Zheng
mesh_terms: D000818:Animals; D006528:Carcinoma, Hepatocellular / Q000235:genetics* / Q000473:pathology*; D045744:Cell Line, Tumor; D006801:Humans; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D008840:Microfilament Proteins / Q000235:genetics*; D009363:Neoplasm Proteins / Q000235:genetics*; D009368:Neoplasm Transplantation; D034741:RNA, Small Interfering / Q000235:genetics; D014162:Transfection
chemical_list: C486778:MTSS1 protein, human; D008840:Microfilament Proteins; D009363:Neoplasm Proteins; D034741:RNA, Small Interfering; D020778:Matrix Metalloproteinase 2