Lentiviral-mediated miRNA against osteopontin suppresses tumor growth and metastasis of human hepatocellular carcinoma.
In our previous study, osteopontin (OPN) was identified as one of the leading genes that promote the metastasis of hepatocellular carcinoma (HCC). However, the mechanism by which OPN promotes metastasis of HCC is not understood. In this study, RNA interference mediated by viral vectors-which could induce a long-lasting down-regulation in gene expression-was applied to analyze the role of OPN in metastasis of HCC. Three lentiviral vectors encoding microRNA against OPN, Lenti.OPNi-1, Lenti.OPNi-2, and Lenti.OPNi-3, were constructed and found to down-regulate the OPN level by 62%, 78%, and 95%, respectively, in HCCLM3 cells which had an overexpression of OPN and a higher metastatic potential. Consequently, both Lenti.OPNi-2 and Lenti.OPNi-3 induced a significant decrease in matrix metalloproteinase (MMP)-2 and urokinase plasminogen activator expression, and led to an obvious inhibition of both in vitro invasion and in vivo lung metastasis of HCCLM3 cells (P < 0.001). Moreover, Lenti.OPNi-3, rather than Lenti.OPNi-2, could also suppress in vitro proliferation and in vivo tumor growth of HCCLM3. Smaller detectable tumors were found in only 50% of mice after implantation of Lenti.OPNi-3-transfected HCCLM3 cells (341 +/- 502.6 mm(3) versus >3500 mm(3) in controls; P < 0.001). Lenti.OPNi-3, not Lenti.OPNi-2, significantly suppressed the MEK/ERK1/2 pathway in HCCLM3 cells. Recombinant OPN was found to induce translocation of p65 into the nucleus of HCC cells and activation of MMP-2 and MEK/ERK/1/2, which were suppressed by the nuclear factor kappaB (NF-kappaB) inhibitor pyrrolidine dithiocarbamate.


CONCLUSIONS
OPN plays an important role in metastasis as well as tumor growth of HCC, in which different minimum threshold levels of OPN are needed. These effects may occur through activation of the mitogen-activated protein kinase and NF-kappaB pathways, and MMP-2. OPN could be a hopeful target for the control of HCC.
Sun
Bing-Sheng
BS
Liver Cancer Institute and Zhongshan Hospital, Institutes of Biomedical Science, Fudan University, Shanghai, China.
Dong
Qiong-Zhu
QZ
Ye
Qing-Hai
QH
Sun
Hai-Jing
HJ
Jia
Hu-Liang
HL
Zhu
Xiao-Qun
XQ
Liu
Dao-Yong
DY
Chen
Jie
J
Xue
Qiong
Q
Zhou
Hai-Jun
HJ
Ren
Ning
N
Qin
Lun-Xiu
LX
D000818:Animals; D006528:Carcinoma, Hepatocellular / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D015536:Down-Regulation / Q000187:drug effects; D006023:Glycoproteins / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D008113:Liver Neoplasms / Q000378:metabolism* / Q000473:pathology; D020930:MAP Kinase Kinase Kinases / Q000378:metabolism; D008297:Male; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D035683:MicroRNAs / Q000235:genetics* / Q000494:pharmacology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D009362:Neoplasm Metastasis; D051996:Transcription Factor RelA / Q000378:metabolism; D014162:Transfection
D006023:Glycoproteins; D035683:MicroRNAs; D051996:Transcription Factor RelA; C024229:osteopoietin; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020930:MAP Kinase Kinase Kinases; D020778:Matrix Metalloproteinase 2