【Abstract】Objective To construct a recombinant adenoviral vector carrying antisense matrix metalloproteinase2 (MMP2) for use in the gene therapy to inhibit the invasiveness and migratory capacity of hepatocellular carcinoma (HCC) cell line HepG2 in vitro and in vivo models.
Methods Total RNA was extracted from HCC, and then a 500 bp fragment at the 5′ end of human MMP2 cDNA was synthesized by polymerase chain reaction (PCR) and was reversely inserted into the multiclone site (MCS) of the shuttle plasmid pAdTrack-CMV,with the resultant plasmid and the backbone plasmid pAdEasy-1,the homologous recombination took place in the E.coli BJ5183 and the recombinant adenoviral plasmid carrying the antisense MMP2 gene was constructed and generated. The adenoviruses(Ad-MMP2AS) were packaged and amplified in the HEK 293 cells.Then the viral titer was checked by GFP.
Results The recombinant adenovirus vector carrying antisense MMP2 was constructed successfully, the b green fluorescence was observed in HEK 293 cells under a fluorescence microscopy. The viral titer was 1×108/ml.
Conclusion The recombinant adenovirus Ad-MMP2AS constructed by us could introduce the antisense MMP2 into HepG2 effectively,which would provide experimental basis for reversing the overexpression of MMP2 in HCC and for inhibiting the invasiveness and migratory capacity of HepG2 in vitro and in vivo models.
Citation：
ZHANG Mingman,LI Dehua,GOU Xinghua,YAN Lnan,HAN Lei,ZHAO Lanying,HU Haiyang.. Construction of Recombinant Adenovirus Vectors Carrying Antisense Matrix Metalloproteinase-2. CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY, 2004, 11(5): 428-432. doi:
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- 2. Niu Q, Tang Z, Ma Z, et al. Relationship between serum matrix metalloproteinase2 and metastasis and recurrence following radical hepatic resection in hepatocellular carcinoma [Ｊ]. Zhonghua Gan Zang Bing Za Zhi, 2001; 9（Suppl）∶58.
- 3. 王爱民，江龙安，梁保忠，等. 原发性肝癌组织中明胶酶A基因表达增强 [Ｊ]. 中华普通外科杂志， 2001； 15（9）∶598.
- 4. He TC, Zhou S, da Costa LT, et al. A simplified system for generating recombinant adenoviruses [Ｊ]. Proc Natl Acad Sci USA， 1998; 95(5)∶ 2509.
- 5. Curran S, Murray GI. Matrix metalloproteinases in tumour invasion and metastasis [Ｊ]. J Pathol， 1999; 189(3)∶ 300.
- 6. Liotta LA, Tryggvason K, Garbisa S, et al. Metastatic potential correlates with enzymatic degradation of basement membrane collagen [Ｊ]. Nature， 1980; 284(5751)∶ 67.
- 7. Deryugina EI, Luo GX, Reisfeld RA, et al. Tumor cell invasion through matrigel is regulated by activated matrix metalloproteinase2 [Ｊ]. Anticancer Res， 1997; 17(5A)∶ 3201.
- 8. Sang QX. Complex role of matrix metalloproteinases in angiogenesis [Ｊ]. Cell Res， 1998; 8(3)∶ 171.
- 9. McKenna GJ, Chen Y, Smith RM, et al. A role for matrix metalloproteinases and tumor host interaction in hepatocellular carcinomas [Ｊ]. Am J Surg， 2002; 183(5)∶ 588.
- 10. Carter G, Lemoine NR. Antisense technology for cancer therapy: does it make sense? [Ｊ]. Br J Cancer, 1993; 67(5)∶ 869.
- 11. Lakka SS, Rajan M, Gondi C, et al. Adenovirusmediated expression of antisense MMP9 in glioma cells inhibits tumor growth and invasion [Ｊ]. Oncogene, 2002; 21(52)∶ 8011.
