Effcts Of Inhibition Of Maspin By ShRNA On Gastric Cancer Cell SGC7901's Ability Of Invasion

Objective: To investigate the effects and molecular mechanism of maspin may be involved in the process of invasion ability in gastric cancer. Lay the academic basis for further implying RNA interference in maspin gene functional research.Methods: To design and synthesis targeting maspin shRNA , clone into plasmid pGenesil-1.1 eukaryotic expression vector .The recombinant expression plasmid pGenesil-1.1/maspin was identified by enzyme restriction and sequence analysis, the identified pGenesil-1.1/maspin plasmid was transfected into gastric cancer cell SGC7901 with LipofactamineTM2000. Fluorescence microscopy was used to observe the efficiency of transfection, the expression of maspin mRNA and protein were detected with reverse transcriptional polymerase chain reaction(RT-PCR) and Western blot assay, respectively, and evaluate the effect of RNA interference; changes in the expression of uPA,VEGF-C,MMP-7 mRNA and protein were detected with reverse transcriptional polymerase chain reaction(RT-PCR) and Western blot assay, respectively; Transwell chamber assay was performed to detect the alteration of SGC7901's ability of invasion.Results: Recombinant expression plasmid pGenesil-1.1/maspin was successfully constructed and it was stably transfected into SGC7901 cell lines, the efficiency of transfection was 85% in SGC7901 cells. In comparison with three control group, pre and post stable transfection the recombinant expression plasmid resulted in reduction of maspin mRNA and protein expressions by 75.8% and 62.7% respectively, and had significant difference (P<0.05). Compared with the group of transfected pGenesil-1.1, the level of mRNA and proteins of uPA ,VEGF-C increased significantly(P<0.05), the level of protein of MMP-7 also increased significantly(P<0.05), but there was no significant difference of MMP-7 mRNA level(P>0.05). Transwell chamber in vitro invasion experiment assay, group of transfected pGenesil-1.1/maspin than group of transfected pGenesil-1.1 significantly increased the number of cell invasion, the results are significant difference(P<0.05).Conclusion: The recombinant plasmid pGenesil-1.1/maspin could effectively inhibit the expression of maspin gene in gastric cancer cell lines SGC7901. The down-regulated expression of maspin gene result in enhancement of gastric cancer invasion ability, the molecular mechanism probably related to up-regulated the expression of uPA,VEGF-C and MMP-7.