AIM:To study the effect of CD73 on breast cancer cell line MB-MDA-231 adhesion to extracellular matrix.METHODS: ① CD73 siRNA plasmid was constructed and transfected into MB-MDA-231 cells by lipofectamine 2000.② The transfection efficiency was analyzed by flow cytometry using eGFP as a marker gene.Stable transfected MB-MDA-231 cells were selected using G418.③ RT-PCR and Western blotting analysis of CD73 expression in MB-MDA-231 cells was performed.④ The effects of CD73 on MB-MDA-231 cells adhesion to extracellular matrix were assessed by cell adhesion assay.RESULTS: ① Transfection of MB-MDA-231 cells with CD73 siRNA plasmid significantly inhibited CD73 expression at mRNA level.The efficiency was up to 91%.② Transfection of MB-MDA-231 cells with CD73 siRNA plasmid significantly inhibited CD73 expression at a protein level.The efficiency was up to 79.3%.③ Treatment of MB-MDA-231 cells with CD73 siRNA resulted in diminished adhesion to extracellular matrix.CONCLUSION: This study demonstrates that CD73 siRNA effectively inhibits CD73 gene expression in MB-MDA-231 cells leading to adhesion to extracellular matrix suppression.

ObjectiveTo study the biological characterization and the genetic background of circulating CA16 strains in mainland of China for the purpose of CA16 vaccine development in the future.MethodsCA16 strains were isolated from throat swabs of patients with hand-foot-mouth disease and identified by neutralization assay and RT-PCR.The genotype of these isolates were determined by sequence alignment and phylogenetic analysis of VP1 gene.The proliferation dynamics and the plaque morphology were observed when propagated in Vero cells.The pathogenicity of these CA16 isolates was evaluated by challenging newborn mice.ResultsIn this study,six CA16 circulating isolates,BJ-1-6 were obtained.The RT-PCR products were 150 bp amplified with the general enterovirus primers and 210 bp with CA16 primers respectively,which cannot be amplified by EV71 primers.Additionally,these isolates were identified to display some obvious proliferation dynamics and plaque morphology when propagated for 96 h in Vero cells.The diameter of plaques were about 1.5 to 2 mm for BJ-1,BJ-2,BJ-4,BJ-6,4-5 mm for BJ-3 and 3 mm for BJ5,the plaques were regular except BJ-3.All the six isolates can be neutralized by the convalescent serum of patient infected with CA16.The virus titer of different isolates propagated for five passages in Vero cells was 7.0LgCCID50/ml.The sequence alignment of VP1 gene demonstrated that the genotypes of BJ-2,BJ-4,BJ5 were C1 and BJ-1,BJ-3,B J-6 were C,3 comparatively.The genetic distance of the VPI gene from theseisolates suggested that they were highly genetic identity with the homology of 90％ in nucleotide and 99％ in dedicated amino acid respectively.However,a distinctive difference in pathogenic ability in neonatal mice was found that the suckling mice challenged with BJ-3 & BJ-5 were paralyzed 4-5 d and dead 6-7d postchallenge,compared with the control group without any abnormality in the during of 14 d.ConclusionThe circulating CA16 isolates in China have different biological characteristics,different pathogenic ability and similar genetic backgrounds,which is helpful for the development of a CA16 vaccine in the future.