| With the development of society and the progress of science, many diseases have been ableto cure. Malignant tumor has become the number one killer of human death, tumor treatment wasbecome one of the current challenges for medical and biology. The morbidity and mortality oftumor patient has increased year by year, tumor treated by traditional methods has been difficult.Looking for a kind of treatment concepts and methods which were safer and effective becomethe primary task of the medical and biological fields. The new concept of gene-virus therapy hasbringing cancer treatment a new prospect. The strategy of targeting gene-viro therapy wasadopted to construct recombinant adenovirus Ad-pShuttle-CMV-p14ARF (PCA) by homologousrecombination method.The lectins are un-enzymatic glycoproteins which can specifically agglutinin cells. Duo tothe binding function towards monosaccharides or polysaccharides, they are very important inimmunoregulation, plant defence, signaling pathways and inducing apoptosis. Themannose-binding lectin is the most commonly studied in the field of plant lectins. The snowdropbulb lectin, which named Galanthus nivalis agglutinin (GNA) is the frst monocotmannose-binding lectin isolated and characterized in details. Nowadays the monocotmannose-binding lectins have been generally classified as GNA (Galanthus nivalisagglutinin)-related lectin, which are distributed ubiquitously in plants and play a significant rolein recognition of the high mannose type glycans. Pinellia ternata is a traditional Chinesemedicinal plant belonging to Pinellia, Araceae species. Pinellia ternata agglutinin (PTA), astorage protein, accumulates in the tissues of P. ternate is a traditional herb with a more thantwo-thousand year history. PTA specifically recognizes and binds to mannan, but not to simplemannose, glucose and maltose. PTA agglutinins mice erythocyte, but not human’s. A number ofinvestigations have suggested that PTA has anti-tumor activity, anti-fugal activity, and anti-virusactivity. This research constructed a recombinant of adenovirus with p14ARF gene byhomologous recombination, named Ad-pShuttle-CMV-p14ARF (PCA). The aim of this study isto research the growth inhibiting effect to colon cancer cell line SW480and HT-29of PCA andPTA.Methods: Adenovirus Ad-pShuttle-CMV-p14ARF (PCA) was constructed following themanufacturer protocol of Adeasy system; PCR method for identification of positive recombinantvirus; Verified adenoviruses were amplified in293A cells and purified by CsCl density-gradientcentrifuge; We titered purified adenovirus according to TCID50protocol. PTA was purified by affinity chomatography and the concentration was determined subsequently. The proliferationeffect of the cell treated with PCA, PTA and their complex (PCA/PTA) was determined by MTTmethod; Crystal violet staining was carried out to analyse the killing effect of PCA, PTA andPCA/PTA on two colon cancer cells qualitatively. Hoechst33258staining assay was alsoperformed to observe the cell morphology changes.Results: MTT results showed that PCA, PTA and PCA/PTA exhibited dose-andtime-dependent effect on colon cancer cells SW480and HT-29, but PCA/PTA exhibitedapparently the most anti-proliferative activity towards the cells than another two. Theobservation of crystal violet staining disclosed dose-dependent killing effect of PCA, PTA andPCA/PTA on the cells. Typical apoptosis bodies were observed in the cells which were treatedwith PCA, PTA and PCA/PTA, but PCA/PTA showed more obvious apoptosis.Conclusion: Adenovirus and native Pinellia ternata agglutinin was firstly combined in thiswork to research whether they had a synergistic effect. The results revealed: both adenovirusPCA and PTA could inhibit the proliferation of colon cancer cell lines, and PCA combined PTA,however, exhibited the most anti-proliferative effect than PCA or PTA. |
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