ABSTRACT
Adipose-derived stem cells (ADSCs) may be useful as an efficient vehicle in cell-based gene therapy of human diseases due to their ability to migrate to disease lesions. This study investigated the ability of ADSCharbored human tumor necrosis factorrelated apoptosisinducing ligand (TRAIL) cDNA to facilitate TRAIL expression and induce A375 melanoma cell apoptosis as observed using a Transwell coculture system. A cell migration assay was used to observe ADSC migration ability. In addition, TRAIL protein expression was successfully detected by western blot analysis in ADSCs after stable transfection of TRAIL cDNA. The Transwell coculture system data showed that TRAIL-ADSCs could induce A375 cell apoptosis in a dosedependent manner. At the gene level, the killing activity of TRAIL-ADSCs was associated with activation of caspase4 and caspase8. Collectively, the data from the current study provides preclinical support of ADSCfacilitated TRAIL expression in the treatment of melanoma. Further investigation is required to evaluate and confirm the in vivo ability of TRAIL-ADSCs in therapy of melanoma in animal models.
Subject(s)
Adipose Tissue/cytology , Gene Expression Regulation, Neoplastic , Melanoma/genetics , Melanoma/metabolism , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/metabolism , TNF-Related Apoptosis-Inducing Ligand/genetics , Animals , Apoptosis/genetics , Caspases/metabolism , Cell Differentiation , Cell Line, Tumor , Cell Movement/genetics , Cell Survival/genetics , Disease Models, Animal , Gene Expression , Humans , Melanoma/therapy , TNF-Related Apoptosis-Inducing Ligand/metabolism , TransfectionABSTRACT
OBJECTIVE: To purify and identify recombinant Varicella-Zoster Virus Glycoprotein E. METHODS: The recombinant plasmid pGEX-VZVgE was induced by isopropyl-beta-D-thiogalactoside (IPTG), the fusion protein was purified with affinity chromatography column; then the purified fusion protein was cleaved by thrombin, and the product's antigenicity was examined by Western blot. RESULTS: The product of pGEX-VZVgE induced by IPTG was separated from the mixture proteins by the affinity chromatography column, the expressed fusion protein's relative molecular mass was about 98 x 10(3). After cleavage, the obtained VZV Glycoprotein E's relative molecular mass was about 72 x 10(3); the purified fusion protein and VZV Glycoprotein E were single band by SDS-PAGE. The available antigenicity of Glycoprotein E was confirmed by Western blot. CONCLUSION: Purification of VZV Glycoprotein E with affinity chromatography is an effective method. It provides a foreground for studies on the application of VZV gE.
