Enhancing infection affinity of therapeutic recombinant viral vectors to targeted cells in regenerative nanomedicine

Abstract

Hyaline articular cartilage trauma and degeneration are major causes of suffering and pose great threat to the life quality of human beings worldwide. In cell-based therapy, TGF-β3 could be dosed to synovium-derived mesenchymal stem cells to induce differentiation into chondrocytes. However, type I collagen is intrinsically expressed and upregulated during monolayer culture by the introduction of TGF-β3, which would result in the formation of fibrous cartilage. Therefore, RNA interference strategy was adopted to suppress the expression of type I collagen by degrading its mRNA in a posttranscriptional pathway. In our study, we constructed a dual-functioning lentiviral vector that can express TGF-β3 and inhibit type I collagen expression. Based on this lentiviral vector, protocols for lentiviral titration, transduction and TGF-β3 quantification in the active form with modified ELISA would be specifically formulated and provided. Optimal medium would also be selected for the culture of transduced cells. Particularly, as the lentiviral nanoparticles are encapsulated in negative-charged envelope, there would be electrical repulsion between the lentiviral nanoparticles and the cells during viral transduction. Therefore, positive-charged polybrene has been added to mitigate the repulsion and facilitate lentiviral transduction. In our study, we tried several polybrene concentrations and determined the optimal one for lentiviral transduction.