Abstract:

To observe the effect of herpes simplex virusthymidine kinase/cytosine deaminase fusion suicide gene on MG63 cells. MethodsThe target cells were infected by recombinant adenovirus expressed TK and/or CD and the efficiency of infection was measured with green fluorescent protein. MG63 cells were treated with different prodrug concentrations and multiple of infection(m.o.i). Cell survival rate and proliferation inhibiting rate were measured. The killing mechanism was also investigated with flow cytometry (FCM) and Hochest 33342 fluorescent staining. Results100% MG63 cells could be infected with  recombinant adenovirus vector either single or double genes as determined by GFP detection. Survival of MG63 cells in AdCDTK infected group was 31% when GCV was used and 44% to 5Fc. Combination with two prodrugs, cell survival was reduced to 90%, lower than the AdTK(79%), AdCD(63%) or AdTK+AdCD(66%) groups with the same prodrug treatmemt (P＜0.05). Concentration of prodrugs or m.o.i of adenovirus in the fusion gene group was lower than that of the single gene groups with respect to the same inhibition effect on MG63 cells. The infected MG63 cells with the double gene group had greater bystander effect than both  AdCD and AdTK groups. The mechanism of killing MG63 cells demonstrated by FCM and fluorescent staining was cytotoxic action and apoptosis. ConclusionThese results prove that the double gene expressed recombinant adenovirus could be used successfully to inhibit the proliferation of MG63 cells and has more powerful killing effect than the single gene group.

Key words: Osteosarcoma; Thymidine kinase gene; Cytosine deaminase gene; Fusion suicide gene