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Objective: To investigate the role of simultaneous blockade of CD40/CD40L and B7/CD28 pathways in the immune tolerance via co-expression of sCD40LIg and CTLA4Ig mediated by replication-defective adenovirus. Methods: Ad-sCD40LIg- IRES2-CTLA4Ig, replication-defective adenovirus co-expressing sCD40LIg and CTLA4Ig, was constructed and identified. The co-expression of sCD40LIg and CTLA4Ig was evaluated with confocal laser scanning microscope and Western blotting. Skin transplantations of C57BL/6 to BALB/c mice were performed. PBS, Ad-Shuttle-CMV and Ad-sCD40LIg-IRES2-CTLA4Ig were administered. Skin graft survival was monitored and the mRNA expression of both genes was evaluated in the skin allografts. Results: Ad-sCD40LIg-IRES2-CTLA4Ig was constructed successfully and identified. The co-expression of sCD40LIg and CTLA4Ig was identified with confocal laser scanning microscopy and Western blotting. Compared to the skin graft mean survival time (MST) of non-treated group ((5.75±0.71) d) or Ad-Shuttle-CMV-treated group ((5.50±0.53) d), the skin graft MST was dramatically prolonged in the Ad-sCD40LIg-IRES2-CTLA4Ig-treated group ((16.38±1.19) d, P<0.001). The mRNA expression of both genes was detected. Conclusion: Ad-sCD40LIg-IRES2-CTLA4Ig, a replication-defective adenovirus carrying genes encod-ing sCD40LIg and CTLA4Ig, was constructed. Simultaneous blockade of CD40/CD40L and B7/CD28 costimulatory pathway mediated by replication-defective adenovirus significantly prolonged skin allograft survival in mice.
Methods: Ad-sCD40LIg-IRES2-CTLA4Ig, replication-defective; Antibodies against CD40 / CD40L and B7 / CD28 pathways in the immune tolerance via co- expression of sCD40LIg and CTLA4Ig mediated by replication- defective adenovirus. Methods: The co-expression of sCD40LIg and CTLA4Ig was evaluated with confocal laser scanning microscopy and Western blotting. PBS, Ad-expression of sCD40LIg and CTLA4Ig were constructed and identified. Shuttle-CMV and Ad-sCD40LIg-IRES2-CTLA4Ig were administered. Skin graft survival was monitored and the mRNA expression of both genes were evaluated in the skin allografts. Results: Ad-sCD40LIg-IRES2- CTLA4Ig was constructed successfully and identified. The co -expression of sCD40LIg and CTLA4Ig was identified with confocal laser scanning microscopy and Western blotting. Compared to the skin graft mean survival time (MST) of non-treated group ((5.75 ± 0.71) d) or Ad The skin graft MST was significantly prolonged in the Ad-sCD40LIg-IRES2-CTLA4Ig-treated group ((16.38 ± 1.19) d, P <0.001) Expression of both genes were detected. Conclusion: Ad-sCD40LIg-IRES2-CTLA4Ig, a replication-defective adenovirus carrying genes encod-ing sCD40LIg and CTLA4Ig, was constructed. Simultaneous blockade of CD40 / CD40L and B7 / CD28 costimulatory pathway mediated by replication- defective adenovirus significantly prolonged skin allograft survival in mice.