used poxvirus Crizotinib for boosting and the soluble factor(s) secreted by MVA may not or less affect the expression of
poxvirus itself. Viral interference was discovered several decades ago. Co-infection of cells with two replication-competent viruses results in suppression of replication of both viruses. The Ad and MVA vectors used in this study were not capable of replicating in mice and human. Therefore, we infected A549 cells (a human epithelial cell line, which can be infected by both Ad and MVA vectors without viral replication) with a GFP-expressing MVA vector and an mCherry-expressing Ad vector. We found that most of the cells were infected with only an individual virus (Fig. 3d), indicating that interference caused by the co-administration of the Ad vector and MVA vector may be different from that caused by dual replication-competent find more viral infection. To explore transgene expression, we co-infected A549 cells with a SEAP-expressing Ad vector and a GFP-expressing MVA vector. As shown in Fig. 3a and b, the MVA vector down-regulated the transgene expression produced by the Ad vector. Furthermore,
similar results were observed when the Ad-SEAP-infected A549 cells were incubated with a supernatant of the MVA-GFP-infected cells (Fig. 3c). This indicated that MVA vector-infected A549 may secrete soluble factor(s) that would cause suppression of Ad vector transgene expression. Recent studies have shown that bacterial and viral infection in cells results in the secretion of type I IFN via toll-like receptor, dependant or independent of the innate immune pathway [31], [32] and [33]. To explore whether innate immunity is involved in viral interference, we infected the A549 cells with Ad or MVA and detected the mRNA of IFNα, IFNβ, and IFNγ at various time points between 0 and 96 h post infection (Fig. 4a). The mRNA of IFNα and IFNγ MycoClean Mycoplasma Removal Kit was not detected at any point of time; however, only a small amount of IFNβ mRNA was detected after 40 cycles of PCR, indicating that type I IFN may have not had much influence on our results. A further study confirmed our conclusion, since blocking of IFNβ in the supernatant of the MVA-infected cells did not bring about recovery of Ad transgene expression
(Fig. 4b). In summary, we co-administered Ad-HIV and MVA-HIV or their mock vectors to mice, and observed the suppression of HIV-specific effector T-cell responses and a part of memory T cell responses, compared to vaccination with either of the vaccines alone. An in vitro experiment indicated that viral interference may involve other soluble factor(s) besides type I IFN. Our study may help in designing a vaccination regimen and in investigating viral interference in the future. We thank NIH Tetramer Core Facility (Atlanta, GA) for tetramers. This work was partially supported by a Grant-in-Aid from the Ministry of Education, Science, Sports and Culture of Japan. “
“The past 5 years have been a period of extraordinary achievement in the rotavirus field.