|MONTIEL, NESTOR - Oak Ridge Institute For Science And Education (ORISE)|
Submitted to: Veterinary Immunology and Immunopathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/19/2012
Publication Date: 11/14/2012
Citation: Montiel, N.A., Smoliga, G.R., Arzt, J. 2012. Time-dependent biodistribution and transgene expression of a recombinant human adenovirus serotype 5-luciferase vector as a surrogate agent for rAd5-FMDV vaccines in cattle. Veterinary Immunology and Immunopathology. 151(1-2):37-48. DOI: 110.1016/j.vetimm.2012.10.003.
Interpretive Summary: The purpose of this study was to characterize biodistribution and gene expression of a recombinant adenovirus vaccine vector in tissues of vaccinated cattle. In preliminary work, adenovirus infection and gene expression were characterized during time-course experiments in cell culture. This preliminary data was used to conduct subsequent experiments in which steers were injected with the same adenovirus construct and euthanized at various time-points for the purpose of tissue collection and analysis. Several novel techniques were used to analyze the harvested tissue samples including: real-time PCR, luminometry, and microscopy. The substantial findings were that 1) subsequent to inoculation with the recombinant vector, a peak of virus protein and gene expression occurred at 24 hours post injection (hpi) at the injection site, 2) cells containing adenovirus and/or transgenic proteins were mostly consistent with macrophages and dendritic cells, and 3) adenovirus DNA, RNA, and proteins were detected (by PCR and microscopy respectively) at local lymph nodes at 24hpi. This information is critical to determine the ideal route of inoculation for Ad5-based vaccines.
Technical Abstract: Replication-defective recombinant adenovirus 5 (rAd5) vectors carrying foot-and-mouth disease virus (FMDV) transgenes elicit a robust immune response to FMDV challenge in cattle; however vaccine function mechanisms are incompletely understood. Recent efforts addressing critical interactions of rAd5 vectors with components of the immune system have elucidated important aspects of induction of protective immunity against FMDV. In the current study, a surrogate rAd5-Luciferase (rAd5-Luc) vector was utilized for indirect assessment of rAd5-FMDV infection of cattle during the first 48 h post inoculation. To compare infection dynamics and time-dependent transgene expression, bovine cells were inoculated in vitro with rAd5-FMDV and rAd5-Luc vectors. Superior transgene expression was detected in cells infected with rAd5-Luc compared to rAd5-FMDV. However, both vectors behaved remarkably similar in demonstrating elevated mRNA transcription at 24 and 48 hpi and the occurrence of a peak of transgene expression at 48 hpi. Injection sites of cattle inoculated with rAd5-Luc contained mononuclear inflammatory infiltrates with hexon and transgene proteins associated with antigen-presenting cells. Luciferase activity, as well as microscopic detection of luciferase antigens, peaked at 24 hpi. Presence of mRNA transcripts also peaked at 24 hpi but unlike luciferase, remained strongly detected at 48 hpi. Luciferase antigens were detected at the cortical interfolicullar areas of local LN, indicating active trafficking of antigen-presenting cells to lymphoid tissues. This work provides mechanistic insights on rAd5-mediated immunity against FMDV infection and may contribute to ongoing efforts to enhance vaccine efficacy.