M.J. Welsh and A.E. Smith, RAC Application 
days. At the time of sacrifice of each animal, lung lavage and blood samples were taken for 
analysis. The lungs were fixed and processed for normal light-level histology. 
Blood and lavage fluid were evaluated for total leukocyte count and leukocyte differential. 
As an addition^ measure of the inflammatory process, lavage fluid was evaluated for total 
protein. Lung lavage fluid was also evaluated for the presence of infectious viral particles. 
Following embedding, sectioning and hematoxylin/eosin staining, lung sections were 
evaluated for signs of inflammation and airway epithelial damage. 
Results 
With the small sample size, the data from this preliminary study were not amenable to 
statistical analyses, however, some general trends could be ascertained. In the peripheral 
blood samples, total leukocyte counts showed no apparent dose- or time-dependent 
changes. In the blood leukocyte differential counts, there was a minor dose-related 
elevation in percentage of neutrophils at 6 hours, however, data from all other time points 
showed no elevation in neutrophil percentages. Taken together, these data suggest a 
minimal and transient systemic inflammatory response to viral administration. 
From the lung lavage, some elevation in total neutrophil and percentage neutrophil counts 
were observed at the first three time points (6 hr, 24 hr, 48 hr). By seven days, both total 
and percentage of neutrophil values had returned to the normal range. The trends in lung 
lavage protein levels were more difficult to assess due to inter-animal variability, however, 
no obvious dose- or time-dependent effects were readily apparent. From the lung 
histology, two clear observations were apparent. First, no damage to airway epithelium 
was observed at any time point or virus dose level. Second, a time- and dose-dependent 
mild inflammatory response was observed, being maximal in the 48 hr high virus dose 
animals. By seven days, the inflammatory response had completely resolved, such that the 
lungs from animals in all treatment groups were indistinguishable. 
In the virus titer assay of the lung lavage fluid, cytopathic effect (CPE) was observed in one 
of two control animals from the 6 hr and 24 hr time points, and in one of two high dose 
animals at all four time points. Culture supernatants from the initial CPE positive plates 
were reinoculated onto 293 cells, and examined by immunofluorescence for adenovirus 
structural proteins. DNA was extracted and characterized by restriction enzyme analysis. 
Only two of the CPE positive samples stained with adenovirus antibody (6 hr high dose, 48 
hrs high dose) and of these the 6 hr sample has been confirmed as Ad2/BGal, presumably 
residual imput virus. No sample has been confirmed to have wild-type virus. We assume 
that the majority of samples giving CPE, contained other viruses or agents, perhaps 
originating from the hamsters. 
Conclusion 
In summary, a mild, transient, inflammatory response appears to be associated with the 
intratracheal administration of the described doses of adenoviral vector in the Syrian 
hamster. Of note, the high dose hamsters received 100 times the number of viral PFUs as 
we propose to deliver to the highest dose human. 
b. Study Two. 
Procedures 
A single intratracheal Ad2/BGal-l administration hamster study is in progress. This study 
is designed to assess the possibility of productive infection of organs outside of the lung, 
the possibility of a long term pulmonary inflammatory response, and the antibody response 
of the animals to the adenoviral vector. In this study, the three treatment groups are: 
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Recombinant DNA Research, Volume 16 
