cytokines (Boat et al . , 1989; McElvaney et al . , 1991; McElvaney et al . , 
1992; Suter et al . , 1988; Suter et al . , 1989a; Welsh and Fick, 1987). To 
insure that a replication deficient recombinant adenovirus can survive in 
this hostile environment, AdRSV.^gal, a recombinant vector containing the 
E. coli lacZ gene coding for ^-galactosidase (see Appendix 2 for details) 
was incubated at 37° in respiratory epithelial lining fluid from individu- 
als with CF for varying times and the fluid (with the vector) then placed 
on HeLa cells. Evaluation of the cells after 24 hr demonstrated the vector 
had infected the cells and the /?-galactosidase gene was expressed (Figure 
3.1.4-A). 
3.2 In Vivo Evidence 
The evidence that a recombinant vector containing the CFTR cDNA will trans- 
fer the cDNA to the airway epithelium in vivo comes from studies in cotton 
rats [a rodent with susceptibility to human adenovirus infection similar to 
that of humans (Ginsberg et al . , 1989)] and in non-human primates (rhesus). 
3.2.1 Expression In Vivo in Cotton Rats 
Studies with cotton rats have demonstrated that the intratracheal adminis- 
tration of Ad-CFTR results in expression of the human CFTR cDNA as evi- 
denced by: (a) PCR analysis showing human CFTR mRNA expression in the lung 
for at least 6 weeks; (b) in situ hybridization analysis demonstrating 
human CFTR mRNA transcripts in airway epithelial cells; (c) quantitative 
northern analysis showing that lung levels of human CFTR mRNA transcripts 
at 6 weeks are approximately 42% of that at 2 days after instillation; and 
(d) immunohistochemical evidence of human CFTR protein in airway epithelial 
cells. Details of these studies can be found in Appendix 1. 
In regards to the relative infectivity and resulting expression of the 
exogenous gene in various airway epithelial cells in vivo . studies were 
carried out in cotton rats receiving intratracheal AdRSV.^S gal, the recom- 
binant vector containing the /?-galactosidase gene. The results demonstrate 
that all major categories in the airway epithelium of large and small 
airways are equally infectable, including ciliated, basal, secretory and 
undifferentiated columnar cells (see Appendix 2 for details). 
3.2.2 Expression In Vivo in Non-Human Primates 
Intratracheal administration of the recombinant adenovirus vector contain- 
ing the lacZ gene to rhesus demonstrated expression of /3-galactosidase 
protein in airway epithelial cells at 3 days (Figure 3. 2. 2 -A) and 7 days 
(not shown) after administration. 
Recombinant DNA Research, Volume 16 
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