Protocol 
RAC Application 
Kenneth L. Brigham, M.D. 
monitoring during an intervention done with a fully awake, unmedicated animal. After the 
sheep recovered from surgery, a baseline response to endotoxin infusion (0.5|ig/kg) was 
measured. One week later, we infused intravenously 8mg DNA of the same pCMV^-PGH 
plasmid which we used in the rabbit studies described above and in the appended manuscript 
(PI 3). The plasmid was CO mplexed with 24 mg cationic liposome (L ipo fee tin ’^) prior to inf usion. 
Pulmonary and systemic hemodynamics and lung mechanics were monitored throughout and 
following the infusion. During plasmid/liposome administration there was no evidence of 
distress in the animal. The only physiologic effect observed during infusion of the 
plasmid/liposome complex was a slight increase in pulmonary artery pressure which was 
directly related to the infusion rate. Slowing the infusion rate eliminated this response. 
Twenty four hours following 
administration of the plasmid/liposome 
complex, endotoxin (0.5/ig/kg) infusion was 
repeated and the two responses compared. 
Figure 4 shows the alveolar to arterial 
oxygen gradient over the course of the 
endotoxin response in the two studies 
conducted in this animal (i.e. before and 
after PGG/H gene transfer). Following 
transfection, the hypoxemia consequent to 
endotoxin infusion was substantially 
decreased. Similar results were seen for 
comparisons of lung mechanics. As shown in 
Figures 5 and 6, lung tissue removed from 
this animal at the end of the endotoxin 
experiment generated increased amounts of 
PGE, and decreased amounts of 
thromboxane. These results are like those 
which we saw in the rabbit experiments. 
During the DNA/liposome administration, 
there was no evidence of any distress in 
the animal. 
Figure 4: A-a DOj Responses to Endotoxin in a Sheep before and 
after PGH Synthase Gene Transfection 
Organ Culture 
Recombinant DNA Research, Volume 19 
[181] 
