Effects of Nebulized Diethylenetetraamine-NONOate in a Mouse Model of Acute Pseudomonas aeruginosa Pneumonia: Effects Of DETA-NO and iNO
Effects Of DETA-NO and iNO on Endogenous NO Levels
In pilot studies, exposure to a single dose of 125 ^mol nebulized DETA-NO was associated with increased eNO and plasma NOx~ levels but no change in NOx~ levels in BAL fluid (Fig 4). eNO levels increased markedly within 30 min, which is consistent with intrapulmonary NO release, and then declined to baseline levels after > 24 h. Plasma levels of NOx~ also were markedly increased within 30 min of DETA-NO nebulization and then declined to baseline within 12 h (Fig 4B) other ventolin inhaler. Plasma NOx~ levels were significantly greater in mice with untreated pneumonia than in sham mice (Fig 5). Continuous exposure to iNO during the 24 h of the protocol increased plasma NOx~ levels in both sham and pneumonia mice but had a greater effect in mice with pneumonia. Moreover, doses of 10 and 40 ppm had similar effects in sham mice but plasma NOx~ levels were greater following administration of a dose of 40 ppm iNO than of 10 ppm iNO in mice with pneumonia.
When sham mice were twice exposed (at 4 and 12 h) to nebulized DETA-NO (both 12.5 and 125 ^mol doses) during the 24-h protocol, there was no effect on plasma NOx~ levels assessed at 24 h vs those in untreated sham mice (Fig 5). In mice with pneumonia that had been exposed to DETA-NO at 4 and 12 h, plasma NOx~ levels at 24 h were slightly reduced after exposure to 125 ^mol DETA-NO and were slightly higher after exposure to 12.5 ^mol DETA-NO compared to those in mice with untreated pneumonia.
Effects of DETA-NO Exposure on P aeruginosa Growth In Vitro
Exposure to DETA-NO at 5 and 50 mM (which are equivalent to nebulized doses of 12.5 and 125 ^mol, respectively) completely inhibited the growth of P aeruginosa, whereas exposure to DETA-NO at 0.5 mM had a lesser inhibitory effect (Table 2). Exposure to the DETA nucleophile alone (ie, exhausted DETA-NO) inhibited bacterial growth at all concentrations, which is similar to DETA-NO, NaNO2 had only a slight inhibitory effect on bacterial growth at a dose of 50 mM.
Figure 4. Effects of exposure of mice to a single dose of nebulized DETA-NO (125 |j,mol) on (top) eNO levels in parts per billion (ppb) and (bottom) plasma and BAL NOx~ levels (6 to 12 animals per group). * = p < 0.05 (vs respective preDETA-NO baseline); ** = p < 0.01 (vs respective pre-DETA-NO baseline).
Figure 5. Effect of nebulized DETA-NO or iNO on plasma NOx~ levels in sham mice and mice with P aeruginosa pneumonia. Untreated sham group, 19 mice; untreated pneumonia group, 28 mice; other treatment groups, 6 to 12 mice per group. * = p < 0.01 (pneumonia vs sham mice); # = p < 0.05 (DETA-NO or iNO-treated groups vs respective untreated control animals); ## = p < 0.01 (DETA-NO or iNO-treated groups vs respective untreated control animals).
Table 2—In vitro Growth of P aeruginosa During Exposure to DETA-NO, Exhausted DETA-NO, and NaNO2
|ExperimentalTreatment||ConcentrationmM||Initial Bacterial Concentration, cfu/mL|
|Control||100 ± 8||100 ± 2|
|DETA-NO||50||0 ± 0t||1 ± 1t|
|5||1 ± 0t||0 ± 0t|
|0.5||79 ± 4||79 ± 7|
|Exhausted DETA-NO||50||0±0||1 ± 1t|
|5||7 ± 2||4 ± 1t|
|0.5||54 ± 8||40 ± 8|
|NaNO2||50||68 ± 4||91 ± 2|
|5||97 ± 6||103 ± 5|
|0.5||72 ± 13||82 ± 17|