Omeprazole attenuated hyperoxic lung injury

1
Omeprazole induced CYP1A1 enzyme activity in Lung
and Liver
Omeprazole attenuated lung edema and hemorrhage

• Animals used
• C57BL/6J adult mice from Harlan.
• Treatment
• Omeprazole (100 mg/kg) or vehicle (Trioctanoin)
  intraperitoneally once daily for 4 days.
• Exposure
• Room air or hyperoxia (FiO2gt95) for 48 hours or
  72 hours.
• Animal groups
• Omeprazole hyperoxia
• Trioctanoin hyperoxia
• Omeprazole room air
• Trioctanoin room air
• Mice in each of the above groups were sacrificed
  at 48 hours and 72 hours of their assigned
  exposure.
• Estimation of lung injury
• Lung weight/Body weight ratio (LW/BW) and lung
  histopathology.
• Estimation of CYP1A1/1A2 induction
• Enzyme activity
• Ethoxyresorufin-O-deethylase (EROD) and
  methoxyresorufin-O-deethylase (MROD) assay for
  CYP1A1 and CYP1A2 activities respectively.
• Apoprotein levels Western Blotting.
• Statistical Analysis Data are expressed as mean
  SE.

Background Excessive supplemental oxygen
contributes to lung injury in experimental
animals and bronchopulmonary dysplasia (BPD) in
preterm infants. We showed earlier that inducers
of cytochrome P 450 1A (CYP1A) attenuate
hyperoxic lung injury in rodents. Omeprazole, a
proton pump inhibitor, induces CYP1A in human and
animal cells in vitro. However, whether
omeprazole induces CYP1A in vivo is
unknown. Hypothesis To test the hypothesis that
omeprazole pretreatment of wild type mice will
induce CYP1A expression and attenuate hyperoxic
lung injury. Materials/Methods Adult C57BL/6J
mice were treated i.p. with omeprazole (100
mg/kg) or vehicle (trioctanoin) once daily for 4
days and exposed to room air or hyperoxia (FiO2
gt0.95) for 48 hours or 72 hours. The lung
weight/body weight (LW/BW) ratio and lung
histopathology were used to determine the extent
of lung injury. Pulmonary and hepatic CYP1A1 and
CYP1A2 activities were determined by measuring
the activities of ethoxyresorufin O-deethylase
(EROD) and methoxyresorufin O-deethylase (MROD)
respectively. CYP1A1 and 1A2 apoprotein levels
were determined by Western blotting. Results In
mice exposed to hyperoxia, only eight of twenty
trioctanoin-treated mice survived at 72 hours of
oxygen exposure compared to all of
omeprazole-treated mice. The LW/BW ratio was 22
lower in the omeprazole-treated mice at 72 hours
of hyperoxia compared to those treated with
trioctanoin hyperoxia. In mice exposed to
hyperoxia, omeprazole significantly induced EROD
(CYP1A1) and MROD (CYP1A2) activities, compared
to trioctanoin controls. CYP1A1 and CYP1A2
apoprotein levels were also significantly greater
in omeprazolehyperoxia group compared to
trioctanoin hyperoxia group. Conclusions
Omeprazole attenuated hyperoxic lung injury in
mice. The mechanism of protection by omeprazole
may involve CYP1A1 and CYP1A2 enzyme induction
since activities/levels of these enzymes were
augmented by omeprazole. Thus, omeprazole may be
a potential candidate for clinical trials for the
prevention/treatment of BPD in premature infants.




?
?
Hyperoxia Trioctanion
Hyperoxia Omeprazole
Figure3. Gross pathology of the lungs. The lungs
(?) of mice treated with omeprazole and exposed
to hyperoxia were significantly less edematous
and hemorrhagic compared to the lungs of mice
treated with trioctanoin and exposed to hyperoxia.
.
Omeprazole attenuated hyperoxic lung injury
Figure 6. EROD assay reflecting CYP1A1 activity
in liver and lung. CYP1A activity in lung and
liver was significantly higher in omeprazole
treated mice compared to trioctanoin treated mice
at 48 hrs (lung p0.03, liver plt 0.0005)
and 72 hrs (lung and liver plt 0.0005) of
exposure to hyperoxia. Lung CYP1A1 activity was
also significantly higher in omeprazole treated
mice compared to trioctanoin treated mice in room
air at 48 hrs ( plt 0.0005) after completion of
treatment.
Omeprazole treated mice showed improved survival
Omeprazole induced CYP1A2 enzyme activity in
Liver
Figure 4. HE stain of lung sections. Omeprazole
treated mice showed significantly less
perivascular edema (?), peribronchiolar edema and
disruption (?) and alveolar infiltrates and
hemorrhages (?) compared to trioctanoin treated
mice at 48 and 72 hours of hyperoxia.The lung
injury significantly worsened at 72 hours of
hyperoxia in the trioctanoin treated mice.

• Bronchopulmonary Dysplasia (BPD) is the most
  common
• complication of preterm infants and is
  associated with significant
• long-term pulmonary and neurodevelopmental
  problems.
• Excessive supplemental oxygen contributes to
  lung injury in
• experimental animals and BPD in preterm
  infants.
• CYP1A inducers attenuate hyperoxic lung injury
  in rodents1.
• Omeprazole, a proton pump inhibitor, induces
  CYP1A in human
• and animal cells in vitro2.

Figure 1. Survival of mice in percentage at 72
hours of hyperoxia. All the omeprazole treated
mice exposed to 72 hrs hyperoxia survived,
whereas only 40 of trioctanoin treated mice
survived at a similar point of time.
Omeprazole treated mice had less lung edema
Omeprazole induced CYP1A apoprotein expression
CONCLUSIONS/SPECULATIONS

• Omeprazole attenuates hyperoxic lung injury in WT
  mice by inducing CYP1A.
• 2. The results suggest that omeprazole may act
  as an anti-oxidant.
• Omeprazole may be a candidate for clinical trials
  for the prevention/treatment of BPD in premature
  infants.

Omeprazole pretreatment of wild type mice will
induce CYP1A expression and attenuate hyperoxic
lung injury.
REFERENCES
1. A. Sinha. et al., Toxicol.Sci.,2005,87,204. 2
. D. Diaz. et al., Gastroenterology,1990,99,737
.
Figure 2. The mean lung weight (mg)/body weight
(gm) ratio. The mean LW/BW ratio reflecting the
extent of lung edema was considerably less in
omeprazole treated mice compared to trioctanoin
treated mice at 72 hours of hyperoxia (plt
0.0005).
This work was completed with grant support from
Ikaria Therapeutics.