DEVELOPMENTAL SUSCEPTIBILITY TO HYPEROXIC LUNG INJURYSara Berkelhamer, MDNorthwestern UniversityLurie Children’s Hospital
Background: Oxidative Lung Injury
Reactive oxygen species (ROS) are generated with exposure to supraphysiologic oxygen
ROS cause damage to proteins, lipids and DNA Animal models demonstrate that high FiO2 causes
lung injury and arrested lung development Lung injury is greatest in the most premature infants Even brief exposure to 90% O2 during resuscitation
increased the risk of BPD in 24-28w infants (Vento, 2009)
Modified from Li, et al, Current Respiratory Medicine Reviews, 2005.
plasma membraneO2
mitochondria
O2- H2O2 OH-
death factors
O2-
H2O2
OH−
JNK
BAX
BAX
SOD1 catalase
SOD2 catalase
NOX
O2
O2
ETC
1. Hypothesis:
developmental differences exist in ROS generation and cellular response to supraphysiologic oxygen
Methods In vitro studies
Intact lung slices and isolated alveolar cells Live cell imaging using a redox sensor, RoGFP Whole lung isolation for protein and mRNA analysis
In vivo exposures to 75% O2 (Biospherix Chamber) Lung analysis
Protein and mRNA Morphometry
Heart analysis RVH by Fulton’s Index (RV/LV+S)
Immature Lung ModelsMurine Intact Lung Slices
Rat Primary AT2 Cells
RoGFP: Sensor of redox state
Reduced
Oxidized
ReducedOxidized
Hyperoxia Induces Exaggerated Mitochondrial Oxidative Stress in AEC of Immature Lung Slices
Free Radical Biology, in revision
RoGFP: Alveolar Cells
Hyperoxia Induces Exaggerated Mitochondrial Oxidative Stress Response in Immature AT2
Free Radical Biology, in revision
Hyperoxia Induces Comparable Oxidative StressResponse in Immature and Adult PASMC of Lung Slices
A
Free Radical Biology, in revision
RoGFP: PA Smooth Muscle Cells
Antioxidant Expression Matures Postnatally
Free Radical Biology, in revision
24h of Hyperoxia Induces Increased SOD Activity in Adult but Not Immature Lung
SOD MnSOD
Free Radical Biology, in revision
24h Hyperoxia Induces Increased NOX1 mRNA and Protein Expression in Immature but Not Adult Lung
Free Radical Biology, in revision
mRNA protein
EUK134 Attenuates Hyperoxia-Induced
Increase in NOX1 Expression in Immature Lung
Free Radical Biology, in revision
Hyperoxia Induces Increased Cell Death in Immature but not Adult Lung Slices
Free Radical Biology, in revision
Summary Mitochondrial ROS are generated within
minutes of exposure to supraphysiologic oxygen
Immature alveolar lung cells demonstrate exaggerated mitochondrial oxidative stress in hyperoxia
ROS generation may be further amplified in the immature lung via up-regulation of NOX1
Immature lung slices demonstrate increased susceptibility to hyperoxia-induced cell death
2. Hypothesis:
A critical developmental window may exist in which exposure to supraphysiologicoxygen results in compromised lungdevelopment
Model: In vivo exposure
0 1 2 3 4 5 6 7 14
= 21% O2
= 75% O2
Control
P0-3
P4-7
BPD
Day
72h of Hyperoxia at P0-3 but not P4-7 Induces Persistent RVH
*
Control
P0-3
P4-7
*
72h of Hyperoxia at P0-3 but not P4-7 Disrupts Alveolarization
* *
Model: Developmental Susceptibility
Hyperoxia
ROS ROS
NOX1 antioxidantenzymes
NOX1
lung injuryBPD
antioxidantenzymes
Clinical Implications Supraphysiologic oxygen should be used
with great caution in premature infants A critical window may exist during which
there is increased susceptibility to oxidative injury
Antioxidants targeted to the mitochondria or delivered directly to alveoli may represent novel therapeutic approaches
AcknowledgementsPaul Schumacker, PhD Molly BallAnkur DattaDanijela DokicGina KimPaul MungaiSimran SabharwalJacqueline SchreiwerGregory Waypa
Kathryn Farrow, MD, PhDLyubov CzechRobin Steinhorn, MDStephen Wedgwood
Neonatal Resuscitation Program Young Investigator’s Award, 2011