GBSN - Microbiology (Unit 7) Microbiology in Everyday Life
Sigma Xi
1. I S A B E L D I P I R R O
T H E B R O N X H I G H S C H O O L O F S C I E N C E
Carbon monoxide protects lungs
from hyperoxia induced arrest of
alveolarization in neonatal mice
2. Introduction Methodology Results Discussion
BACKGROUND
Bronchopulmonary dysplasia: A developmental disorder from
lack of supplemental oxygen for at least 28 days after birth.
Social Impact and Development of Bronchopulmonary
Dysplasia (BPD): An arrest in alveolar development as a case of
neonatal chronic lung disease, BPD is the leading cause of
morbidity and mortality among survivors of severe prematurity.
The Frontier in BPD Development Mechanism: BPD is
linked to unrelated to ante- and post-natal conditions. This
presents difficulties in developing effective therapeutic strategy
for the prevention or the treatment of BPD.
3. Introduction Methodology Results Discussion
LITERATURE REVIEW
Risk Factors’ Mechanistic Effects on Mitochondrial Function: Pyridaben
renders mitochondrial function. The lung development of pyridaben-treated
mice demonstrated similar effects to hyperoxic lungs.
Direct Effect of Mitochondrial Function on Alveolar Development:
Dinitrophenol (DNP) was used to decrease mitochondrial function in mouse
lungs. These lungs exhibited decreased alveolar development, strengthening
the association between mitochondrial function and alveolar development.
Therapeutic Role of Carbon Monoxide on Oxidant Stress: As an effective
agent against oxidant stress through its anti-inflammatory behavior, CO
protects against pulmonary underdevelopment, an outcome of BPD. Studies
have yet to identify how CO plays a role in lung development through
inflammation resolution.
4. Introduction Methodology Results Discussion
RESEARCH PROBLEM
To investigate whether inhaled intermittent low doses of
carbon monoxide can protect neonatal mice against
hyperoxia-induced lung injury.
5. Introduction Methodology Results Discussion
SIGNIFICANCE
Prevention of BPD allows future generations to live
their lives independently and sufficiently.
Avoids extensive lifelong burden as BPD
negatively affects the individual’s quality of life,
relations and economic stability of themselves and
their families in all stages of their life.
6. Introduction Methodology Results Discussion
RESEARCH HYPOTHESIS
Inhaled CO can rescue pulmonary mitochondria from the O2
dysfunction and preserve alveolar growth and development
7. Introduction Methodology Results Discussion
PRIOR METHODOLOGY
To properly analyze lung development and function,
neonatal lungs were harvested for morphometric
analysis and mitochondrial respiration.
8. Introduction Methodology Results Discussion
EXPERIMENTAL MODEL
Alveolar developmental arrest was produced by prolonged (10 days) exposure
of 3 days old neonatal mice to a hyperoxic (70-75%O2) environment.
9. Introduction Methodology Results Discussion
EXPERIMENTAL METHODOLOGY
Methods of Lung Morphometry:
Cavalieri's principle utilization for digital lung volume
Image analysis used to determine the alveolar surface area,
parenchyma lung volume and septum thickness.
Mitochondrial Respiration and Analysis
Clark’s electrode used to measure oxygen concentration inside
the closed system of the chamber of mitochondria to quantify
respiratory rate.
Respiratory Control Ratio (RCR): The ratio between the
initiated states of state 3 (oxidative phosphorylation) and 4
(resting state) rates of respiration that reflects the efficiency of
mitochondrial respiration.
10. Introduction Methodology Results Discussion
ANALYSIS METHODOLOGY
To compare the results between the three study
groups I used one-way ANOVA with Fisher’s post
hoc analysis throughout the experiment. A
difference was identified as significant using a p-
value of less than 0.05.
11. Introduction Methodology Results Discussion
Hyperoxic mice
demonstrated
significantly (p = 0.02)
less rate weight gain
compared to untreated
naive mice. O2+CO mice
treated had the rate of
weight gain similar to
that of control mice and
significantly (p = 0.02)
faster than O2 mice.
12. Introduction Methodology Results Discussion
The representative images of 5μm lung section obtained at 2 weeks
of life (scale bar = 50μm). Hyperoxic lungs showed alveolar arrest
with (i) fewer and larger alveoli, (ii) shorter and less numerous
alveolar secondary buds and (iii) drastically thicker alveolar septum
compared to that in control mouse. The CO treated group was found
to have lung histology very similar to the control group.
13. Introduction Methodology Results Discussion
Morphometry data
(a) Total lung volume was significantly (p=0.01) decreased in O2 mice compared to the
control. O2+CO mice preserved lung volume compared to exposure to O2 mice.
(b) The alveolar surface area (p=0.007) decreased only in O2 mice.
(c) There was drastic (p > 0.02) increase of thickness of alveolar septum in both O2
groups compared to the control. Thickness of the alveolar septum was significantly
(p=0.008) smaller in O2+CO mice compared to that in O2 mice.
14. Introduction Methodology Results Discussion
(a) Representative mitochondrial
respiration showed a large disparity in
the mitochondrial function of O2 mice
compared to O2+CO and naïve mice.
(b) O2 exposure associated with a
significantly (p=0.03) decreased in the
rate of state 3. The rate of state 3 of CO
was (p=0.02) higher compared to that
of untreated O2 mice.
(c) State 4 of mitochondria respiration
was similar among all groups.
(d) Mitochondria of O2 mice exhibited a
significantly (p = 0.001) decreased
RCR, indicating that their
mitochondria were uncoupled and
therefore not functioning efficiently as
pulmonary mitochondria in naïve
mice.
15. Introduction Methodology Results Discussion
The mice within the O2+CO group produced increased levels of
Resolvin D1 compared to the naïve and O2 groups.
The O2 groups measured to have significantly larger measure of
macrophage-derived chemokine (MDC) (p=0.002).
Hyperoxic mice measured to have elevated levels keratinocyte-
derived chemokine (KC) from naïve mice (p=0.02).
16. Introduction Methodology Results Discussion
INTERPRETATION OF RESULTS
Weight gain preserved in O2+CO mice
Improved lung function and development of O2+CO
mice
O2 lungs produced high levels cytokines due to
prolonged stress
Elevated levels of resolvin in O2+CO mice indicate
increased reaction to stress
17. Introduction Methodology Results Discussion
MAIN DISCOVERY
The outcome of this study enables one to conclude
that inhaled carbon monoxide is in fact protective
against hyperoxia-induced (i) alveolar
developmental arrest and (ii) mitochondrial
dysfunction.
18. References
1. Zysman-Colman Z, Tremblay GM, Bandeali S, Landry JS. (2013) Bronchopulmonary
dysplasia - trends over three decades. Paediatr Child Health. 18(2):86-90.
2. Baraldi E, Carraro S, Filippone M. (2009) Bronchopulmonary dysplasia: definitions and
long-term respiratory outcome. Early Hum Dev. 85 (10 Suppl)
3. Jobe AH, Bancalari E. (2001) Bronchopulmonary dysplasia. Am J Respir Crit Care Med.
163(7):1723-9
4. Northway, W. H., Rosan, R. C., & Porter, D. Y. (1967). Pulmonary Disease Following
Respirator Therapy of Hyaline-Membrane Disease. New England Journal of Medicine,
276(7):357-368.
5. Ratner V, Slinko S, Utkina-Sosunova I, Starkov A, Polin RA, Ten VS. (2009) Hypoxic stress
exacerbates hyperoxia-induced lung injury in a neonatal mouse model of bronchopulmonary
dysplasia. Neonatology. 95(4):299-305.
6. Ratner V, Starkov A, Matsiukevich D, Polin RA, Ten VS. (2009) Mitochondrial dysfunction
contributes to alveolar developmental arrest in hyperoxia-exposed mice. Am J Respir Cell Mol
Biol., 40(5):511-8.
7. Mishra S, Fujita T, Lama V N, Nam D, Liao H, Okada M, Minamoto K, Yoshikawa Y,
Harada H & Pinsky D J. (2006). CO rescues ischemic lungs by interrupting MAPK-driven
expression of Egr-1 and its downstream target genes. Proc Natl. Acad Sci,103(13):5191-6
