Journal of Pediatric Gastroenterology and Nutrition
43:336Y341 Ó September 2006 Lippincott Williams & Wilkins, Philadelphia
Pepsin, a Reliable Marker of Gastric Aspiration, Is
Frequently Detected in Tracheal Aspirates From
Premature Ventilated Neonates: Relationship With Feeding
and Methylxanthine Therapy
*Sabeena Farhath, †Zubair H. Aghai, †Tarek Nakhla, †Judy Saslow, *Zhaoping He,
*Sam Soundar and ‡Devendra I. Mehta
*Division of Gastroenterology and Nutrition, and Nemours Biomedical Research, Alfred I. duPont Hospital for Children,
Wilmington, DE; ÞDepartment of Pediatrics/Neonatology, Cooper Hospital-UMDNJ-Robert Wood Johnson Medical School,
Camden, NJ; and þDivision of Gastroenterology and Nutrition, Nemours Children’s Clinic, Orlando, FL
ABSTRACT
Objectives: To determine the frequency of pepsin detection in
tracheal aspirate (TA) samples of mechanically ventilated premature neonates and its association with feedings and methylxanthine therapy.
Patients and Methods: Serial TA samples (days 1, 3, 5, 7, 14,
21, 28 and 928 days) were collected from premature neonates
receiving ventilatory support. An enzymatic assay with a fluorescent substrate was used to detect pepsin. Pepsin was also
measured in 10 serum samples collected in conjunction with the
TA samples from 8 neonates.
Results: A total of 239 TA samples was collected from 45 premature neonates (mean birth weight, 762 T 166 g; mean gestational age, 25.5 T 1.5 wk). Pepsin was detectable in 222 of 239
TA samples (92.8%) and in none of the serum samples. Pepsin
was significantly lower on day 1 (mean, 170 T 216 ng/mL) when
compared with all other time points (P G 0.05). Mean concentration of pepsin was significantly lower when infants were
unfed (265 T 209 ng/mL) compared with levels during feeding
(390 T 260 ng/mL, P = 0.02). The mean level of pepsin was
significantly higher in infants during xanthine therapy (419 T
370 ng/mL) compared with no xanthine therapy (295 T 231 ng/mL,
P = 0.037).
Conclusion: Pepsin, a marker of gastric contents, was detected
in more than 92% of TA samples from premature infants on
mechanical ventilation. The level of pepsin was higher in fed
infants when compared with unfed infants. Xanthine therapy
was also associated with increased pepsin in TA samples. Chronic
aspiration of gastric contents may worsen lung disease in premature infants. JPGN 43:336Y341, 2006. Key Words: Gastric
aspirationVGastroesophageal refluxVMethylxanthineVPepsinV
Premature infants. Ó 2006 Lippincott Williams & Wilkins
INTRODUCTION
ventilated neonates in whom uncuffed ET tubes are used.
Gastroesophageal reflux may also precipitate esophagitis,
which lowers the esophageal sphincteric pressure and
further augments reflux, increasing the risk of aspiration
(13). However, literature to support the aspiration of gastric contents in premature neonates is sparse.
Low gastric pH along with pepsin and bile acid in
gastric contents can damage lung tissue (14,15). Aspiration due to GER in premature neonates on ventilatory
support can worsen lung disease and may contribute to
the development of bronchopulmonary dysplasia (BPD)
(16,17). The true prevalence of aspiration is difficult to
determine because of vague definitions, nonspecific signs
and symptoms, poor assessment methods and varying
levels of clinical recognition. There is no reliable method
to detect gastric contents in tracheal aspirate (TA)
samples in premature neonates. Measurement of pH in
TA samples is not a reliable marker of gastric contents
because gastric pH is greater than 4 for 90% of the time in
premature neonates (18,19). Hopper et al. (20) measured
Gastroesophageal reflux (GER) is very common in
premature neonates (1Y3). Multiple factors, including
immature tone of the lower esophageal sphincter (LES),
supine positioning, small stomach capacity, delayed
gastric emptying, decreased gastrointestinal motility and
the presence of a nasogastric tube, contribute to GER in
preterm infants (4Y7). In neonates, GER can be associated with apnea, bradycardia and aspiration of gastric
contents into the lungs (8Y10). Cuffed endotracheal (ET)
tubes in mechanically ventilated adults and children minimize the aspiration of gastric contents (11,12). However,
microaspiration may not be preventable in mechanically
Received March 23, 2006; accepted May 16, 2006.
Address correspondence and reprint requests to Sabeena Farhath,
MD, Division of Gastroenterology and Nutrition, Alfred I. duPont
Hospital for Children, PO Box 269, Wilmington, DE 19899 (e-mail:
[email protected]).
336
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PEPSIN DETECTION IN TA FROM VENTILATED NEONATES
TABLE 1. Demographic and clinical characteristics
(mean T SD) of study population (n = 45)
Birth weight (g)
Gestational age (wk)
Sex (M/F)
Race (black)
Prenatal steroids
Days on mechanical ventilation
Survived
762 T 166
25.5 T 1.5
27 (60%)
23 (51.1%)
40/45 (88.8%)
52 T 48
34 (75%)
lactose in TA samples as a marker of aspiration of
gastric contents. Lipid-laden macrophages in TA
samples are increased in premature neonates receiving
intravenous lipids (21). A specific marker of GERrelated aspiration should originate in the stomach, not
the lung. Detection of pepsin in TA samples is a new
reliable marker of gastric contents and microaspiration
and has been used as a marker of aspiration in adults
(22,23). Meert et al. (24) and Krishnan et al. (25) used a
similar technique to measure microaspiration in children.
However, to our knowledge, no study has been done to
detect pepsin in TA samples of premature neonates.
Methylxanthines (aminophylline and caffeine) are commonly used in premature neonates to prevent and treat
apnea of prematurity. Xanthine derivatives are known to
increase GER by reducing LES tone and increasing gastric acid secretion (1,4,26).
This study was performed to determine the frequency
of pepsin detection in TA samples from mechanically
ventilated premature neonates and its association with
feeding and xanthine therapy. We hypothesized that pepsin,
a marker of gastric contents, is detectable in TA samples
from premature ventilated neonates and that its level increases with increased feedings and with xanthine therapy.
337
Additional samples were obtained after 28 days in some infants
if ventilatory support continued. Tracheal aspirate samples were
obtained by instilling 0.5 mL of normal saline into the infant’s
endotracheal tube and suctioning the residue with a 5F suction
catheter after 2 or 3 ventilator breaths. The suction catheter was
passed to a standardized length of 0.5 to 1 cm beyond the tip of
the ET tube. This method of collection is used widely in
neonates to collect TA samples (27Y29), and it is well tolerated
by even the most critically ill neonates. The procedure was
repeated 3 times and replicates were pooled. The suction catheter was flushed with 0.5 mL of normal saline after each suctioning episode to collect the residual sample in the catheter.
The samples were immediately transported to the laboratory
on ice and processed within 30 minutes in the laboratory. The
samples were centrifuged at 4-C for 10 minutes at 300g. The
supernatant was collected, divided into aliquots and stored at
j70-C for future use.
Serum samples (n = 10) from 8 neonates were also collected
at the same time as the TA collection.
Pepsin Enzymatic Method
The enzymatic method was modified according to the assay
developed by Krishnan et al. (25). Porcine pepsin (Sigma-Aldrich,
St Louis, MO) standards (12.5Y400 ng/mL) were prepared in
0.1 mg/mL bovine serum albumin (BSA) with saline. Gastric
fluid from positive control patients was diluted in the same
BSA/saline solution, and the enzymatic reactions were carried
out in a 96-well microplate. Fifty microliters of standard or
sample was pipetted to microplate wells. Sample blanks were
prepared by incubating standard or sample on a 100-C dry
block for 5 minutes to inactivate the enzymatic activity. To each
well, 23 KL of 129 mM HCl was added to adjust the pH to 2.0
and left on ice for 15 minutes to inactivate lysosomal acid
hydrolase (cathepsin D) and to convert pepsinogen to active
pepsin (25). Next, 20 KL of 0.5% fluorescein isothiocyanate
PATIENTS AND METHODS
Study Population
The study was conducted in a 39-bed, level III neonatal
intensive care unit at Cooper University Hospital in Camden,
NJ, between March 2003 and October 2004. The Institutional
Review Committee approved the study, and parents signed a
written informed consent. Infants born before 32 weeks_ gestation and requiring mechanical ventilatory support were eligible
for participation. The decision to treat each infant with xanthine
derivatives was made by the attending neonatologist. Aminophylline was given as a loading dose of 8 mg/kg, and the maintenance dose was adjusted to keep the serum level between 7
and 15 Kg/mL. The initial loading dose of caffeine citrate was
20 mg/kg, and the maintenance dose was adjusted to keep the
serum level between 5 and 25 Kg/mL. Relevant clinical data
including the infant’s demographics, clinical parameters and
feedings were collected from the patient’s chart.
Sample Collections
Tracheal aspirate samples were collected on days 1, 3, 5, 7,
14, 21 and 28 while the infant was mechanically ventilated.
FIG. 1. The levels of pepsin over different time points (days 1, 3,
5, 7, 14, 21, 28 and 928). The level of pepsin was significantly
lower on day 1 when compared with all other time points (days 3,
5, 7, 14, 21, 28 and 928, all P G 0.05). Box plot: the boundary of
the box indicates the 25th and 75th percentiles, the line within the
box marks the median value and the error bars indicate the 5th
and 95th percentiles.
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338
FARHATH ET AL.
of pepsin was also measured in 10 serum samples from 8
premature neonates; samples were collected at the same
time as the TA. Pepsin was undetectable in all serum
samples and detectable in all TA samples.
Levels of Pepsin Over Time
FIG. 2. The levels of pepsin in preterm infant with gestational age
23 to 25 weeks (GA 23Y25, n = 23) and 26 to 31 weeks (GA
26Y31, n = 22). The level of pepsin was significantly higher on
days 5 and 7 in premature neonates GA 23Y25.
casein (Sigma-Aldrich) was added to each well and incubated
for 3 hours at 37-C. The plates were transferred back to the ice
tray, and 90 KL of 1.2 mg/mL BSA and 30 KL of 20%
trichloroacetic acid were added to each well for trichloroacetic
acid precipitation. The plates were centrifuged for 90 minutes at
3500 rpm and 8-C in a Sorvall centrifuge. Thirty-eight microliters of the supernatant was transferred to a new, clean, flatbottomed microplate, and 212 KL of 500 mmol/L Tris was added
to each well. The plate was read in a spectrofluorometer at
excitation (485 nm) and emission (530 nm), and the net fluorescent intensity was subtracted from the blanks. The final
pepsin concentration of the sample was determined based on the
net fluorescent intensity of the known concentrations of the
standards. The subjective pepsin level (defining positive from
negative) of an aspirate was set at the lower limit (12.5 ng/mL)
of the sensitivity of the assay.
The levels of pepsin were compared across different
time points (days 1, 3, 5, 7, 14, 21, 28 and 928). The
numbers of TA samples collected on days 1, 3, 5, 7, 14,
21, 28 and 928 were 30, 30, 34, 35, 28, 25, 23 and 34,
respectively. The concentration of pepsin in TA samples
was significantly lower on day 1 (mean T SD, 170 T
216 ng/mL) when compared with all other time points
(days 3, 5, 7, 14, 21, 28 and 928, all P G 0.05). There was
no significant difference in pepsin level among other
time points (Fig. 1).
Pepsin and Gestational Age
The levels of pepsin were compared in preterm infant
with gestational age 23 to 25 weeks (GA 23Y25, n = 23)
and 26 to 31 weeks (GA 26Y31, n = 22). The level of
pepsin was significantly higher on days 5 and 7 in
premature neonates GA 23Y25 compared with GA
26Y31 (Fig. 2).
Feeding and Pepsin Level
Thirty-four infants received enteral feeding during the
study period. The data were available on 30 infants to
compare the pepsin levels before and after feeding. If
more than 1 sample was collected from an infant before
or during feeding, the mean level of pepsin was used
for statistical analysis. The mean concentration of
pepsin was significantly lower when infants were unfed
Statistical Analysis
Statistics were performed using Sigma Stat 3.1 for Windows
statistical package (Systat Software, Inc, Point Richmond, CA).
A comparison of pepsin levels over time was done by KruskalWallis 1-way ANOVA. The comparisons between groups
(before and after feeding, xanthine therapy) were performed
using the paired t test and Wilcoxon signed rank test. The difference was considered significant for P G 0.05.
RESULTS
A total of 239 TA samples were collected from 45
premature neonates (mean birth weight, 762 T 166 g;
gestational age, 25.5 T 1.5 wk). Clinical characteristics of
the study population are summarized in Table 1.
Pepsin was detectable in 222 of 239 (92.8%) TA
samples. The median pepsin level of all aspirates was
283.21 ng/mL (range, 0Y2441 ng/mL). Eight of 17 negative samples were from day 1. After day 1, 200 of 209
(95.7%) TA samples were positive for pepsin. The level
FIG. 3. The levels of pepsin when infants were unfed (nil per os)
and during the feed. Median concentration of pepsin was significantly lower when infants were unfed when compared with
during feeding.
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PEPSIN DETECTION IN TA FROM VENTILATED NEONATES
FIG. 4. The level of pepsin and treatment with xanthine derivatives. Treatment with xanthine increased pepsin level.
(265 T 209 ng/mL) when compared with levels during
feeding (390 T 260 ng/mL, P = 0.02) (Fig. 3).
Xanthine and Pepsin
Thirty-six infants received xanthine derivatives during
the study period. The data were available on 23 infants to
compare the pepsin levels before and during the xanthine
therapy. The mean level of pepsin was used for statistical
analysis if more than 1 sample was collected from an
infant before or during the xanthine therapy. The mean
level of pepsin was significantly higher in infants during
the xanthine therapy (419 T 370 ng/mL) when compared
with no xanthine therapy (295 T 231 ng/mL, P = 0.037).
Because the level of pepsin was significantly lower on
day 1 in our cohort of population, we again analyzed the
data after excluding the samples from day 1. The data
from all 23 infants were available for this analysis.
Again, the pepsin level was significantly higher during
the xanthine therapy (no xanthine, 341 T 232 ng/mL;
xanthine, 523 T 370 ng/mL; P = 0.036) (Fig. 4).
DISCUSSION
Premature infants are predisposed to GER and are at
increased risk for pulmonary aspiration. Several factors
that lead to aspiration of gastric contents in premature
neonates are intubations and mechanical ventilation, use
of sedation, immature swallowing mechanism, reduced
muscle tone and suppression of reflexes that protect the
airways (4Y7). In infants, physiological functions seem
to be impaired at different swallowing stages, increasing
the predisposition to aspiration. At the same time, the
control over the systems that protect against aspiration
is lost due to a direct action of the endotracheal tube.
In the past, many tests have been done to look at
aspiration of gastric contents such as dye studies, lipidladen macrophages and glucose and lactose assays. In
clinical practice, technetium scintigraphy is used to
339
detect pulmonary aspiration. However, this study is performed during and after a single feed for 2 to 3 hours and,
therefore, has limited ability to detect the aspiration of
gastric contents (30). Hopper et al. (20) used the detection
of lactose in TAs to diagnose GER aspiration in neonates.
Conclusions from these studies usually are assumptions,
which can give false-negative results. Recent studies have
shown that modified pepsin assay has more sensitivity and
specificity in detecting aspiration (23Y25). An advantage
of the tracheal pepsin test over other methods for
assessing aspiration is that tracheal secretion samples
can be collected easily at the bedside as a noninvasive
procedure that does not interfere with the patient care.
In the current study, detection of pepsin was investigated as a marker for gastric contents in TA samples from
premature ventilated infants. The results demonstrated that
pepsin is detectable in 992% of all TA samples and in
995% of samples collected after day 1, suggesting significant aspiration of gastric contents in premature infants.
In 30 acutely ill, tube-fed, mechanically ventilated adults,
pepsin was detectable in only 14 of 136 TA samples (31).
Meert et al. (24) reported positive pepsin in only 9 of 100
TA samples from 37 children. They also found that children with pepsin-positive TA are more likely to have clinical evidence of GER (24). Krishnan et al. (25) detected
pepsin in 31 of 37 children with history of GER and respiratory symptoms and none in 26 children without history
of GER or respiratory symptoms. Recently, using intraluminal impedance technique, Lopez et al. (32) reported
that 979% of acid and nonacid refluxes reach the
proximal esophagus in premature neonates. Valat et al.
(33) measured radioactivity in ET after injecting solution
of 99mTc-sulfur colloid in the gastric tube in neonates.
Forty-two percent of neonates had positive radioactivity
in the tracheal tube, suggesting aspiration of gastric contents. Our data indicate that aspiration of gastric contents
into the lung is a widespread phenomenon in ventilated
premature infants. Apart from factors discussed earlier,
uncuffed ETs used in ventilated premature infants may
also have a major contribution to the high-pepsin positive
rate. Previous studies support the role of cuffed ET tubes
in the prevention of aspiration (11,12). Small amounts of
pepsinogen are detectable in human blood (34,35). To
exclude a hematogenous source of pepsinogen in TA
samples, pepsin was also measured in serum samples at
the same time of TA collection. Pepsin was undetectable
in all the serum samples.
The presence of pepsin in 995% of samples after day 1
suggests significant aspiration of gastric contents in premature ventilated infants. Aspiration of gastric contents
may play a major role in worsening of lung disease in
premature neonates. Twenty percent to 40% of ventilated
premature neonates develop BPD (36). Multiple factors
including high oxygen, ventilator-induced lung injury,
symptomatic patent ductus arteriosus, various lytic proteinases and nutritional deficiencies are implicated in the
etiopathogenesis of BPD, but the exact etiology is still
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340
FARHATH ET AL.
unknown (36). More, recently, a new chronic lung disease
called Bnew BPD^ is emerging in premature infants, not
related to oxygen therapy and mechanical ventilation (37).
The new BPD develops in premature neonates with no or
minimal initial lung disease and presents with gradual
worsening of respiratory failure. We speculate that chronic
aspiration of gastric contents may be contributing to the
worsening of lung disease in premature infants. The role
of chronic aspiration due to GER warrants more studies in
the development of this new BPD.
Our study shows that pepsin levels in TA samples were
low on day 1, suggesting minimal pulmonary aspiration
of gastric contents. In animal models, after a single episode of aspiration of gastric juice, pepsin was detectable
in all samples at 2 hours, at 4 hours, and in 90% of
samples at 6 hours, suggesting that pepsin can be detectable up to 6 hours after aspiration (38). Low levels of
pepsin on day 1 in our population are more likely related to decreased pepsin secretion immediately after
birth, no feeding and, possibly, lack of sufficient time
for aspiration of gastric contents. However, samples from
day 3 onward showed consistent presence of pepsin in TA,
suggesting ongoing microaspiration of gastric contents.
The level of pepsin was consistently higher after 3 days
and did not significantly change over time.
Our data indicate that the level of pepsin was significantly higher in TA sample in infants when they were fed
when compared with when they were not receiving feeds.
Feeding not only activates pepsin secretion but also increases reflux, leading to increased aspiration of gastric
contents into the lungs. Increased volume of feeds may
increase GER and aspiration. Additional studies are needed
to see the effect of various volumes of feeds on the aspiration of gastric contents in premature infants.
Xanthine derivatives are commonly used in premature
infants to prevent apnea of prematurity. Our data suggest
that the aspiration of gastric contents is increased in premature infants receiving xanthine derivatives. Xanthines
are known to increase GER by reducing LES pressure
(39,40). We were unable to compare the difference in
aspiration with caffeine and aminophylline, as all but 1
infant was treated with aminophylline.
We recognize some important limitations of this study.
Pepsin was measured in TA samples and was potentially
diluted. It is controversial whether TA samples should be
corrected for dilution in neonates (41). We followed the
recommendation of the European Respiratory Task Force
on bronchoalveolar lavage in children and did not correct
our result for dilution (41).
Enzymatic assays in our study measured both pepsin
and pepsinogen. Pepsin is an active form and contributes
to lung injury, whereas pepsinogen is an inactive form.
Secretion of pepsin is inconsistent and low in infants
(42,43), and measuring only pepsin in TA samples may
underscore the severity of aspiration in premature infants.
Our study is an observational study, lacking a control
group of premature infants who are not ventilated. Infants
were not randomized to feed or to receive xanthine therapy.
The decision to treat the infants with xanthine derivatives
was made by the attending neonatologist on service; this
decision may reflect the increased bradycardiac episodes
associated with GER, and the xanthine use may function as
a confounding factor.
Despite the above limitations, this is the first study
demonstrating significant aspiration of gastric contents
in ventilated premature infants. Chronic aspiration of
gastric contents may cause lung injury and is likely to
be important in the pathophysiology of BPD in
premature infants. Additional studies are required to
investigate the role of aspiration of gastric contents in
the development of BPD, its relationship with proinflammatory mediators and the effect of preventing
GER or microaspiration in premature infants.
Acknowledgments: The authors thank Charlene Martin, RN,
Nancy Markiewicz, RN, Valerie Gibson, RN, and Maurine
Remaly, RN, BSN, for their help in screening babies for enrollment and collecting tracheal aspirates. They also thank Kee Pyon,
PhD, for her help in analyzing data and reviewing the article.
REFERENCES
1. Newell SJ, Booth IW, Morgan ME, et al. Gastroesophageal reflux
in preterm infants. Arch Dis Child 1989;64:780Y6.
2. Ewer AK, Durbin GM, Morgan ME, et al. Gastric emptying and
gastroesophageal reflux in preterm infants. Arch Dis Child Fetal
Neonatal Ed 1996;75:F117Y21.
3. Peter CS, Sprodowski N, Bohnhorst B, et al. Gastroesophageal
reflux and apnea of prematurity: no temporal relationship. Pediatrics
2002;109:8Y11.
4. Omari TI, Barnett CP, Snel A, et al. Mechanisms of gastroesophageal reflux in healthy premature infants. J Pediatr 1998;
133:650Y4.
5. Omari TI, Rommel N, Staunton E, et al. Paradoxical impact of
body positioning on gastroesophageal reflux and gastric emptying
in the premature neonate. J Pediatr 2004;145:194Y200.
6. Omari TI, Barnett CP, Benninga MA, et al. Mechanisms of gastrooesophageal reflux in preterm and term infants with reflux
disease. Gut 2002;51:475Y9.
7. Siegel M. Gastric emptying time in premature and compromised
infants. J Pediatr Gastroenterol Nutr 1983;2:S136Y40.
8. Sheikh S, Stephen TC, Sisson B. Prevalence of gastroesophageal
reflux in infants with recurrent brief apneic episodes. Can Respir J
1999;6:401Y4.
9. Jolley SG, Halpern CT, Sterling CE, et al. The relationship of
respiratory complications from gastroesophageal reflux to prematurity in infants. J Pediatr Surg 1999;25:755Y7.
10. Herbst JJ, Minton SD, Book LS. Gastroesophageal reflux causing
respiratory distress and apnea in newborn infants. J Pediatr 1979;
95:763Y8.
11. Spray SB, Zuidema GD, Cameron JL. Aspiration pneumoni:
incidence of aspiration with endotracheal tubes. Am J Surg 1976;
131:701Y3.
12. Young PJ, Basson C, Hamilton D, et al. Prevention of tracheal
aspiration using the pressure-limited tracheal tube cuff. Anaesthesia 1999;54:559Y63.
13. Cucchiara S, Staiano A, Di Lorenzo C, et al. Pathophysiology of
gastroesophageal reflux and distal esophageal motility in children
with gastroesophageal reflux disease. J Pediatr Gastroenterol
Nutr 1988;7:830Y6.
J Pediatr Gastroenterol Nutr, Vol. 43, No. 3, September 2006
Copyr ight © Lippincott Williams & Wilkins. Unauthorized reproduction of this article is prohibited.
PEPSIN DETECTION IN TA FROM VENTILATED NEONATES
14. Knight PR, Davidson BA, Nader ND, et al. Progressive, severe lung
injury secondary to the interaction of insults in gastric aspiration.
Exp Lung Res 2004;30:535Y57.
15. Porembka DT, Kier A, Sehlhorst S, et al. The pathophysiologic
changes following bile aspiration in a porcine lung model. Chest
1993;104:919Y24.
16. Radford PJ, Stillwell PC, Blue B, et al. Aspiration complicating
bronchopulmonary dysplasia. Chest 1995;107:185Y8.
17. Fuloria M, Hiatt D, Dillard RG, et al. Gastroesophageal reflux in
very low birth weight infants: association with chronic lung
disease and outcomes through 1 year of age. J Perinatol 2000;20:
235Y9.
18. Mitchell DJ, McClure BG, Tubman TR. Simultaneous monitoring
of gastric and oesophageal pH reveals limitations of conventional
oesophageal pH monitoring in milk-fed infants. Arch Dis Child
2001;84:273Y6.
19. Grant L, Cochran D. Can pH monitoring reliably detect gastroesophageal reflux in preterm infants? Arch Dis Child Fetal
Neonatal Ed 2001;85:F155Y7.
20. Hopper AO, Kwong LK, Stevenson DK, et al. Detection of gastric
contents in tracheal fluid of infants by lactose assay. J Pediatr
1983;102:415Y8.
21. Kajetanowicz A, Stinson D, Laybolt KS, et al. Lipid-laden macrophages in the tracheal aspirate of ventilated neonates receiving
Intralipid: a pilot study. Pediatr Pulmonol 1999;28:101Y8.
22. Ward C, Forrest IA, Brownlee IA, et al. Pepsin-like activity in
bronchoalveolar lavage fluid is suggestive of gastric aspiration in
lung allografts. Thorax 2005;60:872Y4.
23. Ufberg JW, Bushra JS, Patel D, et al. A new pepsin assay to detect
pulmonary aspiration of gastric contents among newly intubated
patients. Am J Emerg Med 2004;22:612Y4.
24. Meert KL, Daphtary KM, Metheny NA. Detection of pepsin and
glucose in tracheal secretions as indicators of aspiration in mechanically ventilated children. Pediatr Crit Care Med 2002;3:19Y22.
25. Krishnan U, Mitchell JD, Messina I, et al. Assay of tracheal
pepsin as a marker of reflux aspiration. J Pediatr Gastroenterol
Nutr 2002;35:303Y8.
26. Vandenplas Y, De Wolf D, Sacre L. Influence of xanthines on
gastroesophageal reflux in infants at risk for sudden infant death
syndrome. Pediatrics 1986;77:807Y10.
27. Gupta GK, Cole CH, Abbasi S, et al. Effects of early inhaled
beclomethasone therapy on tracheal aspirate inflammatory
mediators IL-8 and IL-1ra in ventilated preterm infants at risk
for bronchopulmonary dysplasia. Pediatr Pulmonol 2000;30:
275Y81.
341
28. Munshi UK, Niu JO, Siddiq MM, et al. Elevation of interleukin-8
and interleukin-6 precedes the influx of neutrophils in tracheal
aspirates from preterm infants who develop bronchopulmonary
dysplasia. Pediatr Pulmonol 1997;5:331Y6.
29. Tullus K, Noack GW, Burman LG, et al. Elevated cytokine levels
in tracheobronchial aspirate fluids from ventilator treated neonates
with BPD. Eur J Pediatr 1996;155:112Y6.
30. Orenstein SR, Klein HA, Rosenthal MS. Scintigraphy versus pH
probe for quantification of pediatric gastroesophageal reflux: a
study using concurrent multiplexed data and acid feedings. J Nucl
Med 1993;34:1228Y34.
31. Metheny NA, Chang YH, Ye JS, et al. Pepsin as a marker for
pulmonary aspiration. Am J Crit Care 2002;11:150Y4.
32. Lopez Alonzo M, Moya MJ, Cabo JA, et al. Acid and non-acid
gastroesophageal reflux in newborns. Preliminary results using
intraluminal impedance. Cir Pediatr 2005;18:121Y6.
33. Valat C, Demont F, Pegat MA, et al. Radionuclide study of
bronchial aspiration in intensive care newborn children. Nucl Med
Commun 1986;7:593Y8.
34. Defize J. Development of pepsinogens. In: Lebenthal E ed.
Human Gastrointestinal Development. New York: Raven Press,
1989:299Y324.
35. Samloff IM, Liebman WM. Radioimmunoassay of group I pepsinogens in serum. Gastroenterology 1974;66:494Y502.
36. Fanarrof, Avroy: Neonatal-Perinatal Medicine, Neonatal Chronic
Lung disease, 6th ed, Vol 2. St Louis, MO: Mosby, 1997;1074Y89.
37. Jobe A. The new BPD: an arrest of lung development. Pediatr Res
1999;46:641Y3.
38. Metheny NA, Dahms TE, Chang YH, et al. Detection of pepsin in
tracheal secretions after forced small-volume aspirations of gastric
juice. JPEN J Parenter Enteral Nutr 2004;28:79Y84.
39. Dennish GW, Castell DO. Caffeine and the lower esophageal
sphincter. Am J Dig Dis 1972;17:993Y6.
40. Cohen S, Booth GH Jr. Gastric acid secretion and loweresophageal-sphincter pressure in response to coffee and caffeine.
N Engl J Med 1975;293:897Y9.
41. de Blic J, Midulla F, Barbato A, et al. Bronchoalveolar lavage in
children. ERS Task Force on bronchoalveolar lavage in children.
European Respiratory Society. Eur Respir J 2000;15:217Y31.
42. Mouterde O, Dacher JN, Basuyau JP, et al. Gastric secretion in
infants. Application to the study of sudden infant death syndrome
and apparently life-threatening events. Biol Neonate 1992;62:15Y22.
43. Gharpure V, Meert KL, Sarnaik AP, et al. Indicators of postpyloric feeding tube placement in children. Crit Care Med 2000;
28:2962Y6.
J Pediatr Gastroenterol Nutr, Vol. 43, No. 3, September 2006
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