ORIGINAL ARTICLES
The impact of proton-pump inhibitors on intraperitoneal sepsis:a word of caution for transgastric NOTES procedures
Sonia L. Ramamoorthy Æ Jeffrey K. Lee Æ Yoav Mintz Æ John Cullen ÆMichelle K. Savu Æ David W. Easter Æ Alana Chock Æ Ravi Mittal ÆSantiago Horgan Æ Mark A. Talamini
Received: 6 December 2008 / Accepted: 1 May 2009 / Published online: 24 June 2009
� The Author(s) 2009. This article is published with open access at Springerlink.com
Abstract
Background During transgastric natural orifice translu-
minal endoscopic surgery (NOTES), there is an iatrogenic
perforation of the gastric wall with leakage of gastric
contents into the peritoneal cavity. The aim of this study is
to determine the effect of proton-pump inhibitors (PPI) and
alterations of gastric pH on infection during transgastric
surgery.
Methods Thirty 250-g male Sprague–Dawley rats were
divided into a study group (SG, n = 15) and a control group
(CG, n =15). SG were given 5 mg/kg pantoprazole for
3 days before procedure and another dose 1 h before. CG
received saline at similar time points. A mini-laparotomy
with gastrotomy was performed. Aspiration of 2.0 cc gas-
tric contents was removed from the stomach and injected
into the peritoneal cavity of both groups. Gastric pH and
peritoneal pH levels were obtained. Gastric aspirate was
sent for culture. White blood cell counts (WBC) were
obtained on postoperative days 1, 7, and 14, and C-reactive
protein (CRP) levels were obtained on postoperative day 1.
At day 14, a necropsy was performed and aerobic and
anaerobic cultures of the peritoneal cavity were obtained.
Results There were no deaths in either group. The aver-
age gastric pH in the SG was 5.13 versus 3.26 (p = 0.03) in
the CG. The average peritoneal pH was similar in both
groups. The WBC in the SG was 4.5 vs. 3.5 (1,000 cells/
mm) in the CG. There was no elevation in CRP levels in
either group. Bacterial cultures were positive in 3/15 (20%)
rats in the CG and in 9/15 (60%) in the SG (p = 0.008).
Intra-abdominal abscesses were found in 2/15 (13%) rats in
the CG and in 5/15 (33%) in the SG (p = 0.08).
Conclusions Pretreatment with a PPI resulted in a higher
rate of peritoneal bacterial contamination and abscess
formation. The acidic environment of the stomach appears
to be protective against infection when intraperitoneal
contamination occurs as a result of gastrotomy.
Keywords Transgastric surgery � Natural orifice surgery �NOTES � Proton-pump inhibitors
Transgastric surgery is a form of natural orifice surgery that
involves endoscopic gastrotomy followed by intraperito-
neal surgery and gastric closure. The first reported cases of
natural orifice transluminal endoscopic surgery (NOTES)
were appendectomies performed by Reddy and Rao in
India (personal communication with co-author, Talamini).
The field of transgastric surgery is cautiously emerging as
issues of safety and efficacy are investigated. A concern of
transgastric surgery is spillage of gastric contents during
transgastric passage. Much of the recent interest in ‘‘dual-
lumen’’ and ‘‘hybrid’’ approaches to NOTES procedures
stems from the concern over leakage and reliable closure.
Historically, the leakage of gastric contents into the
Supported by Ethicon/NOSCAR 2007 research grant
Presented at the Society of American Gastrointestinal and Endoscopic
Surgeons, Philadelphia, PA, 2008.
S. L. Ramamoorthy (&) � Y. Mintz � J. Cullen �M. K. Savu � D. W. Easter � S. Horgan � M. A. Talamini
Center for the Future of Surgery- Department of Surgery, UC
San Diego Medical Center, 3855 Health Sciences Drive, Suite
2073, La Jolla, CA 92093-0987, USA
e-mail: [emailprotected]
J. K. Lee � R. Mittal
Department of Medicine, San Diego Medical Center, University
of California, San Diego, CA, USA
A. Chock
Northwest Weight Loss Surgery, 125 30th St. SE, Everett, WA
98208, USA
123
Surg Endosc (2010) 24:16–20
DOI 10.1007/s00464-009-0559-3
peritoneal cavity is thought to lead to peritonitis from both
chemical and bacterial exposure. Much of these concerns
arise from the surgical experience with perforated gastro-
duodenal ulcers. In most cases, patients are in extremis
when presenting with perforated gastric disease and require
emergent exploration and repair. Recent studies however
point towards conservative management in a select popu-
lation of patients who are believed to have the perforation
‘‘sealed’’. This management includes the use of antisecre-
tory drugs such as proton-pump inhibitors (PPI), antibiotics
directed at H. Pylori, and bowel rest [1]. PPIs are also
prescribed for a variety of other reasons including reflux
disease and chronic gastritis, post bariatric surgery, and are
often used by patients on an intermittent basis for symp-
tomatic relief from any number of upper gastrointestinal
(GI) complaints [2]. In 2005, spending on PPIs exceeded
US $12 billion in the USA, making prescription PPIs the
second largest therapeutic drug class in terms of sales [3].
The impact of PPIs on transgastric NOTES procedures is of
particular interest as one of the anticipated benefits of
natural orifice surgery is to minimize the physiologic
impact of surgical intervention. The aim of this study
therefore is to determine the effect of PPIs and alterations
in gastric pH on infection during transgastric surgery.
Materials and methods
The following study was conducted after approval from
and under supervision of the University of California, San
Diego (UCSD) Institute for Animal Care and Use Com-
mittee (IACUC protocol #S07035).
Animals
Thirty male Sprague–Dawley rats (250–275 g) were
obtained from a US Department of Agriculture (USDA)-
approved vendor (Charles-Rivers Labs, Boston, MA). The
rats were acclimatized to their environment for 3–5 days
upon arrival, per IACUC protocol. During this period all
animals were caged per UCSD animal care protocol with
standard husbandry procedures, and given food and water
ad libitum, overseen by UCSD Animal Care. The animals
were kept in the institutional animal care facilities with
automated 12 h of light–dark cycle maintained at 25�C
room temperature.
Protocol
Animals were equally divided into a study group (SG, n =
15) and a control group (CG, n = 15). The study group
received subcutaneous pantoprazole (Wyeth Pharmaceuti-
cals Inc., Philadelphia, PA) at a dose of 5 mg/kg injected
subcutaneously under the neck fur every 8 h for 3 days and
1 h before the procedure. Standard PPI doses for rats vary in
the literature between 5 to 20 mg/kg [4, 5]. The current dose
was chosen to obtain noticeable change in gastric pH in this
rat model. The control group was injected in similar fashion
with sterile 0.9% normal saline at the same time points.
Preoperative care and anesthesia
The animals were weighed and given a study number. Three
to four hours prior to procedure animals were fasted, placed
in wire-based cages to prevent ingestion of bedding/stool/
food, and given access to water ad libitum. The animals were
transferred to a surgical laboratory facility on the day of the
procedure. Anesthesia was administered via a precision
vaporizer. Induction was obtained with 4–5% inhaled iso-
flurane (IsoFlo; Abbot Laboratories, North Chicago, IL) and
maintained with 2–3% isoflurane with 100% oxygen carry-
ing gas. Depth of anesthesia was determined by spontaneous
movement, twitching, increased respiratory rate, increased
work of breathing, movement of extremities upon stimula-
tion of plantar surface, and tail and toe pinch. After induc-
tion, the left upper quadrant was shaved using electric
clippers, and sterilely prepped and draped. At the minilapa-
rotomy incision site, 0.25% Marcaine 1 mg/kg (Astra-Zen-
eca, Wilmington, DE) was injected for pain control.
Surgery
After sterile draping of the animals, baseline serum com-
plete blood counts were obtained using ventral tail artery or
the lateral saphenous vein as access. Local anesthetic was
then administered to the incision site in the left upper
quadrant of the animal. A small (\1 cm) incision was made
using sterile instruments (Fig. 1). After gaining access to
the peritoneal cavity, baseline peritoneal pH levels were
measured using an Accumet AB15 Basic benchtop pH
meter (Fisher Scientific International Inc., Hampton, NH)
and a MI-508 pH microelectrode (Microelectrodes Inc.,
Bedford, NH). This was done by placing the probe in a
dependent portion of the peritoneal cavity posterior to the
liver such that the tip of the catheter is constantly bathed in
the small amount of peritoneal fluid present there. This
technique has been previously described by Hanly et al.
[6]. A reference electrode (Microelectrodes Inc.) was
inserted into the right ear. The system was calibrated
before each animal by immersing the tips of the pH and
reference electrodes in sterile commercially prepared buf-
fer solutions (Fisher Scientific, Fair Lawn, NJ) of pH 7.0
and pH 4.0. Once the baseline peritoneal pH values for
each animal were obtained, the stomach was exposed and
baseline gastric pH values (an average of three readings)
were recorded. Needle aspiration (16G) of gastric contents
Surg Endosc (2010) 24:16–20 17
123
was performed into a syringe containing 2 cc 0.9% normal
saline. As a control, a sample of the gastric/saline mix was
rechecked for changes in pH; none was noted. This mate-
rial was then injected into the peritoneal cavity of both the
study and control groups to mimic gastric spillage during
transgastric natural orifice surgery. The needle insertion
site in the stomach was closed using an absorbable stitch.
The fascia and muscle were closed using an absorbable
suture and the skin reapproximated using Indermil� tissue
adhesive. Animals were given buprenorphine 0.05 mg/kg
subcutaneously prior to recovery (Reckitt Benckiser Phar-
maceuticals, Richmond, VA.) for pain control. Total sur-
gery time ranged from 10 to 15 min. Animals were
recovered in postprocedure cage, with a heating source and
monitoring. Animals were monitored until they were able
to maintain sternal recumbency and were normothermic.
Recovery and postoperative observation
The animals were returned to feed ad libitum. White cell
count was drawn on postoperative days (POD) 1, 7, and 14.
Daily weights, eating habits, clinical signs of sepsis (pilo-
erection, abdominal tenderness, and lethargy), and bowel
movements were assessed on each rat. On POD 14, the rats
were sacrificed and subject to gross examination. Abscess
was defined as the presence of purulent material within the
peritoneal cavity at the time of necropsy.
Cultures
Aerobic and anaerobic cultures were sent from the gastric
aspirate and again at the time of necropsy. Cultures were
obtained by swab and delivered to the UCSD microbiology
laboratory within 1 h of procedure. The swabs were
applied to aerobic and anaerobic culture media using
standard CLIA-approved procedure. The swabs were
placed on appropriate agar plates and three streaks made
per plate. Growth on only the first streak, which is con-
sidered ‘‘scant’’ or (1?) culture, was quantified as\10–20
colony forming units (CFUs). Growth on streaks 2–3 is
considered moderate (2?) to heavy growth (3–4?). The
cultures were then speciated by UCSD microbiology lab-
oratory based on microscopic evaluation.
Statistical analysis
Descriptive statistics were used throughout. Nonparametric
Student’s t-test was used to analyze differences between
the control and study group. Statistical significance was
reported as those p values of less than 0.05.
Results
There were no deaths or leaks in either group. The average
intragastric pH in the study group was 5.13 versus 3.26 (p =
0.03) in the control group. The average intraperitoneal pH
was similar in both groups (7.09 versus 7.19, p = 0.34). The
white blood cell count in the study group increased by an
average of 4.5 versus 3.5 (1,000 cells/mm) in the control
group. There was no elevation in CRP levels in either
group. Bacterial cultures were positive in 3/15 (20%) rats
in the control group and in 9/15 (60%) in the group treated
with PPI (p = 0.008). Intra-abdominal abscesses were
found in 2/15 (13%) rats in the control group and in 5/15
(33%) in the group treated with PPI (p = 0.08). The most
common isolate in the rat cultures were E. coli, Proteus
mirabilis, and Staphylococcus xylosus. There was a higher
rate of E. coli infection ([3?) in the PPI treated group 8/9
(88%) compared with the control group 1/3 (33%) (Table 1).
Discussion
In this rodent model, we studied the effect of PPIs on intra-
abdominal sepsis after gastric spillage. From our results,
we saw a higher rate of bacterial contamination and abscess
formation in the study group. While our data for abscess
was not statistically significant, there appeared to be a trend
towards higher rate of infection. Of the cultures that were
positive, there was a higher rate of Gram-negative (enteric)
Fig. 1 Animal surgery: minilaparotomy in left upper quadrant
18 Surg Endosc (2010) 24:16–20
123
infection in the PPI-treated group versus the control group.
This data supports the conclusion of several studies on
rodents and humans that found an association between acid
suppression and bacterial overgrowth/infection [6–9]. In a
landmark study by Franklin et al., it was found that the
previously held notion of a ‘‘sterile’’ foregut was chal-
lenged by the finding of polymicrobial gastric flora. This
study was the first to show an increase in colony-forming
units (CFUs) and an increase in enteric pathogens as the
gastric pH increases [10]. Our study observed similar
findings with loss of comensural organisms such as Lac-
tobacillus, and increased growth of enterics such as E. coli
and Proteus species. More recently, the chronic use of PPI
has been associated with community-acquired Clostridium
difficile colitis [11] and a meta-analysis found an associa-
tion with acid suppression and enteric infection [12].
Similarly, in two retrospective human studies, the crude
odds ratio for development of spontaneous bacterial peri-
tonitis among PPI users versus nonusers ranged from 1.22
to 7.0 [8, 13]. A human study by Narula et al. found that
transgastric instrumentation contaminates the peritoneal
cavity and those patients on PPIs have a higher bacterial
load compared with nonusers [7]. In this small series of 50
patients, there was no observable adverse clinical outcome
as a result of a higher bacterial load; however these patients
were all treated with preoperative antibiotics. Our study
demonstrates similar clinical findings to those of Narula,
although with the added benefit of the animal model, we
were able to demonstrate a higher rate of peritoneal
infection with a trend towards significance (p = 0.08) in the
study group. Despite these findings, there was no obser-
vable difference between the groups with regards to sub-
jective or objective criteria such as behavior, activity level,
absolute neutrophil count or C-reactive proteins levels. Our
rat model raises the question as to whether the alterations in
gastric pH as a result of PPIs and subsequent changes in gut
flora are of clinical significance necessitating changes in
clinical practice.
The limitations of our study was that we worked with a
set ‘‘volume’’ of gastric aspirate that was chosen as a
percentage of total body weight of the rat model, i.e., 2 cc/
250 g rat = 320 cc/60 kg human. It is difficult to assess
whether an increase in bacterial load or ‘‘volume’’ would
have led to a different outcome, or whether one can
extrapolate human outcomes from a rodent model. In
addition, our experiment design could have more closely
approximated the clinical scenario of NOTES by using
open or closed gastrotomy with peritoneal contamination
instead of directly injecting gastric contents into the peri-
toneum, which may have caused an increase rate of abscess
formation.
The results of this study are suggestive of an increased
risk of intraperitoneal infection with proton-pump inhibi-
tors, but it remains unclear if simple measures such as
appropriate antibiotic coverage, aggressive sterile tech-
nique, and better clearance of secretions will reduce this
risk. More investigations are warranted to address this issue
before alterations in clinical practice can be recommended.
Open Access This article is distributed under the terms of the
Creative Commons Attribution Noncommercial License which per-
mits any noncommercial use, distribution, and reproduction in any
medium, provided the original author(s) and source are credited.
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Proteus Mirabilis 4? 4?
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Enterobacter sp. 1? 2?
Lactobacillus 4? 1?
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