Mechanical ventilation during experimental sepsis increases deposition of advanced glycation end products and myocardial inflammation

Kneyber, Gazendam, Niessen, Kuiper Jan-Willem, Dos, Slutsky, Plötz

Description

Increasing evidence links advanced glycation end products (AGE) including N ε -(carboxymethyl)lysine (CML) to the development of heart failure. Accumulation of AGE leads to myocardial inflammation, which is considered as one of the possible mechanisms underlying sepsis-induced cardiac dysfunction. We hypothesized that mechanical ventilation (MV) augmented sepsis-induced myocardial CML deposition and inflammation. Methods Sepsis was induced using a modified cecal ligation and perforation (CLP) technique in 36 male adult Sprague Dawley rats. Rats were randomized to four hours of MV with low tidal volume (LTV: 6 ml/kg, PEEP 5 cmH 2 O, n = 10) or high tidal volume (HTV: 15 ml/kg, PEEP 3 cmH 2 O, n = 10) 24 hours after the induction of sepsis. Eight rats served as septic, non-ventilated controls and eight as non-septic, non-ventilated controls. After 28 hours all rats were killed. The number of extravascular polymorphonuclear (PMN) leucocytes, macrophages, and lymphocytes was measured as the number of positive cells/mm 2 . The number of CML positive endothelial cells were semi-quantified based upon an intensity score. The CML intensity score was correlated with the number of inflammatory cells to study the association between CML depositions and inflammation. Results Gas exchange was comparable between the ventilated groups. Sepsis induced a significant increase in CML deposition in both ventricles that was significantly augmented by MV compared with non-ventilated septic controls (left ventricle 1.1 ± 1.0 vs 0.7 ± 0.1, P = 0.030; right ventricle 2.5 ± 0.5 vs 0.6 ± 0.1, P = 0.037), irrespective of ventilatory strategy. In the right ventricle there was a non-significant tendency towards increased CML deposition in the HTV group compared with septic, non-ventilated controls (1.0 ± 0.1 vs 0.7 ± 0.09, P = 0.07). Sepsis induced a significant increase in the number of macrophages and PMNs compared with non-ventilated septic controls that was augmented by MV, irrespective of ventilatory strategy. CML deposition was significantly correlated with the number of macrophages and PMNs in the heart. Conclusions Sepsis induces CML deposition in the heart with a predominant right ventricular inflammation that is significantly augmented by MV, irrespective of the ventilatory strategy.

Informations

• Publié par: biomed
• Publié le: 01 janvier 2009
• Nombre de lectures: 13
• Langue: English

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Available online

http://ccforum.com/content/13/3/R87

Vo l13
R
No 3
esearch
Open Access
Mechanical ventilation during experimental sepsis increases
deposition of advanced glycation end products and myocardial
inflammation
MartinCJKneyber
1,2,3
, RoelPGazendam
1,2
, HansWMNiessen
2,4,5
, Jan-WillemKuiper
1,6,2
,
ClaudiaCDos Santos
6
, ArthurSSlutsky
6
and FransBPlötz
1,2

1
Department of Pediatric Intensive Care, VU university medical center, P.O. Box 7057, 1007 MB Amsterdam, The Netherlands
2
Institute for Cardiovascular Research (ICaR-VU), VU university medical center, P.O. Box 7057, 1007 MB, Amsterdam, The Netherlands
3
Beatrix Children's Hospital/University Medical Center, P.O. Box 30001, 9700 RB Groningen, The Netherlands
4
Department of Pathology, VU university medical center, P.O. Box 7057, 1007 MB Amsterdam, The Netherlands
5
Department of Cardiac Surgery, VU university medical center, P.O. Box 7057, 1007 MB, Amsterdam, The Netherlands
6
Keenan Research Centre, Li Ka Shing Knowledge Institute, St. Michael's Hospital, 30 Bond Street, Toronto, Ontario, M5B 1W8; University of
Toronto, Toronto, Ontario, Canada
Corresponding author: MartinCJKneyber,[e-mail]
Received: 2 Apr 2009Revisions requested: 24 Apr 2009Revisions received: 2 Jun 2009Accepted: 9 Jun 2009Published: 9 Jun 2009
Critical Care
2009,
13
:R87 (doi:10.1186/cc7911)
This article is online at: http://ccforum.com/content/13/3/R87
© 2009 Kneyber
et al
.; licensee BioMed Central Ltd.
This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0),
which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract
Introduction
Increasing evidence links advanced glycation endinflammatory cells to study the association between CML
products (AGE) including N
ε
-(carboxymethyl)lysine (CML) to thedepositions and inflammation.
development of heart failure. Accumulation of AGE leads to
myocardial inflammation, which is considered as one of the
Results
Gas exchange was comparable between the ventilated
possible mechanisms underlying sepsis-induced cardiac
dysfunction. We hypothesized that mechanical ventilation (MV)groups. Sepsis induced a significant increase in CML
augmented sepsis-induced myocardial CML deposition anddeposition in both ventricles that was significantly augmented by
MV compared with non-ventilated septic controls (left ventricle
inflammation.1.1 ± 1.0 vs 0.7 ± 0.1,
P
= 0.030; right ventricle 2.5 ± 0.5 vs
Methods
Sepsis was induced using a modified cecal ligation0.6 ± 0.1,
P
= 0.037), irrespective of ventilatory strategy. In the
right ventricle there was a non-significant tendency towards
and perforation (CLP) technique in 36 male adult Spragueincreased CML deposition in the HTV group compared with
Dawley rats. Rats were randomized to four hours of MV with low
tidal volume (LTV: 6 ml/kg, PEEP 5 cmH
2
O, n = 10) or high tidalseptic, non-ventilated controls (1.0 ± 0.1 vs 0.7 ± 0.09,
P
=
0.07). Sepsis induced a significant increase in the number of
volume (HTV: 15 ml/kg, PEEP 3 cmH
2
O, n = 10) 24 hours aftermacrophages and PMNs compared with non-ventilated septic
the induction of sepsis. Eight rats served as septic, non-
ventilated controls and eight as non-septic, non-ventilatedcontrols that was augmented by MV, irrespective of ventilatory
controls. After 28 hours all rats were killed. The number ofstrategy. CML deposition was significantly correlated with the
extravascular polymorphonuclear (PMN) leucocytes,number of macrophages and PMNs in the heart.
macrophages, and lymphocytes was measured as the number of
positive cells/mm
2
. The number of CML positive endothelial
Conclusions
Sepsis induces CML deposition in the heart with
cells were semi-quantified based upon an intensity score. Thea predominant right ventricular inflammation that is significantly
CML intensity score was correlated with the number ofaugmented by MV, irrespective of the ventilatory strategy.

AGE: advanced glycation end product; BSA: bovine serum albumin; CLP: cecal ligation and perforation; CML: N
ε
-(carboxymethyl)lysine; HTV: high
tidal volume; LTV: low tidal volume; LV: light ventricle; MPO: myeloperoxidase; MV: mechanical ventilation; PaCO
2
: partial pressure of arterial carbon
dioxide; PaO
2
: partial pressure of arterial oxygen; PBS: phosphate-buffered saline; PEEP: positive end-expiratory pressure; PMN: polymorphonuclear
leucocytes; RAGE: receptor for advanced glycation end product; RV: right ventricle; sRAGE: soluble receptor for advanced glycation end product;
VCAM-1: vascular cell adhesion molecule-1.

Page 1 of 7
(page number not for citation purposes)

Critical Care
Vol 13 No 3 Kneyber
et al.

Introduction
Sepsis-induced cardiac dysfunction occurs in approximately
40 to 50% of patients with prolonged septic shock and is
associated with increased mortality. Various possible underly-
ing mechanisms have been reviewed extensively [1-6]. Myo-
cardial inflammation is one of these mechanisms as sepsis-
induced cellular infiltration of the myocardium has been found
in experimental studies and in humans dying from sepsis [7-9].
Advanced glycation end products (AGE) such as N
ε
-(car-
boxymethyl)lysine (CML) may play an important role in this
inflammation [10]. AGE are formed during oxidative stress,
acting as ligands for AGE receptors (RAGE) [11]. These
receptors trigger a cascade of signaling mechanisms with
subsequent expression of vascular cell adhesion molecule-1
(VCAM-1), induction of vascular leakage, and increased
chemotaxis of mononuclear phagocytes and release of pro-
inflammatory mediators resulting in cellular dysfunction [12-
15]. The soluble form of the receptor (sRAGE) was found to
be elevated in septic patients and associated with worsened
outcome [16].
The detrimental effect of AGE may be enhanced by mechani-
cal ventilation (MV) (double-hit principle). MV induces inflam-
mation of healthy lungs or aggravates pre-existing lung injury
(ventilator-induced lung injury); various mediators produced
during this inflammation including AGE formed during MV-
induced oxidative stress may contribute to distant organ fail-
ure, including the heart [17-21].
We hypothesized that sepsis led to myocardial CML deposi-
tion resulting in inflammation. In addition, we hypothesized that
this effect was augmented following four hours of MV. To test
this hypothesis we therefore designed a study in which rats
were subjected to two different ventilatory strategies in a
model of sepsis induced by cecal ligation and puncture (CLP).
Materials and methods
Animal preparation and experimental protocol
All animals were treated according to the Canadian national
guidelines and with approval of the Institutional Animal Care
and Use Committee of St Michael's Hospital. Sepsis was
induced in Sprague Dawley rats (weight ± 300 g; Charles Riv-
ers, St Constan, QC, Canada) using a modification of the
cecal ligation and perforation technique [22,23]. Twenty-four
hours later, rats were randomized to one of two strategies and
ventilated for four hours: low tidal volume (LTV) of 6 ml/kg and
positive end-expiratory pressure (PEEP) 5 cm H
2
O (n = 10);
or high tidal volume (HTV) of 15 ml/kg and PEEP 3 cm H
2
O (n
= 10). Normocapnia (partial pressure of arterial carbon dioxide
(PaCO
2
) 35 to 45 mmHg) was maintained by adjusting respi-
ratory rate. Inspiratory to expiration time was set to 1:2. The
fraction of inspired oxygen was 0.4 in both ventilated groups.
Anesthesia was maintained with intravenous xylazine 1 mg/kg/
hr and ketamine 20 mg/kg/hr; muscle relaxation was achieved

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by continuous intravenous administration of pancuronium bro-
mide (Sabex Inc, QC, Canada) 0.6 mg/kg/hr. For blood sam-
pling and arterial blood pressure measurements, a catheter
was inserted into the right carotid artery. All rats received a
continuous infusion of normal saline at a rate of 10 ml/kg/hr to
keep mean arterial pressure above 60 mmHg. At the end of the
experiment animals were sacrificed with an overdose of
anesthesia. Part of the lung was weighed and heated over-
night to determine lung wet-to-dry ratio.
Eight rats that underwent the CLP procedure were not sub-
jected to MV. Another eight rats were not subjected to CLP or
MV. All of these animals were sacrificed after 28 hours; the
first group served as non-ventilated septic controls, and the
second group as non-ventilated, non-septic controls. The
reported investigations were performed as part of experimen-
tal studies investigating the effects of MV during sepsis on
renal function.
Immunohistochemistry
Antibodies used were monoclonal mouse anti-rat CD68
(Serotec, Kidlington, UK), monoclonal mouse anti-rat CD45
(BD Pharmingen, Breda, The Netherlands), polyclonal rabbit
anti-human myeloperoxid