Leukocyte filtration of blood cardioplegia
attenuates myocardial damage and inflammation†
Francesco Onorati*, Francesco Santini, Tiziano Menon, Enrico Amoncelli, Francesco Campanella,
Kostantinos Pechlivanidis, Giuseppe Faggian and Alessandro Mazzucco
Division of Cardiac Surgery, University of Verona Medical School, Verona, Italy
* Corresponding author. Division of Cardiac Surgery, University of Verona, Piazzale Stefani 1, 37126 Verona, Italy. Tel: +39-045-8123853; fax: +39-045-8123308;
e-mail: firstname.lastname@example.org (F. Onorati).
Received 20 October 2011; received in revised form 3 January 2012; accepted 5 January 2012
OBJECTIVES: Leukocyte filtration of blood cardioplegia (cLkF) is postulated to reduce ischaemia–reperfusion myocardial injury.
Contradictory results have been published and few studies have addressed perioperative cytokine leakage and haemodynamic status
METHODS: Thirty patients undergoing isolated aortic valve replacement were randomized to cLkF (cLkF-Group) or to standard cold
blood cardioplegia (S-Group). Troponin I (TnI) and lactate were sampled from the coronary sinus at reperfusion. Peripheral TnI and
lactate were collected preoperatively at admission, and in the intensive care unit (ICU) at 8, 12, 36 and 60 h postoperatively. Cardiac
index (CI), indexed systemic vascular resistances, cardiac cycle efficiency (CCE) and central venous pressure (CVP) were registered pre-
operatively, at admission to the ICU and at the 6th, 12th, 18th, 24th and 36th postoperative hour. IL-6, IL-8, TNF-alpha and IL-10 were
sampled preoperatively, at reperfusion, on admission to the ICU and the 6th, 18th and 24th postoperative hours.
RESULTS: The cLkF group showed lower TnI (2.4±0.4 vs. 5.1± 0.8 μg/l, P=0.0001) and lactate (0.9±0.1 vs. 1.6 ±0.2 mmol/l, P=0.0001)
from the coronary sinus at reperfusion. TnI levels (group-P= 0.0001, group× time-P< 0.0001) and lactate (group×time-P= 0.001)
remained lower postoperatively after cLkF. Ventricular defibrillation at aortic declamping was less common in the cLkF-Group (33.3% vs.
S-Group: 93.3%; P= 0.002). Cytokines demonstrated significant postoperative leakage (time-P=0.0001 in both groups for IL-6, IL-8, TNF-
alpha, IL-10), with lower pro-inflammatory (IL-6 group-P= 0.0001, group×time-P=0.0001; IL-8 group-P= 0.0001, group×time-P=0.007;
TNF-alpha group-P=0.0001; group× time-P= 0.012) and higher anti-inflammatory cytokine secretion after cLkF (IL-10 group-P= 0.005).
Perioperative haemodynamic indices proved to be similar between the two groups (group-P=NS for CI, SVRI, CCE and CVP).
CONCLUSIONS: cLkF during blood cardioplegia attenuates myocardial ischaemia/reperfusion injury and reduces perioperative leakage of
TnI, lactate and pro-inflammatory cytokines. These data did not result in a better haemodynamic status.
Keywords: Myocardial protection • Leukocyte filtration • Ischaemia–reperfusion injury • Inflammation
Since Buckberg and Calafiore demonstrated the superiority of
blood cardioplegia over crystalloid cardioplegia with regard to
oxygen and substrate supply, buffering systems and antioxidant
scavengers, its use has widened to become the most employed
solution to achieve myocardial protection in cardiac surgery .
Despite its well-recognized superiority, however, the presence of
leukocytes and platelets was soon proved able to worsen to
some extent myocardial ischaemia/reperfusion injury either by
direct tissue damage or by indirect micro-vascular plugging [1,
2]. Accordingly, leukocyte depleting filters were introduced to
reduce myocardial damage in view of their ability to selectively
catch circulating activated neutrophils and platelets [2, 3].
Different strategies, based on the location on the cardiopulmon-
ary bypass (CPB) system of the leukocyte-depleting filters and
the duration of filtration, have been suggested: accordingly, sys-
temic filters were used to remove activated leukocytes from the
systemic arterial line or from the venous line of the CPB, either
throughout the CPB or only during reperfusion after aortic
declamping; cardioplegic filters were employed to remove leu-
kocytes from blood cardioplegia throughout its administration or
only at reperfusion before aortic declamping; other dedicated
devices were tested to filter the salvaged blood or the allogeneic
blood transfusions regardless of the timing of CPB [2, 4–9];
finally, some authors have suggested the concept of ‘total leuko-
cyte control’ by a multilevel continuous filtering of the arterial
line, the cardioplegic line and of all the external blood products
. Despite encouraging findings in animal models, however ,
contradictory results were reached in the clinical practice, mostly
due to the heterogeneity of patients, interventions and strategies
†Presented at the 6th Biennial Meeting of the Society for Heart Valve Disease &
Heart Valve Society of America, Barcelona, Spain, 25–28 June 2011.
© The Author 2012. Published by Oxford University Press on behalf of the European Association for Cardio-Thoracic Surgery. All rights reserved.
European Journal of Cardio-Thoracic Surgery 43 (2013) 81–89
doi:10.1093/ejcts/ezs145Advance Access publication 30 March 2012
in cardioplegic leukocyte filtration (cLkF) usage, the presence of
coronary disease, the case mix in terms of myocardial metabolic
demands, cardiac diseases, indications, etc. . Furthermore, al-
though the majority of these studies focused on the biochemical
markers of myocardial damage (i.e. CPK, troponins) and on the
incidence of myocardialstunning/perioperative
infarctions, no one paralleled the latter outcome with actual
haemodynamic indices and/or with perioperative cytokine
leakage . Another variable systematically overlooked in previ-
ous studies concerns the hypertrophied heart, notoriously more
susceptible to ischaemia/reperfusion injury than the normal
heart during cardioplegic arrest and more prone to neutrophil-
mediated reperfusion damage afterwards . Accordingly, the
higher the ischaemic damage in the hypertrophied heart, the
more the potential benefit achievable by cLkF usage .
Therefore, it was the aim of this study to investigate the
impact of cLkF during cardiopulmonary bypass (CPB) on the
myocardial outcome of patients undergoing aortic valve replace-
ment for severe aortic stenosis, and therefore at increased risk of
myocardial ischaemia/reperfusion injury in view of the left ven-
tricular hypertrophy, expressed as biochemical markers of myo-
performance and pro-inflammatory and anti-inflammatory cyto-
MATERIALS AND METHODS
Patients and study design
Between April 2010 and January 2011, 30 consecutive patients
admitted to our institution because of degenerative calcified
severe aortic valve stenosis and scheduled for elective primary
isolated valve replacement, were enrolled in the study. The study
protocol was approved by the institution’s Ethical Committee/
Institutional Review Board. Informed consent was obtained from
To avoid the risk of bias related to jeopardized distribution of
cardioplegic solutions, patients with coronary disease were
excluded from the study. Other exclusion criteria included: age
<60 or >85 years, emergent/urgent/salvage procedures, left ven-
tricular ejection fraction <35% at preoperative echocardiography,
associated cardiac or vascular surgical procedures, redo surgery,
recent (<8 weeks) acute myocardial infarction (AMI), preopera-
tive IABP assistance, severe chronic obstructive pulmonary
disease (≥ stage IIIa of GOLD classification), obesity (BMI>30 kg/
m2), renal disease [Kidney Disease Outcome Quality Initiative
(KDOQI) class ≥2], previous irradiation, previous thoracic surgery,
previous transfusion (within 6 months), recent (<30 days) infec-
tions, liver dysfunction, ongoing steroids or statin therapy, drug
abuse, neurological diseases or recent (<3 months) stroke,
acquired/congenital deficits of the immune system, autoimmune
diseases or cancer. Owing to the tertiary nature of our hospital
with admission to our institution the same day of surgery for
elective cases, as those enrolled in the present study, patients
were randomized by lottery on the day of surgery (drawing pre-
prepared sealed envelopes containing the group assignment) to
either cLkF during CPB (15 patients, cLkF-Group) or standard
CPB without cLkF (15 patients, S-Group). With the exception of
the main investigator and the CPB technician, all other physi-
cians (surgeons, anaesthesiologists, intensivists, biochemists, etc.)
dealing with perioperative patient care, biological sampling and/
or collection and analysis of clinical data were blinded to the
group assignment until the end of the study.
Anaesthesia and surgery
Anaesthesia was standardized: anaesthetic induction consisted of
intravenous propofol infusion at 2 mg/kg combined with fentanyl
administration at 0.010 mg/kg. Propofol infusion (150–200 μg/kg
per min) and isoflurane (0.8% inspired concentration) maintained
anaesthesia in the operating room. Neuromuscular blockade was
achieved by 4 mg/h of vecuronium bromide, and lungs were
ventilated to normocapnia with a volume-controlled ventilation,
at a frequency of 12 per min, a tidal volume of 8 ml/kg (of ideal
body weight) and an FiO2of 0.5. A positive end-expiratory pres-
sure (PEEP) was set at 5 cm/H2O. During CPB, mechanical venti-
lation was discontinued. Arterial and central venous catheters
were the standard. During the stay in the intensive care unit
(ICU), the lungs were ventilated mechanically for at least 4 h after
surgery, with volume-controlled ventilation, at an initial fre-
quency of 12 per min, a tidal volume of 8 ml/kg of ideal body
weight and a PEEP maintained at 5 cm/H2O during the entire
study period. FiO2and ventilation rates were then adjusted to
keep PaO2> 120 mm/Hg and PaCO2between 30 and 35 mm/Hg.
Airway clearance was maintained by means of closed routine tra-
cheal suctioning. Anaesthesia was maintained with continuous
propofol infusion (180–200 μg/kg per min), plus i.v. fentanyl ad-
ministration if requested, until the start of weaning from mechan-
ical ventilation. Mechanical ventilation (FiO2and ventilation rate)
was adjusted during the stay in the ICU to keep acid–base
balance and PaCO2between 40 and 45 mm/Hg. Analgesia was
guaranteed by routine opioid administration.
Surgery was always performed through a median sternotomy.
Bioprosthetic or mechanical aortic valve replacement was
accomplished according to the conventional guidelines. In all
the cases, supraannular prosthetic anchorage was achieved by
interrupted pledgeted non-everting sutures. Management of
CPB was standardized. Heparin was given at a dose of 300 IU/kg
to achieve a target-activated clotting time of 480 s or above. Left
ventricular venting through the right superior pulmonary vein
and cardiotomy suction, until protamine administration, was
routinely employed. Returned blood was always reinfused to
the patient. Blood recovery with an autotransfusion device
(Autotrans Dideco, Mirandola, Modena, Italy) was also per-
formed intraoperatively in all cases. A level of haemoglobin
lower than 8 g/dl prompted blood transfusion. The extracorpor-
eal circuit was primed with 1400 ml of Ringer’s lactate solution
and 5000 IU of heparin. A non-pulsatile CPB flow was estab-
lished at 2.4 l min–1m–2. All patients were cooled to moderate
hypothermia ranging from 32 to 34°C. Cardiac arrest and myo-
cardial protection were accomplished by means of aortic cross-
clamping coupled with intermittent antegrade and retrograde
hyperkalemic cold blood cardioplegia (1:4 ratio), followed by a
retrograde ‘hot shot’ immediately before aortic declamping .
Repeated doses were administered via the sole retrograde route
every 20 min. Composition of the crystalloid solutions mixed
with blood in a 1:4 ratio for antegrade bolus, retrograde main-
tenance and pre-declamping ‘hot shot’ are reported in Table 1.
The CPB circuit was similarly standardized and included a Sorin
phosphorylcholine-coated tubing set (Sorin Group SpA, Milano,
Italy), a Jostra roller pump (Jostra, Maquet Cardiopulmonary,
Hirrlingen, Germany) and a hollow fibre membrane-coated
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