CordBlood and Heart Damage an 2009)

8/6/2019 CordBlood and Heart Damage an 2009)

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Cell Transplantation, Vol. 18, pp. 855868, 2009 0963-6897/09 $90.00 + .00Printed in the USA. All rights reserved. DOI: 10.3727/096368909X471170Copyright 2009 Cognizant Comm. Corp. E-ISSN 1555-3892

www.cognizantcommunication.com

Autologous Umbilical Cord Blood Mononuclear Cell

Transplantation Preserves Right Ventricular Function in a NovelModel of Chronic Right Ventricular Volume Overload

Can Yerebakan,* Eugen Sandica,* Stephanie Prietz,* Christian Klopsch,* Murat Ugurlucan,*Alexander Kaminski,* Sefer Abdija, Bjorn Lorenzen, Johannes Boltze,

Bjorn Nitzsche, Dietmar Egger, Malte Barten,# Dario Furlani,* Nan Ma,*Brigitte Vollmar,** Andreas Liebold,* and Gustav Steinhoff*

*Department of Cardiac Surgery, Medical Faculty, University of Rostock, Rostock, Germany

Department of Cardiology, Ruppiner Clinics, Neuruppin, Germany

Department of Diagnostic and Interventional Radiology, Medical Faculty, University of Rostock, Rostock, Germany

Fraunhofer Institute for Cell Therapy and Immunology, Translational Center of Regenerative Medicine, Leipzig, Germany

Vita 34 AG, Leipzig, Germany#Institute for Pathology, Medical Faculty, University of Rostock, Rostock, Germany

**Institute for Experimental Surgery, Medical Faculty, University of Rostock, Rostock, Germany

We aimed to evaluate the feasibility and efficacy of autologous umbilical cord blood mononuclear cell(UCMNC) transplantation on right ventricular (RV) function in a novel model of chronic RV volume over-load. Four-month-old sheep (n = 20) were randomized into cell (n = 10) and control groups (n = 10). Afterassessment of baseline RV function by the conductance catheter method, a transannular patch (TAP) wassutured to the right ventricular outflow tract (RVOT). Following infundibulotomy the ring of the pulmonaryvalve was transected without cardiopulmonary bypass. UCMNC implantation (8.22 6.28 107) in the cellgroup and medium injection in the control group were performed into the RV myocardium around the TAP.UCMNCs were cultured for 2 weeks after fluorescence-activated cell sorting (FACS) analysis for CD34antigen. Transthoracic echocardiography (TTE) and computed tomography were performed after 6 weeksand 3 months, respectively. RV function was assessed 3 months postoperatively before the hearts were

excised for immunohistological examinations. FACS analysis revealed 1.2 0.22% CD34+ cells within theisolated UCMNCs from which AcLDL+ endothelial cells were cultured in vitro. All animals survived sur-gery. TTE revealed grade IIIII pulmonary regurgitation in both groups. Pressurevolume loops under dobu-tamine stress showed significantly improved RV diastolic function in the cell group (dP/dtmin: p = 0.043; Eed:

p = 0.009). CD31 staining indicated a significantly enhanced number of microvessels in the region of UC-MNC implantation in the cell group (p < 0.001). No adverse tissue changes were observed. TAP augmen-tation and pulmonary annulus distortion without cardiopulmonary bypass constitutes a valid large animalmodel mimicking the surgical repair of tetralogy of Fallot. Our results indicate that the chronically volume-overloaded RV profits from autologous UCMNC implantation by enhanced diastolic properties with a proba-ble underlying mechanism of increased angiogenesis.

Key words: Tetralogy of Fallot; Right ventricular dysfunction; Pulmonary insufficiency;Umbilical cord blood; Stem cells

INTRODUCTION right ventricular outflow tract (RVOT) caused by ante-rior and leftward displacement of the infundibular sep-

tum. Hence, the closure of ventricular septal defect(s)Tetralogy of Fallot (TOF) is one of the most common

cyanotic congenital heart defects. Surgical repair is inev- and the resection of the hypertrophied trabecular muscle

bands in the right ventricle (RV) followed by transannu-itable and aims to normalize jeopardized pulmonary

blood flow by correcting the abnormal anatomy of the lar patching is the standard corrective surgery. However,

Received November 3, 2008; final acceptance April 6, 2009. Online prepub date: April 9, 2009.Address correspondence to Can Yerebakan, M.D., Department of Cardiac Surgery, Medical Faculty, University of Rostock, Schillingallee 35,18057, Rostock, Germany. Tel: +49 494 381 6101; Fax: +49 494 381 6102; E-mail: [email protected]

855


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856 YEREBAKAN ET AL.

additional infundibulectomy with RVOT enlargement prepared by the Institute of Laboratory Animal Re-

sources and the US National Institutes of Health.performed during the correction may predispose to RVOT

aneurysms, akinetic myocardial regions, and pulmonary Four-month-old domestic sheep (n = 20) were used.

Animals were randomly assigned to two groups as, theregurgitation with chronic right ventricular (RV) volume

overload. The latter, though well tolerated in the short cell group (n = 10, mean weight: 41.1 5.2 kg), which

received intramyocardial UCMNC implantation, and theterm, is the most dominant factor contributing to late

morbidity and mortality of patients (18). Currently, rou- control group, which received medium only (n = 10,

mean weight: 37.9 4.9 kg).tine and generous transannular patch type of repair has

therefore been abandoned. Rather limited RVOT patch-Autologous UCB Collection and Isolation of UCMNCsing with the preservation of pulmonary valve function,

if possible, is the preferred strategy (16). Nevertheless, Collection and cryopreservation of autologous ovine

UCB was performed by VITA 34 (Vita 34 AG, Leipzig,development of RV dysfunction in the long term still

remains a challenge and requires further animal and hu- Germany). After the collection of the UCB samples, di-

methyl sulfoxide was added and the UCB was storedman studies to establish protective strategies. To date,

however, the literature lacks a valid in vivo experimental in liquid nitrogen tanks with a temperature of 141C

according to standard operating procedures.model mimicking the postsurgical scenario of TOF pa-

tients. Mononuclear cells (MNC) were isolated from UCB

using the Pancoll (density 1.086 g/ml, PAN-BiothechRecent discoveries seem to overcome the historicalconsideration of the heart as a postmitotic organ, and the GmbH, Aidenbach, Germany) density gradient separa-

tion technique according to the manufacturers protocol.detection of residing cardiac stem cells able to regener-

ate damaged myocardium has fueled great attention for Thereafter, definite cell count was determined. Total

mononuclear cell suspension in IMDM of 1 ml volumecellular cardiomyoplasty (5,7,34,42). Although different

stem cells have been used in experimental and clinical was prepared as final product for autologous transplanta-

tion on the day of the operation.settings, the ideal source for stem cell harvesting is still

in debate. The ideal source of stem cells should possessFlow Cytometry of the Mononuclear Cell Fractionproperties such as genetic compatibility, pluripotency,and Cell Cultures From UCMNCsand, especially for cardiac therapies, the ability to gain

cardiomyocyte traits. Umbilical cord blood (UCB) is Only for quantification of antigen expression within

the isolated mononuclear cell fraction, fluorescence-acti-known to accommodate a considerable number of multi-

potent stem/progenitor cells (19,26,29,32,40) and can vated cell sorting (FACS, FACS Scan flow cytometer;

Becton Dickinson, San Jose, CA, USA) analysis waseasily be obtained during birth. Moreover, mononuclearcells isolated from the UCB have been successfully em- performed. Cells were incubated with antibodies against

CD34 (CD34 goat polyclonal IgG, Santa Cruz, Santaployed in experimental approaches for myocardial repair

and regeneration, which can be expanded in vitro and Cruz, CA, USA). Subsequently, donkey anti-rabbit Alexa-

Fluor 488 or secondary antibody (Invitrogen, Carlsbad,then transplanted (20,21,23,25,30,38).

Latest literature provides only a limited number of CA, USA) was applied. Samples lacking primary anti-

body incubation were used as negative controls.studies that investigate the effects of stem cells on RV

dysfunction (9,46). No experimental approach has been After isolation from umbilical cord blood the UCMNC

fraction was resuspended in endothelial media (MCDBconducted to study the potential of stem cell treatment

in case of RV volume overload. Hence, the aim of our 131 Medium; Sigma Aldrich) and transferred to a cul-

ture flask. After 14 days of cultivation in endothelialexperimental design was to evaluate the effects of intra-

myocardial transplantation of autologous umbilical cord media culture, AcLDL (low-density lipoprotein from hu-

man plasma, acetylated, Alexa-Fluor 488 conjugated;blood mononuclear cells (UCMNCs) on RV function in

a novel experimental surgical model of RV volume Molecular Probes) staining was performed according tothe manufacturers instruction. These investigations wereoverload in sheep.

instituted to define the cell surface markers within theMATERIALS AND METHODS

total mononuclear cell suspension, which was isolatedAnimals from the umbilical cord blood and was delivered intra-

myocardially.All procedures were approved by the local Animal

Care Committee of Mecklenburg/Vorpommern in Rostock,Surgical ProcedureGermany. Animals received humane care in compliance

with the Principles of Laboratory Animal Care formu- Animals were premedicated with an intramuscular in-

jection of 0.10.5 mg/kg xylazin (Rompun 2%, Bayerlated by the National Society for Medical Research and

the Guide for the Care and Use of Laboratory Animals Vital GmbH, Leverkusen, Germany) and 1020 mg/kg


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UCMNC TREATMENT IN CHRONIC RV VOLUME OVERLOAD 857

Ketamin 10% (Bela-Pharm GmbH & Co. KG, Vechta, and the pulmonary valve ring was transected bluntly

with scissors via the infundibulotomy without the needGermany). After intubation anesthesia was maintained

by inhalation of 1.5% to 2.5% isoflurane delivered through for extracorporal circulation.

a ventilator (Excel 210 SE, Ohnmeda-BOC Group, Mad- Experimental Designison, WI, USA). Oxygen was added to the respiratory

circle with the aim of a peripheral arterial saturation of Each animal underwent two surgeries. During theover 94%. Invasive arterial blood pressure was mea- first operation as described above, a TAP with infun-sured through right femoral artery line, central venous dibulotomy and a transsection of the pulmonary valvepressure was measured through the right jugular vein, ring with the aim of creating pulmonary regurgitationand peripheral arterial saturation was monitored with a were conducted following baseline hemodynamic mea-pulse oxymeter in a continuous fashion. surements with conductance catheters. Postprocedural

hemodynamic parameters were assessed to validate acuteCreation of Chronic Right Ventricular pulmonary regurgitation. Thereafter, 1 ml of UCMNCVolume Overload suspension for the animals of the cell group and 1 ml of

medium (10 injections of 0.1 ml) to each control animalA novel model for chronic RV volume overload

mimicking hemodynamic properties after the correction were injected intramyocardially into the RV free wall

below the inferior border of the transannular patch inof TOF was created by transannular patch (TAP) im-

plantation and pulmonary valve distortion in the RVOT two rows with a special self-constructed tuberculin sy-ringe with an epicardial stopper (a shorter 21-gauge nee-(Fig. 1af). For this purpose, a left anterior thoracotomy

was performed along the 5th or 6th intercostal space and dle slided over the original 30-gauge needle) to prevent

ventricular perforation and outflow of the cell sus-the RVOT was exposed. A TAP (Gelweave woven vas-

cular prosthesis, Vacutek Ltd., Terumo, Refrewshire, pension during injection. All animals received a pre-

operative single dose of 0.1 mg/kg body weight dexa-Scotland) was sutured from the infundibulum to the

main pulmonary artery reaching approximately 2 cm be- methasone (Dexa-ratiopharm, Ratiopharm GmbH, Ulm,

Germany) intravenously to prevent inflammation in-low and 2 cm above the pulmonary annulus on the

RVOT with continuous 5/0 polypropylene (Ethicon, duced by multiple intramyocardial injections as described

before by Borenstein and collegues (9). Although prolifer-Norderstedt, Germany). Thereafter, an incision in the

RVOT was performed through an opening in the patch ative effects of dexamethasone on UCMNC have been

Figure 1. Steps of the novel technique for the creation of the pulmonary insufficiency with TAP implantation. (a) Exposure of theRVOT through a left anterior thoracotomy. (b) Preoperative conductance catheter measurements via main pulmonary artery. (c)TAP implantation on the RVOT. (d) Completed TAP on the RVOT. (e) RVOT incision and pulmonary valve ring transsectionthrough an opening on the TAP. (f) Closure of the patch incision after side clamping of the TAP and postoperative conductancecatheter measurements. Scale bars: 1 cm.


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described in vitro (24), the single preoperative dose that values for volumetric analysis were obtained in triplicate

via the thermodilution method using a Swan-Ganz-cath-was instituted in our series in vivo did not target a prolif-

erative response of the administered cells. eter (Arrow International Inc., Reading, PA, USA). The

mean was saved for offline factor calibration. ParallelEchocardiography was performed 6 weeks after the

first operation to assess the grade of pulmonary regurgi- conductance was measured by using hypertonic saline

(injection of 5 ml of 10% NaCl into the right atrium).tation. Thirty-two-slice cardiac computed tomography

(CT) (Toshiba, Aquilion 32, Toshiba Medical Systems Thereafter a 23-mm balloon catheter (Fogarty Occlusion

Catheter, 8-22 F, Edward Lifesciences LLC, Irvine, CA,Corp., Tochigi, Japan) was performed for the detection

of any adverse tissue formation (e.g., calcification, tu- USA) was advanced through the right atrium into the

inferior vena cava for standardized preload reductionmor formation) 1012 weeks postoperatively. The blinded

radiologists were asked to indicate any presence and lo- maneuvers. A series of three caval occlusions of 10

pressurevolume loops was gained during apnea. Maxi-cation of possible myocardial calcifications and tissue

changes. mum pressure (Pmax), end-diastolic pressure (EDP), end-

diastolic and end-systolic volume (EDV, ESV), ejectionThe second operation was performed 12 weeks after

the first operation. Online pressurevolume loop analy- fraction (EF), cardiac output (CO), stroke volume (SV),

maximal slope of systolic pressure increment (dP/dtmax),sis was again conducted for each animal under baseline

and dobutamine stress conditions (310 g/kg/min, tar- diastolic pressure decrement (dP/dtmin), end-diastolic

pressure volume relation (EDPVR), and end-systolicget heart rate of 200210/min). pressure volume relation (ESPVR) slopes (Eed and Ees, Analysis of Myocardial Function respectively), and preload recruitable stroke work (PRSW)

were determined. We applied a volume intercept at aPressureVolume Loops. A combined pressurefixed pressure within the pressure range [20 mmHg forvolume conductance catheter 5 F (Millar Instruments,baseline measurements (V20) and 40 mmHg for dobutam-Houston, TX, USA) was inserted into the RV through aine stress examinations (V40), respectively], thus avoid-small stab wound in the pulmonary artery. The conduc-ing the insecurity of a linear extrapolation of ESPVR totance catheter was connected to one pressurevolumezero pressure. Accordingly, for the positioning of thetransducer system for pressure (Millar MPVS 300, EMKAEDPVR, we used a pressure intercept at a fixed volumeTechnologies, Paris, France) and another volume trans-within the volume range to evade linear extrapolation ofducer system for volume analysis (Sigma 5 DF, CDEDPVR to zero volume [40 ml for baseline conditionsLeycom, Zoetermeer, The Netherlands). Transducer sys-(P40) and 20 ml for dobutamine conditions (P20)] (14).tems were linked to the Millar PowerLab data acquisi-

tion hardware (Type ITF 16, EMKA Technologies) and Echocardiography. Transthoracic echocardiography(Vivid, GE Healthcare, Milwaukee, WI, USA) was per-real-time signal processing was performed by IOX

1.8.3.20 software (EMKA Technologies). The reference formed by a cardiologist blinded to the distribution of

Figure 2. Representative FACS plot of CD34+ antigen (A) and subsequent AcLDL+ cells (B) after14 days of cultivation in endothelial cell medium. Red line: positive, black line: control. M1:gating strategy.


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Table 1. Pressure-Volume Loop Measurements With Comparison of the Right Ventricular Function Between the Cell

and Control Groups and Within Each Group for Different Time Points

Day 90

Day 0 Day 0 Day 90 After TAPI

Before TAPI After TAPI After TAPI DobutamineBaseline Baseline Baseline Stress

Control Cell Control Cell Control Cell Control Cell

Parameter (n = 9) (n = 9) (n = 9) (n = 9) (n = 8) (n = 7) (n = 8) (n = 7)

Pmax (mmHg) 26.87 26.82 31.01 29.91 30.43 26.86 46.51 54.12

1.70 1.39 1.64 0.97 3.17 2.29 3.69 5.00

p 0.008* 0.023 0.383* 0.938

p n.s. 0.278 0.235

EDP (mmHg) 11.56 10.08 15.69 12.04 9.38 11.65 12.13 9.53

1.06 0.97 1.33 0.86 0.50 2.33 1.72 1.48

p 0.016* 0.055 0.148* 1.000

p n.s. 0.852 0.279

dp/dtmax (mmHg /s) 415.08 439.19 476.85 508.26 432.26 326.83 1512.79 1881.98

25.21 35.79 43.24 26.08 74.95 47.06 84.81 135.06

p 0.039* 0.039 0.945* 0.078

p n.s. 0.271 0.033

dp/dtmin (mmHg/s) 310.11 329.30 331.37 324.42 246.14 254.44 665.49 902.89

15.49 30.95 25.33 18.28 28.63 25.32 44.94 101.61

p 0.148* 0.641 0.055* 0.078

p n.s. 0.834 0.043

EDV (ml) 67.76 57.12 103.89 90.24 47.81 49.87 27.94 31.09

6.00 2.67 10.83 17.57 7.07 6.67 3.87 1.69

p 0.008* 0.023 0.055* 0.109

p n.s. 0.837 0.491

ESV (ml) 18.93 14.46 43.18 38.86 19.86 18.18 9.45 7.87 3.66 2.47 7.38 9.28 4.84 3.85 2.78 1.30

p 0.008* 0.008 0.641* 0.297

p n.s. 0.794 0.297

EF (%) 72.74 75.40 58.51 54.64 61.56 66.87 69.48 74.75

4.21 3.41 4.35 4.21 4.73 5.10 4.85 4.41

p 0.008* 0.008 0.078* 0.156

p n.s. 0.459 0.441

CO (ml/min) 4273.05 3760.21 5583.95 4955.98 2925.27 3102.04 3809.28 5044.78

349.60 123.40 651.24 1109.70 363.41 269.34 340.04 444.19

p 0.023* 0.078 0.008* 0.016

p n.s. 0.709 0.094

SV (ml) 48.90 42.65 60.76 51.37 27.97 31.70 18.50 23.28 4.86 1.97 7.07 9.58 2.71 3.08 1.67 1.83

p 0.055* 0.195 0.008* 0.031

p n.s. 0.378 0.075

PRSW (ml mmHg) n.a. n.a. n.a. n.a. 16.37 12.41 23.83 26.23

2.57 1.54 2.84 2.69

p 0.224 0.553

Eed (mmHg/ml) n.a. n.a. n.a. n.a. 0.21 0.17 0.67 0.29

0.08 0.04 0.10 0.07

p 0.731 0.009


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Table 1. Continued

Day 90

Day 0 Day 0 Day 90 After TAPI

Before TAPI After TAPI After TAPI Dobutamine

Baseline Baseline Baseline Stress

Control Cell Control Cell Control Cell Control Cell

Parameter (n = 9) (n = 9) (n = 9) (n = 9) (n = 8) (n = 7) (n = 8) (n = 7)

P40 (mmHg) n.a. n.a. n.a. n.a. 5.07 5.67 n.a. n.a.

2.16 1.03

p 0.809

P20 (mmHg) n.a. n.a. n.a. n.a. n.a. n.a. 12.36 7.50

1.69 1.29

p 0.044

Ees (mmHg/ml) n.a. n.a. n.a. n.a. 1.20 0.85 2.78 2.48

0.18 0.21 0.64 0.38

p 0.228 0.699

V20 (ml) n.a. n.a. n.a. n.a. 27.10 30.93 n.a. n.a.

5.53 4.22

p 0.600

V40 (ml) n.a. n.a. n.a. n.a. n.a. n.a. 14.62 13.67

3.04 3.89

p 0.829

Values are presented as mean SEM. n.s.: no significance; Vx, volume intercept for a given pressure of x; Px, pressure intercept for a given volumeof x.*Versus control day 0 before transannular patch implantation (TAPI) under baseline conditions.Versus cell day 0 before TAPI under baseline conditions.Control versus cell day 90 after TAPI under baseline conditions.Control versus cell day 90 after TAPI under dobutamine stress conditions.

the groups 6 weeks postoperatively. Parasternal long and puter-assisted planimetry at the region of TAP in eight

different sections for each animal with 20 different high-short axis views were obtained with both M-mode and

two-dimensional echocardiography images. Right ven- power fields (HPF) of 0.216 mm2 per section.

Immunohistochemical staining with CD-31 antibodytricular end-diastolic and end-systolic volume (RVEDV,

RVESV) as well as right and left ventricular ejection [PECAM-1(M-20) goat polyclonal IgG; Santa Cruz Bio-

technology, CA, USA] followed by donkey anti-goatfraction (RVEF, LVEF) were determined. The grade of

pulmonary insufficiency was scaled in four categories Alexa-Fluor 568-conjugated secondary antibody (Invitro-

gen) was performed around the region of the TAP andas grade I (minimal), II (mild), III (moderate), and IV

(severe). cell/medium injection for comparison of capillary den-

sity between the groups. Sections were then counter- Morphological and Histological Studies stained with DAPI. Four sections of the RV myocardium

of each animal along the TAP implantation region andAfter the measurements in the second surgery heartswere arrested with potassium chloride and rapidly ex- further four sections 1 cm remote from the area of TAP

were analyzed using confocal microscopy. Twenty high-cised. The postmortem RVOT preparations were photo-

graphed for macroscopic assessment. Paraffin-embedded power fields (0.216 mm2) in each section were randomly

selected, and the number of capillaries in each field was10-m sections of the RVOT from the region of UCMNC

transplantation were used for immunohistochemistry. averaged and expressed per mm2.

Hematoxylin-eosin, Goldner, and Kossa stainings wereStatistical Analysisapplied for histological investigation of the myocardium

with regard to morphology of the myocytes, integrity of Data are presented as mean SE. Statistical analysis

was carried out with the SPSS software package (SPSSthe myocardium, detection of fibrosis, and calcifications.

The area of fibrosis (in m2) was analyzed with com- Inc). For time- and procedural-dependent comparison


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between pre- and postoperative RV function within the We were able to create standardized acute pulmonary

regurgitation in all animals with the novel experimentalgroups, the Wilcoxon signed-rank test was chosen. For

overall comparison between the experimental groups surgical technique, which was confirmed by conduc-

tance catheter measurements (see below). In the celldata were subjected to one-way analyses of variance

(ANOVA) method that applies post hoc multiple Holm- group a mean number of 8.22 6.28 107 UCMNC was

successfully transplanted into the RV myocardium aroundSidak tests, the nonparametric Kruskal-Wallis (failing

normality), or post hoc multiple Dunn tests. Values of the area of TAP implantation. In the control group me-

dium was injected in the same way and in the region ofp < 0.05 were considered statistically significant.

RV myocardium. The mean weight of the animals be-

fore the second operation was 40.8 3.5 and 39.7 4.3RESULTSkg, in the cell and control groups, respectively.

Experimental Outcome

No mortality was registered due to the surgical proce- FACS and Cell Culturedure. Three animals died postoperatively. In the cell FACS studies revealed 1.2 0.22% CD34+ cells ofgroup, one animal suffered from severe pneumonia and the total mononuclear cell fraction as the end productdied on the 11th postoperative day, and the other died after isolation from umbilical cord blood before intra-during the transport to the CT investigation, probably myocardial transfer. After 14 days of cultivation ofdue to aspiration following premedication. One animal

UCMNC in endothelial cell medium we were able toin the control group did not survive late pericardial tam- detect cell cultures showing growth of AcLDL+ endothe-ponade in the second postoperative week. lial cells (Fig. 2). This finding confirmed the existence

of the CD34+ cell fraction within the UCMNCs.

Right Ventricular Functional Analysis

Table 1 contains the data from the monitored hemo-

dynamics during a follow-up period of 90 days. Prior to

valvular injury and TAP implantation, RV catheteriza-

tion did not reveal any significant difference between

animals in both experimental groups.

Early Postoperative Right Ventricular Functions

at Day 0

Systolic Functions. RVEF following RVOT incision

and TAP implantation decreased 20% and 28% in con-

trol and cell groups, respectively (both p = 0.008). DP/

dtmax was significantly higher when compared to preop-

erative values in both groups (control group: 15%, cell

group: 16%; both p = 0.039). Right ventricular Pmax in-

creased 15% in the control group (p = 0.008) and 12%

in the cell group (p = 0.023) postoperatively. Right ven-

tricular ESV following the surgical procedure was 2.3-

fold and 2.7-fold increased in control and cell groups

(both p = 0.008), confirming the RV volume overload

due to pulmonary regurgitation. Although there was asignificant 31% increase in CO in the control group

(p = 0.023), the 32% increase in UCMNC group hearts

did not yield a statistically significant change (p = 0.078).

Furthermore, SV was augmented but the differences

were not significant in both groups when compared to

the preoperative values. RV early postoperative systolic

functions were similar in both groups.Figure 3. (A) Slope of end-diastolic pressure volume relation(Eed) and (B) maximal slope of diastolic pressure decrement

Diastolic Functions. Right ventricular EDV, the(dp/dtmin) after 3 months of follow-up under dobutamine stressmain indicator of ventricular volume load, significantlyconditions in comparison of cell and control groups (n = 7 and

n = 8, respectively). *p < 0.05. increased postoperatively in both groups (control group:


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Figure 4. Caval vein occlusions performed 3 months postoperatively reflected the contractility and elasticity of the RV occupyingsurgical PR. The curves of end-diastolic pressure volume relation (EDPVR, green) and end-systolic pressure volume relation(ESPVR, red) were determined under dobutamine-induced stress conditions. The pressure intercept of EDPVR (P20 at a fixedvolume of 20 ml) and its slope significantly reduced under dobutamine stress conditions in the cell group (solid loops) whencompared with the control group (dashed loops). *p < 0.05.

53% increase, p = 0.008; and cell group: 58% increase, Long-Term Right Ventricular Functions at Day 90

p = 0.023). Further, EDP was significantly elevated by

36% in control animals (p = 0.016) after surgery, Systolic Functions. Under baseline conditions in

both groups CO and SV were found to be significantlywhereas the 19% increase found in the cell group was

not found to be significant (p = 0.055). DP/dtmin was not lower than preoperative levels at day 0 (CO in control

group: 32%, p = 0.008; CO in cell group: 18%, p =significantly different from preoperative values in both

groups. Again, the comparison of postoperative parame- 0.016; SV control group: 43%, p = 0.008; SV cell group:

26%, p = 0.031). Remaining monitored parameters EF,ters between the groups was outside of significant vari-

ance. dP/dtmax, Pmax, and ESV were not significantly different


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than the preoperative findings in both groups. However, cantly reduced by 39% (p = 0.044) and 57% (p = 0.009),

respectively (Figs. 3 and 4).although the derangement of right ventricular CO and

SV seemed stronger in the control group than in the cellTransthoracic Echocardiography After 6 Weeksgroup, the comparison of systolic functions in between

both groups at day 90 did not expose any significantEchocardiography revealed pulmonary regurgitationdifferences under baseline conditions. Similarly under in all operated animals. Table 2 shows the absolute val-

dobutamine stress conditions CO and SV elevations in ues of RVEDV, RVESV, the comparison of RVEF andcell group compared with the control group were again LVEF, and pulmonary insufficiency grades in bothnot statistically significant (CO: 32%, p = 0.094; SV: groups. Pulmonary insufficiency grading confirmed a26%, p = 0.075). In contrast, pharmacologically induced similar extent of pulmonary valve injury when bothstress augmented the speed of contraction in the cell study groups are compared (control group: 2.50 0.33;group more prominently when compared to the control cell group: 2.88 0.30; p = 0.19). Figure 5 provides agroup (dP/dtmax: 24%, p = 0.033). visualization of the regurgitant flow in the region of the

TAP. Diastolic Functions. Catheterization after 90 days

follow-up reflected no significant differences in EDPCardiac Computed Tomographyand EDV in both groups under baseline conditions when

compared with preoperative findings at day 0. More- No adverse tissue changes were detected via cardiac

over, both groups revealed moderate decreases in dP/ computed tomography before the second operation. Wedtmin that did not reach statistical significance late after did not add RV functional volumetric parameters thatRVOT incision and TAP implantation (control group: were gained from the CT because breath-holding epi-21%, p = 0.055; cell group: 23%, p = 0.078). Similar to sodes needed for quantitative analysis would have ex-the systolic evaluations, a comparison of diastolic func- posed the animals to a greater risk for hemodynamictions between cell group and control group at day 90 compromise. Therefore, CT data did not reveal represen-did not uncover significant differences under baseline tative values for the assessment of RV function.conditions. Again, dobutamine-induced stress led to sig-

Macroscopic and Microscopic Examinationsnificantly greater speed of relaxation in the cell group

than in the control group (dP/dtmin: 36%, p = 0.043). Macroscopy. All hearts occupied a pulmonary valveESPVR and EDPVR gained by the occlusion of the

annulus defect as expected from echocardiography andvena cava inferior revealed the contractility and elastic-

cardiac CT. At this region fibrinous, reendothelializedity indices of the right heart late after surgical PR induc-

tissue covered the TAP. Further, none of the removedtion. Drawing curvilinear ESPVR also causes a source hearts provided evidence for intravascular thrombus orof error (11). However, the adequate quantification of

tumor formations in the area of interest. Figure 6A rep-the volume intercept of ESPVR within the pressure

resents typical tissue formations and RV organic shaperange did not reveal significant differences between the

late after RVOT incision and TAP implantation. The RVexperimental groups under baseline (V20) and dobutam- sections that were used for microscopic investigationsine stress (V40) conditions. Likewise, Ees did not vary are shown in Figure 6B.between the groups. On the contrary, under dobutamine

Microscopy. The analysis of the area of fibrosis atstress conditions EDPVR shifted rightward downwardtwo different levels did not expose any significant dif-in the cell group compared with the control group. Theferences between the experimental groups at the area ofpressure intercept of EDPVR (P20) and Eed were signifi-TAP (control group: 5523.07 653.43 m2, cell group:

6256.99 1143.99 m2, p = 0.575), and 1 cm away

from the area of TAP (control group: 3826.16 323.87

Table 2. Right and Left Ventricular Function m2, cell group: 3306.87 472.88 m2, p = 0.371). Onin the Transthoracic Echocardiography the contrary, capillary density in the cell group was dra-6 Weeks Postoperatively

matically enhanced at both levels when compared with

the control group at the area of TAP (control group:Cell Control738.83 10.45 capillaries/mm2, cell group: 846.32 (n = 6) (n = 6) p22.71 capillaries/mm2, p < 0.001), and 1 cm away from

RVEDV (ml) 26.6 7.0 26.1 9.4 n.s. the area of TAP (control group: 721.76 10.88 capillar-RVESV (ml) 7.2 3.8 9.0 2.6 n.s. ies/mm2, 918.88 17.69 capillaries/mm2, p < 0.001)RVEF (%) 73.9 9.3 62.6 12.0 n.s. (Fig. 7). Again no adverse tissue changes could be de-LVEF (%) 72.7 3.2 60.3 17.7 n.s.

tected using microscopic and macroscopic investigationsPulmonary insufficiency grade 2.9 0.3 2.5 0.3 n.s.

after 3-month follow-up.


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Figure 5. Representative echocardiographic image showing pulmonary regurgitant flow 6 weeksafter surgery.

DISCUSSION tant improvement of myocardial performance (6,34,35,

42). Unfortunately, congenital cardiac anomalies have

widely been estranged from cell-based cardiac regenera-Diseased myocardium has been one of the most at-

tractive targets of novel regenerative approaches in the tive approaches.TOF belongs to the most common cyanotic congeni-current era. The delivery of stem cells to the myocar-

dium has successfully been performed in a considerable tal heart defects. Although surgical correction reveals

superior early postoperative results (2), chronic pulmo-number of experimental and clinical trials with a resul-

Figure 6. Macroscopic images of the RVOT postmortem 3 months after TAP implantation. (A) Cross-sectional image through theTAP on the RVOT and pulmonary artery (PA) with presentation of successful valvulotomy (*) of the pulmonary valve (PV) andcreation of valvular leakage below the TAP. (B) View from the RV side on the RVOT after TAP implantation. Sections were madein the RVOT at the site of the UCMNC implantation for histological studies.


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those patients with more severe pulmonary regurgita-

tion, may contribute to higher morbidity and mortality

in these patients (47).

The employment of stem cell therapies in the comple-

mentary treatment of congenital heart defects bears ma-

jor potential for regenerative medicine in addition to sur-

gical therapies (43). UCB is rich in stem/progenitor cells

with enhanced potency for angiogenic and myogenic

differentiation and superior proliferative characteristics

(19,33,36). These cells have already been therapeutic

agents in patients suffering from major hematological

disorders (45). UCMNCs have also been used experi-

mentally in several different settings for myocardial re-

generation. These cells were administered systemically

or locally in settings for the treatment of myocardial in-

farction with improvement in cardiac function, reduction

of infarction size and elevation of capillary density (20,

21,30). Recently, immunoselected human UCB CD133+

cells have been reported to possess endothelial and cardio-

myogenic properties expressing VE-cadherin, CD146, or

muscle proteins such as troponin I and myosin ventricu-

lar heavy chains in vitro (8).

Very early efforts aiming at an experimental enlarge-

ment of the pulmonary or aortic orifices were reported

in 1914 by Carrel et al. (12). Our model constitutes a

novel experimental model of pulmonary orifice enlarge-

ment and pulmonary valve distortion without using ex-

tracorporal circulationa first in the literature. To the

best of our knowledge, the present study is one of the

very first investigations in the field combining myocar-

dial regenerative therapy with autologous UCMNCs, theRV and congenital cardiac surgery in a large animal

model. UCMNCs were successfully isolated and trans-

planted into RV myocardium around the transannular

patch and the site of infundibulotomy. Confirming pre-

vious studies using human umbilical cord blood, about

1% of the total sheep UCMNCs expressed CD34, which

is known to be a marker for hematopoietic stem cells,

satellite cells, and endothelial progenitor cells. The con-

tent of endothelial progenitor cells in UCMNCs was alsoFigure 7. Representative confocal microscopic images from

supported by the cultures grown in vitro. By applyingthe cell group (A) and from the control group (B) for the pre-

special endothelial media to the final cell product wesentation capillary density with CD31 staining of the myocar-were able to show a considerable growth of AcLDL+dial regions around the transannular patch where the UCMNC

(cell group) and medium (control group) injections were insti- endothelial cells that originated from UCMNCs.tuted [630 magnification, CD31 antibody (green) for endo- Early conductance catheter analysis revealed alteredthelial cells, nuclear counterstaining with TO-PRO-3 iodide

ventricular volume parameters and acute stress with an(Invitrogen, Carlsbad, CA, USA) (blue)].

enhancement of systolic properties due to acute pulmo-

nary regurgitation, which corroborates with other reports

in the setting of acute volume overload (27,37). Echo-nary regurgitation with late RV dysfunction mainly de-

termines long-term morbidity and mortality of patients cardiography after 6 weeks indeed confirmed the suc-

cess of our experimental strategy by showing pulmonarydespite advanced surgical techniques (4). Patients are

candidates for surgical reintervention due to worsening insufficiency between grades II and III in all operated

animals. The visualization and adequate functionalRV function with a decline in clinical performance, but

also malignant ventricular arrhythmias, especially in quantification of the RV via transthoracic echocardiog-


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raphy is known to be challenging. Therefore, the volu- dynamic analysis. The evidence for a lasting pulmonary

insufficiency of grade IIIII in the echocardiography atmetric parameters gained from the echocardiographic

evaluation should receive cautious acceptance. At the 6 weeks and our postmortem macroscopic investigations

showing clear signs of pulmonary annular distortion inend of the 3-month follow-up we were already able to

observe an alteration of the RV function, which is one all animals contradicts the idea of the resolution of the

pulmonary incompetence. A possible explanation wouldof the main long-term determinants of morbidity and

mortality after TOF correction. Even when the derange- be early remodeling or structural adaptation of the RV

resisting against spatial dilatation (13).ment for CO and SV seemed more prominent in the con-

trol group than in the cell group baseline cardiac func- We observed significantly enhanced capillary forma-

tion in the cell group, which yielded the possible expla-tion did not differ significantly between the groups.

Under dobutamine stress conditions, however, hemody- nation of better diastolic function achieved by cell trans-

plantation. The detection of endothelial cells in thenamic assessment of the cell group uncovered a better

systolic function of the RV with significantly enhanced cultivation of UCMNCs supports the potential role of

these cells in the elevation of angiogenesis. This findingvalues for dP/dtmax compared to the controls. Szabo et al.

conducted an experimental chronic RV volume overload indicates potential importance for RV remodeling after

infundibulotomy and is in concert with other reports instudy by creating a femorofemoral arteriovenous shunt

in dogs for 3 months (44). The authors observed that the the literature (21,30). In our study the area of fibrosis

did not show any significant difference between thechronic volume overload had no effect on RV contractil-ity whereas the inotropic response to an increased after- groups. The average area of fibrosis around the TAP

was about 2.7% in both groups, which is a low valueload was limited. However, the setting of our experi-

mental approach closely mimics the clinical scenario in when compared to the ratio of myocardial fibrosis in

settings for myocardial infarction but slightly over thethat the transannular patch implantation with infundibu-

lectomy was the reason for the volume overloading in normal ratio of fibrotic tissue in healthy myocardium (1).

The route for delivery of UCMNCs seems to be safethe RV. Regional contractile deterioration following the

incision in the RVOT may have partly influenced the and effective at the time of surgery whereas intracoro-

nary delivery has been reported to be unsuitable for theglobal systolic performance that was partly preserved in

the animals of the cell group. Additionally, the reason treatment of myocardial infarction in a swine model (31)

similarly to intravenous delivery of mesenchymal stemfor the alteration in systolic function may be dependent

on cellular changes in calcium homeostasis and contrac- cells in canine model by Freyman and collegues (17).

Moreover, the thin myocardium of the RV free walltile proteins due to chronic volume stress (3). Because

systolic function was not preserved by the increase of must be considered prone to perforation. Therefore, weused a special hand-made double stage needle with anpreload (heterometric autoregulation), the remaining

mechanism for better systolic function in the cell group epicardial stopper to prevent this eventual complication

during the intramyocardial application of UCMNCs. Inmay involve the increase of contractile performance (ho-

meometric autoregulation), which may have been posi- an experimental study by Borenstein et al. myogenic cell

were implanted into the RV myocardium in a setting oftively influenced by the stem cells. On the other hand,

the latter has been reported to be completely absent in pulmonary artery banding and two animals were lost as

a result of the several injections that presumably lead tochronically volume-overloaded hearts without additional

treatment such as stem cells (10). RV edema and failure (9). We did not observe that kind

of complication. Not only the number of injections andIt is known that diastolic dysfunction occurs in

stressed human hearts (10,39). The difference in dia- the amount of injected volume, but also the appropriate

instruments used for injection should be carefully deter-stolic properties between the groups under dobutamine

stress conditions was more exaggerated. Because the mined.

Undoubtedly, there are limitations to our study. Firstly,slope and position of EDPVR represent relatively inde-pendent parameters for the comparison of elasticity of the conductance catheter method for assessment of the

RV function has also been questioned in the literature.the RV, the rightward downward shift of EDPVR slope

and reduced intracavitary pressure load under pharmaco- Nevertheless, it is generally accepted as one of the most

accurate methods for differentiated functional analysislogically induced stress mirrored the enhanced elasticity

of the RV that received UCMNC treatment. Further, dP/ (15,22,28,41). Alternatively, magnetic resonance imaging

would have provided a detailed assessment of RV func-dtmin under dobutamine stress conditions represented pre-

served diastolic function in the cell group compared to tion whereas the requirement of sedation and breath-

holds for qualitative imaging would have exposed ani-the controls. The restoration of RV volume and pressure

loadings under baseline conditions at 3 months in com- mals with RV compromise to a higher risk during the

investigation and eventually impaired our outcome byparison to the immediate postoperative period in both

groups is, however, an interesting feature in our hemo- causing a higher rate of mortality. Secondly, the effects


13/14


elli, L. Human cord blood CD133+ cells immunoselectedof the implanted cells may originate from neovessel for-by a clinical-grade apparatus differentiate in vitro into en-mation in the transplanted heart, although the definitivedothelial- and cardiomyocyte-like cells. Transfusion 47:

evidence is missing if the transplanted autologous cells 280289; 2007.remained in the myocardium after 3 months, even though 9. Borenstein, N.; Jian, Z.; Fromont, G.; Bruneval, P.; Hekmati,

M.; Behr, L.; Laborde, F.; Montarras, D.; Le Bret, E. Non-there is no concern for immunorejection in our autolo-cultured cell transplantation in an ovine model of rightgous setting of UCMNC transplantation. Nevertheless,ventricular preparation. J. Thorac. Cardiovasc. Surg. 129:cell tracking was not applied in our study. Whether the11191127; 2005.

enhanced angiogenesis is a direct effect of the cell im- 10. Brixius, K.; Reuter, H.; Bloch, W.; Schwinger, R. H. Al-plantation or is derived partly by circulating or resident tered hetero- and homeometric autoregulation in the termi-

nally failing human heart. Eur. J. Heart Fail. 7:2935;cardiac stem cells remains to be elucidated.2005.In conclusion, the results of our study have proven a

11. Burkhoff, D.; Sugiura, S.; Yue, D. T.; Sagawa, K. Con-novel and unique experimental model mimicking post-tractility-dependent curvilinearity of end-systolic pressure-

surgical scenario of TOF patients in large animals. Au- volume relations. Am. J. Physiol. 252:12181227; 1987.tologous, intramyocardial UCMNC transplantation has 12. Carrel, A. Experimental operations on the orifices of the

heart. Ann. Surg. 60:16; 1914.been found feasible and safe and seems to positively influ-13. Chaturvedi, R. R.; Shore, D. F.; Lincoln, C.; Mumby, S.;ence the diastolic properties of the RV under chronic vol-

Kemp, M.; Brierly, J.; Petros, A.; Gutteridge, J. M.;ume overload probably through enhanced angiogenesis.Hooper, F; Redington, A. N. Acute right ventricular re-

However, the strategy of cardiac regeneration with strictive physiology after repair of tetralogy of Fallot: As-UCMNCs in TOF still requires further detailed investi- sociation with myocardial injury and oxidative stress. Cir-culation 100:15401547; 1999.gation before possible introduction into a clinical set-

14. de Vroomen, M.; Cardozo, R. H.; Steendijk, P.; van Bel,ting.F.; Baan, J. Improved contractile performance of right

ACKNOWLEDGMENTS: We are grateful to Prof. Matthias ventricle in response to increased RV afterload in new-Peuster from the Department of Pediatric Cardiology of the born lamb. Am. J. Physiol. Heart Circ. Physiol. 278:100University of Rostock for his helpful discussion. We thank Ms. 105; 2000.

Margit Fritsche and Mr. Reinhard Schwarmer for their excel- 15. Dickstein, M. L.; Yano, O.; Spotnitz, H. M.; Burkhoff, D.lent technical assistance. Assessment of right ventricular contractile state with the

conductance catheter technique in the pig. Cardiovasc.REFEENCES

Res. 29:820826; 199516. dUdekem d Acoz, Y.; Pasquet, A.; Lebreux, L.; Ovaert,1. Agata, J.; Chao, L.; Chao, J. Kallikrein gene delivery im-

proves cardiac reserve and attenuates remodeling after C.; Mascart, F.; Robert, A.; Rubay, J. E. Does right ven-tricular outflow tract damage play a role in the genesismyocardial infarction. Hypertension 40:653659; 2002.

2. Alexiou, C.; Mahmoud, H.; Al-Khaddour, A.; Gnanapra- of late right ventricular dilatation after tetralogy of Fallotrepair? Ann. Thorac. Surg. 76:555561; 2003.gasam, J.; Salmon, A. P.; Keeton, B. R.; Monro, J. L.Outcome after repair of tetralogy of Fallot in the first year 17. Freyman, T.; Polin, G.; Osman, H.; Crary, J.; Lu, M.;

Cheng, L.; Palasis, M.; Wilensky, R. L. A quantitative,of life. Ann. Thorac. Surg. 71:494500; 2001.3. Alvarez, B. V.; Perez, N. G.; Ennis, I. L.; Camilion de randomized study evaluating three methods of mesenchy-

mal stem cell delivery following myocardial infarction.Hurtado, M. C.; Cingolani, H. E. Mechanisms underlyingthe increase in force and Ca(2+) transient that follow Eur. Heart J. 27:11141122; 2006.

18. Gregg, D.; Foster, E. Pulmonary insufficiency is the nexusstretch of cardiac muscle: A possible explanation of theAnrep effect. Circ. Res. 85:716722; 1999. of late complications in tetralogy of Fallot. Curr. Cardiol.

Rep. 9:315322; 2007.4. Ammash, N. M.; Dearani, J. A.; Burkhart, H. M.; Connolly,H. M. Pulmonary regurgitation after tetralogy of Fallot re- 19. Harris, D. T.; Rogers, I. Umbilical cord blood: A unique

source of pluripotent stem cells for regenerative medicine.pair: Clinical features, sequelae, and timing of pulmonaryvalve replacement. Congenit. Heart Dis. 2:386403; 2007. Curr. Stem Cell Res. Ther. 2:301309; 2007.

20. Henning, R. J.; Abu-Ali, H.; Balis, J. U.; Morgan, M. B.;5. Anversa, P.; Kajstura, J. Ventricular myocytes are not ter-minally differentiated in the adult mammalian heart. Circ. Willing, A. E.; Sanberg, P. R. Human umbilical cord

blood mononuclear cells for the treatment of acute myo-Res. 83:114; 1998.6. Assmus, B.; Honold, J.; Schachinger, V.; Britten, M. B.; cardial infarction. Cell Transplant. 13:729739; 2004.

21. Hirata, Y.; Sata, M.; Motomura, N.; Takanashi, M.;Fischer-Rasokat, U.; Lehmann, R.; Teupe, C.; Pistorius,K.; Martin, H.; Abolmaali, N. D.; Tonn, T.; Dimmeler, S.; Suematsu, Y.; Ono, M.; Takamoto, S. Human umbilical

cord blood cells improve cardiac function after myocardialZeiher, A. M. Transcoronary transplantation of progenitorcells after myocardial infarction. N. Engl. J. Med. 355: infarction. Biochem. Biophys. Res. Commun. 327:609

614; 2005.12221232; 2006.7. Beltrami, A. P.; Barlucchi, L.; Torella, D.; Baker, M.; 22. Hon, J. K.; Steendijk, P.; Petrou, M.; Wong, K.; Yacoub,

M. H. Influence of clenbuterol treatment during six weeksLimana, F.; Chimenti, S.; Kasahara, H.; Rota, M.; Musso,E.; Urbanek, K.; Leri, A.; Kajstura, J.; Nadal-Ginard, B.; of chronic right ventricular pressure overload as studied

with pressure-volume analysis. J. Thorac. Cardiovasc.Anversa, P. Adult cardiac stem cells are multipotent andsupport myocardial regeneration. Cell 114:763776; Surg. 122:767774; 2001.

23. Hu, C. H.; Wu, G. F.; Wang, X. Q.; Yang, Y. H.; Du,2003.8. Bonanno, G.; Mariotti, A.; Procoli, A.; Corallo, M.; Z. M.; He, X. H.; Xiang, P. Transplanted human umbilical

cord blood mononuclear cells improve left ventricularRutella, S.; Pessina, G.; Scambia, G.; Mancuso, S.; Pier-


14/14


function through angiogenesis in myocardial infarction. Sousa, A. L.; Mesquita, C. T.; Rossi, M. I.; Carvalho,A. C.; Dutra, H. S.; Dohmann, H. J.; Silva, G. V.; Belem,Chin. Med. J. 119:14991506; 2006

24. Jager, M.; Bachmann, R.; Scharfstadt, A.; Krauspe, R. L.; Vivacqua, R.; Rangel, F. O.; Esparcatte, R.; Geng,Y. J.; Vaughn, W. K.; Assad, J. A.; Mesquita, E. T.; Will-Ovine cord blood accommodates multipotent mesenchy-

mal progenitor cells. In Vivo 20:205214; 2006. erson, J. T. Transendocardial, autologous bone marrowcell transplantation for severe, chronic ischemic heart fail-25. Kim, B. O.; Tian, H.; Prasongsukarn, K.; Wu, J.; Angoul-

vant, D.; Wnendt, S.; Muhs, A.; Spitkovsky, D.; Li, R. K. ure. Circulation 107:22942302; 2003.36. Prat-Vidal, C.; Roura, S.; Farre, J.; Galvez, C.; Llach, A.;Cell transplantation improves ventricular function after a

myocardial infarction: A preclinical study of human un- Molina, C. E.; Hove-Madsen, L.; Garcia, J.; Cinca, J.;Bayes-Genis, A. Umbilical cord blood-derived stem cellsrestricted somatic stem cells in a porcine model. Circula-

tion 112:I96104; 2005. spontaneously express cardiomyogenic traits. Transplant.Proc. 39:24342437; 2007.26. Kogler, G.; Sensken, S.; Airey, J. A.; Trapp, T.; Muschen,

M.; Feldhahn, N.; Liedtke, S.; Sorg, R. V.; Fischer, J.; 37. Shah, A. S.; Atkins, B. Z.; Hata, J. A.; Tai, O.; Kypson,A. P.; Lilly, R. E.; Koch, W. J.; Glower, D. D. Early ef-Rosenbaum, C.; Greschat, S.; Knipper, A.; Bender, J.;

Degistirici, O.; Gao, J.; Caplan, A. I.; Colletti, E. J.; fects of right ventricular volume overload on ventricularperformance and beta-adrenergic signaling. J. Thorac.Almeida-Porada, G.; Muller, H. W.; Zanjani, E.; Wernet,

P. A new human somatic stem cell from placental cord Cardiovasc. Surg. 120:342349; 2000.38. Shpall, E. J.; Quinones, R.; Giller, R.; Zeng, C.; Baron,blood with intrinsic pluripotent differentiation potential. J.

Exp. Med. 200:123 135; 2004. A. E.; Jones, R. B.; Bearman, S. I.; Nieto, Y.; Freed, B.;Madinger, N.; Hogan, C. J.; Slat-Vasquez, V.; Russell, P.;27. Lange, P. E.; Onnasch, D. G.; Bernhard, A.; Heintzen,

P. H. Left and right ventricular adaptation to right ventric- Blunk, B.; Schissel, D.; Hild, E.; Malcolm, J.; Ward, W.;

McNiece, I. K. Transplantation of ex vivo expanded cordular overload before and after surgical repair of tetralogyof Fallot. Am. J. Cardiol. 50:786794; 1982 blood. Biol. Blood Marrow Transplant. 8:368376; 2002.

39. Sibbald, W. J.; Driedger, A. A. Right ventricular function28. Leeuwenburgh, B. P.; Helbing, W. A.; Steendijk, P.;Schoof, P. H.; Baan, J. Biventricular systolic function in in acute disease states: Pathophysiologic considerations.

Crit. Care Med. 11:339345; 1983.young lambs subject to chronic systemic right ventricularpressure overload. Am. J. Physiol. Heart Circ. Physiol. 40. Song, S.; Sanchez-Ramos, J. Preparation of neural progen-

itors from bone marrow and umbilical cord blood. Meth-281:H26972704; 2001.29. Lewis, I. D.; Verfaillie, C. M. Multi-lineage expansion po- ods Mol. Biol. 438:123134; 2008.

41. Stamato, T. M.; Szwarc, R. S.; Benson, L. N. Measure-tential of primitive hematopoietic progenitors: Superiorityof umbilical cord blood compared to mobilized peripheral ment of right ventricular volume by conductance catheter

in closed-chest pigs. Am. J. Physiol. 269:H869876;blood. Exp. Hematol. 28:10871095; 2000.30. Ma, N.; Stamm, C.; Kaminski, A.; Li, W.; Kleine, H. D.; 1995.

42. Stamm, C.; Westphal, B.; Kleine, H. D.; Petzsch, M.;Muller-Hilke, B.; Zhang, L.; Ladilov, Y.; Egger, D.; Stein-hoff, G. Human cord blood cells induce angiogenesis fol- Kittner, C.; Schumichen, C.; Nienaber, C. A.; Freund, M.;

Steinhoff, G. Autologous bone marrow stem cell trans-lowing myocardial infarction in NOD/scid-mice. Cardio-

vasc. Res. 66:45 54; 2005. plantation for myocardial regeneration after myocardialinfarction. Lancet 361:4546; 2003.31. Moelker, A. D.; Baks, T.; Wever, K. M.; Spitskovsky, D.;Wielopolski, P. A.; van Beusekom, H. M.; van Geuns, 43. Steinhoff, G. The regenerating hearthope for children

with congenital heart defects. Kinderkrankenschwester 25:R. J.; Wnendt, S.; Duncker, D. J.; van der Giessen, W. J.Intracoronary delivery of umbilical cord blood derived un- 4750; 2006.

44. Szabo, G.; Soos, P.; Bahrle, S.; Radovits, T.; Weigang,restricted somatic stem cells is not suitable to improve LVfunction after myocardial infarction in swine. J. Mol. Cell. E.; Kekesi, V.; Merkely, B.; Hagl, S. Adaptation of the

right ventricle to an increased afterload in the chronicallyCardiol. 42:735745; 2007.32. Nieda, M.; Nicol, A.; Denning-Kendall, P.; Sweetenham, volume overloaded heart. Ann. Thorac. Surg. 82:989995;

2006.J.; Bradley, B.; Hows, J. Endothelial cell precursors arenormal components of human umbilical cord blood. Br. J. 45. Tse, W. W.; Zang, S. L.; Bunting, K. D.; Laughlin, M. J.

Umbilical cord blood transplantation in adult myeloid leu-Haematol. 98:775777; 1997.33. Nishiyama, N.; Miyoshi, S.; Hida, N.; Uyama, T.; Oka- kemia. Bone Marrow Transplant. 41:465472; 2008.

46. Wairiuko, G. M.; Crisostomo, P. R.; Wang, M.; Morrell,moto, K.; Ikegami, Y.; Miyado, K.; Segawa, K.; Terai,M.; Sakamoto, M.; Ogawa, S.; Umezawa, A. The signifi- E. D.; Meldrum, K. K.; Lillemoe, K. D.; Meldrum, D. R.

Stem cells improve right ventricular functional recoverycant cardiomyogenic potential of human umbilical cordblood-derived mesenchymal stem cells in vitro. Stem after acute pressure overload and ischemia reperfusion in-

jury. J. Surg. Res. 141:241246; 2007.Cells 25:20172024; 2007.34. Orlic, D.; Kajstura, J.; Chimenti, S.; Jakoniuk, I.; Ander- 47. Zahka, K. G.; Horneffer, P. J.; Rowe, S. A.; Neill, C. A.;

Manolio, T. A.; Kidd, L.; Gardner, T. J. Long-term valvu-son, S. M.; Li, B.; Pickel, J.; McKay, R.; Nadal-Ginard,B.; Bodine, D. M.; Leri, A.; Anversa, P. Bone marrow lar function after total repair of tetralogy of Fallot. Rela-

tion to ventricular arrhythmias. Circulation 78:III1419;cells regenerate infarcted myocardium. Nature 410:701705; 2001. 1988.

35. Perin, E. C.; Dohmann, H. F.; Borojevic, R.; Silva, S. A.;

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