www.thelancet.com Published online July 26, 2012 http://dx.doi.org/10.1016/S0140-6736(12)60737-5 1
July 26, 2012
See Online for a video interview
with Martin Birchall
Department of Cardiothoracic
Surgery (Prof M J Elliott MD,
S Speggiorin MD, A Fierens RNC,
D Vondrys MD), Department of
Surgery (P De Coppi MD),
Department of Radiology
(D Roebuck MB, C McLaren DCR),
and Ear Nose and Throat
Surgery (L Cochrane MD,
C Jephson FRCS), Great Ormond
Street, Hospital for Children,
London, UK; Centre for
(C R Butler MRCS, S Janes MD),
Centre for Nanotechnology
and Regenerative Medicine
(C Crowley MSc,
Prof A M Seifalian PhD), and
Centre for Molecular Cell
Biology (N J Beaumont PhD,
Prof J J Hsuan PhD), University
College London, London, UK;
Paul O’Gorman Laboratory of
Department of Haematology,
Royal Free Hospital, London,
UK (E Samuel MSc,
M W Lowdell PhD); School of
Veterinary Sciences, University
of Bristol, Bristol, UK
(T Cogan PhD); Department of
Cell Techniques and Applied
Stem Cell Biology, University of
Leipzig, Leipzig, Germany
(Prof A Bader MD); and
University College London Ear
Institute, The Royal National
Throat, Nose and Ear Hospital,
(Prof M A Birchall MD)
Prof Martin A Birchall, University
College London Ear Institute,
Royal National Throat Nose and
Ear Hospital, 330332, Gray’s Inn
Road, London WC1X 8EE, UK
Stem-cell-based, tissue engineered tracheal replacement in
a child: a 2-year follow-up study
Martin J Elliott, Paolo De Coppi, Simone Speggiorin, Derek Roebuck, Colin R Butler, Edward Samuel, Claire Crowley, Clare McLaren, Anja Fierens,
David Vondrys, Lesley Cochrane, Christopher Jephson, Samuel Janes, Nicholas J Beaumont, Tristan Cogan, Augustinus Bader,
Alexander M Seifalian, J Justin Hsuan, Mark W Lowdell, Martin A Birchall
Background Stem-cell-based, tissue engineered transplants might off er new therapeutic options for patients, including
children, with failing organs. The reported replacement of an adult airway using stem cells on a biological scaff old
with good results at 6 months supports this view. We describe the case of a child who received a stem-cell-based
tracheal replacement and report fi ndings after 2 years of follow-up.
Methods A 12-year-old boy was born with long-segment congenital tracheal stenosis and pulmonary sling. His airway
had been maintained by metal stents, but, after failure, a cadaveric donor tracheal scaff old was decellularised. After a
short course of granulocyte colony stimulating factor, bone marrow mesenchymal stem cells were retrieved
preoperatively and seeded onto the scaff old, with patches of autologous epithelium. Topical human recombinant
erythropoietin was applied to encourage angiogenesis, and transforming growth factor β to support chondrogenesis.
Intravenous human recombinant erythropoietin was continued postoperatively. Outcomes were survival, morbidity,
endoscopic appearance, cytology and proteomics of brushings, and peripheral blood counts.
Findings The graft revascularised within 1 week after surgery. A strong neutrophil response was noted locally for the
fi rst 8 weeks after surgery, which generated luminal DNA neutrophil extracellular traps. Cytological evidence of
restoration of the epithelium was not evident until 1 year. The graft did not have biomechanical strength focally
until 18 months, but the patient has not needed any medical intervention since then. 18 months after surgery, he
had a normal chest CT scan and ventilation-perfusion scan and had grown 11 cm in height since the operation. At
2 years follow-up, he had a functional airway and had returned to school.
Interpretation Follow-up of the fi rst paediatric, stem-cell-based, tissue-engineered transplant shows potential for this
technology but also highlights the need for further research.
Funding Great Ormond Street Hospital NHS Trust, The Royal Free Hampstead NHS Trust, University College
Hospital NHS Foundation Trust, and Region of Tuscany.
There are no universally eff ective solutions for the
treatment of advanced structural disorders of the large
airways in children. Such children need frequent stays in
hospital. Although slide tracheoplasty is the primary
treatment of choice for some children, others develop
recurrent stenoses.1 Stent erosion and death can occur.2
Fetuses with laryngotracheal agenesis or severe stenosis
identifi ed before birth might be aborted because these
abnormalities are regarded as fatal.3
In the past decade, tissue-engineered structures re-
populated with cells or stem cells have been used clinically.
Atala and colleagues4 used collagen scaff olds reseeded
with urothelial and muscle cells to repair bladder defects
in patients with myelo meningocele, and the successful
clinical application of a stem-cell-based tracheal replace-
ment in a woman with end-stage airway disease with
6 months follow-up has been reported.5 In 2011, good
short-term (5 months) outcomes were reported in a
patient who received a similar stem-cell-based tracheal
graft, but in this study a nanocomposite trachea was used
as the scaff old.6 However, the long-term outcomes of these
and other patients who have received such grafts on
compassionate grounds have yet to be published pending
adequate follow-up, and there is no previous report of
outcomes after a tracheal graft in a child.
The potential clinical advantages of autologous stem-
cell-derived transplants are that patients who receive
them would not need immunosuppression and that the
transplants are hypothesised to be remodelled by local
stroma to simulate native tissue.5 By contrast, allo-
trans plantation is associated with signifi cant long-term
mortality due to infection and immunosuppression,
especially in the respiratory system.7 There is also a
paucity of donors for transplantation. Thus, there is a
signifi cant unmet need for novel methods of replacing
and regenerating human tissue.
The ideal endpoints for tracheal replacement in children
are normal airway and lung function, appropriate growth,
high quality of life, and the elimination of the need for
repeated surgical inter ventions. Here, we describe the
case of a child who received a stem-cell-based tracheal
replacement as an urgent compassionate-use procedure
and report fi ndings after 2 years of follow-up.
www.thelancet.com Published online July 26, 2012 http://dx.doi.org/10.1016/S0140-6736(12)60737-5
A child born with long-segment congenital tracheal
stenosis and pulmonary sling underwent autologous patch
tracheoplasty at Great Ormond Street Hospital NHS Trust
(London, UK) at 6 days old. He could not be extubated after
surgery because of collapse and scarring of the patch and
severe bilateral bronchomalacia. Balloon-expandable
stainless steel stents (Palmaz, Cordis, Miami Lakes, FL,
USA) were implanted, with marked clinical improvement.
At 3 years old, the child had substantial bleeding into his
airway. Emergency bronchoscopy and CT angi o graphy
revealed erosion by the stent into the aorta. Emergency
aortic repair was done with a bovine pericardial patch
(Synovis, St Paul, MN, USA), and the impacted stents and
trachea were excised and replaced by a tracheal homograft.8
The homograft was replaced 1 week later by another
(stented) homograft after mediastinitis occurred. After
3 months in hospital, the patient made an excellent re-
covery. Over the ensuing years, he needed occasional inter-
ventions, including further stents for recurrent stenosis.
At 10 years old, the patient suff ered a second
haemorrhage. Findings from bronchoscopy and CT scan
suggested erosion of tracheal stents, creating a new
aortotracheal fi stula. Bleeding stopped spontaneously,
which provided us with time to plan urgent reconstruction.
Use of tracheal homografts had been discontinued
and so other options were discussed. Tracheal allo grafting9
was dismissed due to the prospect of lifelong immuno-
suppression. In view of our previous success with an
autologous stem-cell-based tracheal replacement, the
child’s parents were approached and asked to consider the
use of a similar method for their child. The emergent
nature of his disorder, unlike that of the adult recipient,
meant that a more direct protocol for graft preparation was
needed, and so the published technique was adapted using
methods previously applied success fully to bone, skin, and
nerve regeneration.10 After approval by the Medicines and
Healthcare Products Regulatory Agency (MHRA) and the
institutional Clinical Ethics Committee, his parents
consented to the procedure. An appropriate scaff old was
sought and the patient was prepared for surgery.
The patient received 10 mg/kg granulocyte colony
stimulating factor (G-CSF; Chugai, London, UK) daily
for 3 days before surgery to mobilise haemopoietic stem
cells and endothelial progenitors11 and induce mesen-
chymal stem cell (MSC) proliferation.12 We measured
leucocyte counts daily.
After general anaesthesia, 50 mL autologous bone
marrow was aspirated into sterile, heparinised tubes,
diluted 1:1, and mononuclear cells were isolated by
discontinuous density gradient separation. Sterility test-
ing was done on the washings. Cells were re-suspended,
topped up, and counted. The total mononuclear cell count
was 2·56×10⁸, of which 5·51×10⁶ were CD34+ or CD45wk
haematopoietic stem cells and 1·68×10⁶ were CD73+,
CD90+, CD105+, CD117+, or CD45+ MSCs. The cell
suspension was transferred into a 60 mL Cryocyte bag
(Miltenyi, Bisley, UK) supplemented with 10 IU protamin
(Wockhardt, Wrexham, UK) and shipped in a tem-
perature-monitored container (4°C) to the operating room
at Great Ormond Street Hospital NHS Trust.
A CT scan was done to identify the dimensions needed
for the scaff old. An allogeneic trachea of appropriate size
was retrieved by permission of the Tuscany regional
authorities from a 30-year-old female donor. Infectious
disease markers were negative. The scaff old was pre pared
in the Regenerative Surgery Laboratories of the University
Hospital Careggi, Florence, Italy, using a published
protocol,13 and was released by quality control after con-
fi rmation of sterility and absence of HLA-I+ cells. 3 days
before surgery, the scaff old was imported (4°C) in
phosphate-buff ered saline supplemented with penicillin,
streptomycin, and amphotericin B, in compliance with
UK Human Tissue Authority codes and local licence
number 11016. 10 000 IU erythropoietin (Roche, Welwyn,
UK), 200 IU G-CSF (Neupogen, Cambridge, UK) and
50 μg transforming growth factor β (TGFβ; R&D Systems,
Abingdon, UK) were contained in separate syringes and
transported with the scaff old and cells.
Tracheal replacement surgery
During surgery, the head-down tilt position, cardio-
pulmonary bypass, and progressive cooling to 18°C were
used. With the heart decompressed, a resternotomy was
done. Use of right atrium and superior vena cava venous
lines permitted cardiac isolation and great vessels
were mobilised. At 18°C, the aorta was cross-clamped,
anterograde cardioplegia was instilled, head vessels were
snared, and the circulation was stopped. After dissection,
the stent that was entering the aorta was identifi ed, as
were others buried in the tracheal wall (fi gure 1A). The
aortic defect was repaired with bovine pericardium.
Circulation was resumed and re-warming commenced.
The trachea was transected above the upper metal stent
and below the lower metal stent, leaving a 7 cm gap be-
tween the upper trachea and the carina (fi gure 1B). Patches
of tracheal epithelium were removed from the excised
trachea, cut into stamp grafts and retained. The stents in
the main bronchi were trimmed back to provide cuff s of
unstented bronchi for anastomosis. Bronchi were dilated
with an 8 mm Hegar dilator (Lyall Willis, Hastings, UK).
In the operating room, the scaff old was saturated with
the cell suspension. The mucosal stamps were placed as
free grafts at regular intervals within the lumen. An
absorbable polydioxanone (PDO) tracheal stent (Ella-Cs,
Hradec Kralove, Czech Republic) measuring 12×72 mm
was sutured in place (5/0 PDS, II, Ethicon, Edinburgh,
UK). The construct was saturated with human recom-
binant erythropoietin (hrEPO) and G-CSF, and TGFβ was
injected into the tracheal rings (fi gure 1D) to increase
angiogenesis, improve autologous MSC recruit ment, and
www.thelancet.com Published online July 26, 2012 http://dx.doi.org/10.1016/S0140-6736(12)60737-5 3
induce chondrocyte diff erentiation. The construct was
anasto mosed superiorly and inferiorly using horizontal
mattress interrupted sutures (4-0 PDS II, fi gure 1E). Before
completing the anastomoses, a new trans-nasal endo-
tracheal tube was placed under direct vision. Two small air
leaks were sealed and the child was weaned from
cardiopulmonary bypass. The omentum was mobilised
and interposed between the trachea and heart to reduce
the possibility of future fi stulae and increase graft
vascularity (fi gure 1F). On alternate days after surgery,
hrEPO (10 000 IU for 2 weeks) and G-CSF (10 mg/kg for
1 week) were administered.
Role of the funding source
The sponsors of the study had no role in the study design,
data collection, data analysis, data interpretation, or
writing of the report. The corresponding author had full
access to all the data in the study and had fi nal
responsibility for the decision to submit for publication.
Within 3 h of the surgery, ventilation became problematic
and bilateral air trapping
bronchoscopy showed substantial narrowing of the origin
of both bronchi due to the longitudinal rigidity of the
absorbable stent. A temporary stent (Niti-S, Taewoong,
Seoul, South Korea) was implanted in each bronchus
under fl uoroscopy, which resulted in an immediate
improvement in ventilation. These stents were removed
before extubation on day 26 after surgery. There was an
initial increase in the number of circulating leucocytes
(31·1×10⁹/L [SD 6·6×10⁹/L]) between days 2 and 8 after
surgery, which corresponded to the period of application
of G-CSF and hrEPO. This period was also the only time
when circulating CD34+ cells (0·71×10⁹/L [SD 0·05×10⁹/L])
could be detected. Leucocyte counts normalised from
day 9 (9·14 ×10⁹/L [SD 1·18×10⁹/L]; appendix).
There was bleeding on contact from the internal lumen
of the graft by 1 week after surgery, which proved that
angiogenesis was occurring. The patient needed regular
bronchoscopy for removal of dense secretions for 8 weeks
(fi gure 2A). Assessment of the secretions showed that they
included no cells, had a high DNA content, and had a net-
like microscopic appearance. The features were identifi ed
as those of DNA neutrophil extracellular traps (NETs).14,15
SDS gel frac tionation, tryptic digestion, and nano-liquid
chroma tography mass spectrometry identifi ed a protein
profi le consistent with this diagnosis (fi gure 3).14 The
secretions were treated with a combination of DNase and
physiotherapy, and cleared as epithelialisation pro gressed.
The patient was discharged on day 63.
6 weeks after surgery the stent had dissolved and there
was mild collapse of the proximal graft. A shorter
(10×45 mm) PDO stent was implanted under fl uoroscopy.
The patient underwent bronchoscopy or balloon dilatation
under fl uoroscopy, or both, regularly for 6 months (fi gure
2B). The major reason for further bronchoscopy or balloon
dilation was mucus retention and crusting within the
native bronchi in which there were still embedded metal
stents. At 5 months, after dissolution of the latest stent, we
remained concerned about the rigidity of the proximal
graft, and so overlapping, self-expanding Nitinol stents
(S.M.A.R.T. Control, Cordis, Waterloo, Belgium) were
implanted into the trachea. At 6 months after the initial
surgery the graft seemed stable, the patient’s airway was
patent, and he returned to school.
The patient’s last endoscopy (15 months after surgery)
showed complete epithelialisation (fi gure 2C), and
Figure 1: Surgical procedure
(A) During surgery the airway was found to be severely stenotic with multiple stents including one entering the
ascending aorta. (B and C) The old homograft trachea was removed and replaced by the engineered graft.
(C) The aortic defect was closed with a bovine pericardial patch and air leaks sealed. (D) Transforming growth
factor β was injected into tracheal rings in the operating theatre before (E) implantation of the recellularised
graft. (F) Before closing, an omental wrap was brought up to cover the graft. The graft sits in the anatomical
position to the right of the ascending aorta.
Figure 2: Bronchoscopic appearances
(A) Microlaryngobronchoscopy 15 days after the transplant showing a dense web covering the stent and partially
occluding the lumen (A), which was cleared by regular bronchoscopies and DNAase. (B) Image at 6 months,
showing that reabsorption of the stent (white areas) caused so-called cobblestones of granulation tissue with little
normal epithelium. (C) At 15 months after surgery, the graft seemed to be patent, with healthy mucosa.
See Online for appendix
www.thelancet.com Published online July 26, 2012 http://dx.doi.org/10.1016/S0140-6736(12)60737-5
cytology of tracheal brushings showed healthy, ciliated
respiratory epithelial cells (fi gure 4D). At 18 months, he
had his last fl uoroscopic balloon dilatation because the
malacic seg ment had strengthened such that he had not
needed any further admissions to hospital. As of May 7,
2012, he was well, active, and had grown 11 cm and 5 kg
since graft im plantation. His lungs appeared normal on
CT scan, without bronchiectasis or air trapping, and a
ventilation-perfusion scan at 12 months was normal
(fi gure 5). As of May 13, 2012, there has been no
serological or clinical evidence of rejection of the graft
and a comprehensive screen of his serum at 15 months
showed no anti-HLA antibodies.
Histological assessment of the homograft trachea
removed at the time of surgery showed an infl amed
mucosa overlying dense fi brous tissue and islands of
cartilage unlike normal tracheal architecture (fi gure 4B).
Histology of the decellularised scaff old showed complete
decellularisation with good retention of tracheal archi-
tecture (fi gure 4C) and absence of MHC expression (not
shown). High-resolution proteomic analysis of the scaff old
by ion-trap mass spectrometry identifi ed 166 proteins,
including several extracellular matrix components.
Bioinformatic analysis (IPA, Ingenuity Systems, Redwood
City, CA, USA) identifi ed a broad range of potential
biological roles for these proteins.
We report a stem-cell-based tissue replacement in a
child and long-term follow-up of a stem-cell-based
tissue-engineered graft (panel). The child is well,
growing, and had not needed medical intervention for
6 months by May 5, 2012.
Because the protocol used in this study was devised in an
emergency, we applied empirically a new combination of
technologies on the basis of previous clinical successes in
non-airway settings (ie, bone, skin, and nerve regeneration).
Thus, to minimise delays, there was no previous expansion
of epithelial cells and MSCs, nor any chondrocytic
DescriptionMw (Da) PLGS scorePeptides Coverage (%)
Alpha 2 macroglobulin
Ig alpha 1 chain C region
Ig alpha 2 chain C region
Ig gamma 2 chain C region
Ig kappa chain C region
Ig kappa chain V III region SIE
Ig kappa chain V III region WOL
Ig mu chain C region
Polymeric immunoglobulin receptor
Figure 3: Identifi cation of protein in the tracheal exudate
Proteins in the tracheal exudate identifi ed in the early weeks (sampled postoperative week 2) were separated using SDS-PAGE and stained with colloidal Coomassie
Blue (A). Destained gel slices were digested with trypsin (Promega, Southampton, UK), fractionated by high-performance liquid chromatography (NanoAcquity,
Waters, Manchester, UK), and analysed using an in-line Q-TOF mass spectrometer (Waters). (B) The table shows the proteins identifi ed from at least two peptides and
with a PLGS score greater than 10. PLGS=Protein Lynx Global Server.
100 μm20 μm
Figure 4: Findings on cytology
Haematoxylin and eosin staining of (A) normal trachea compared with (B) the patient’s previous tracheal homograft
removed at the time of surgery, which shows an epithelialised lining but atypical gland formation. (C) A sample of the
decellularised tracheal graft used in this study shows loss of cells but preservation of normal architecture. (D) Bronchial
brushing taken from the middle of the graft 1 year after surgery shows a cluster of ciliated cells.
www.thelancet.com Published online July 26, 2012 http://dx.doi.org/10.1016/S0140-6736(12)60737-5 5
diff erentiation of MSCs.5 Instead, we used an intraoperative
protocol, which was similar to those used in clinical trials
of MSCs for patients with myocardial infarction.17 Not
undertaking long-term culture of MSCs also has the
potential advantage of avoiding the risk of malignancy.18
We aimed to create an in-vivo microenvironment that
represented some of the events that occur during the
normal physiological response to injury. A similar method
is in phase 2 clinical trials of bone, skin, and nerve
regeneration.10 We hypothesise that this altered protocol, in
addition to the length of the graft, the presence of an
absorbable stent, and the underlying diff erent physiology
and regenerative potential of children’s compared with
adults’ tissues, were responsible for the diff erences in
clinical course and outcomes from the published adult
case, at least at the 6-month timepoint.5 Specifi cally, the
graft in the present study took longer to epithelialise and
did not have proximal rigidity until almost 2 years.
However, at last follow-up the boy was alive, growing, had
normal lung function, and had returned to school.
A key criterion for paediatric implants is that of
growth potential. In this study, although we were
unable to measure graft length, there was no CT evidence
of shortening of the graft, as has been previously
described, for example, with an alternative aortic allo-
graft approach.19 At age 13 years the child’s torso is not
expected to elongate much further as his height increases
and so the growth demands on this graft are limited.
However, experimental evidence of graft growth is crucial
for the clinical use of similar protocols of transplantation
for children of all ages. Equally crucial is rapid
vascularisation. As with the adult case,5 touch bleeding
on the internal graft surface was visible by 1 week, which
proved that rapid angiogenesis was occurring.
In both this study and a previous case,5 a cadaveric
donor trachea was decellularised, with successful removal
of cellular components including MHCs. Neither patient
had developed rejection by May, 2012, and the child had
not developed anti-donor antibodies by 20 months. These
fi ndings, in addition to reports of preclinical success with
similar methodologies for heart and lung grafts,20,21
suggest that decellularised scaff old-based technologies
could be an immuno suppression-free alternative to
In the UK, patients operated upon under a Hospitals
Exemption Certifi cate on compassionate grounds, as was
the case with the patient in this study, are not treated as
research patients. Thus, we did not label the applied cells
and so cannot comment on whether the eventual stromal
and epithelial cells originated from those implanted or
from cells recruited from neighbouring tissues. Future
preclinical and clinical trials should incorporate markers
that will answer the question of the exact contribution of
applied cells to the fi nal result.
Many clinicians assume that decellularised scaff olds
are inert composites of structural proteins. Proteomic
measurement of non-structural, or minor structural,
proteins has been diffi cult because of the dominance of
collagen and elastin in protein preparations. In this
study, with new techniques we identifi ed 166 proteins
with diverse functions relevant to regenerative medicine
(eg, angiogenesis and immunity) that were preserved
despite decellularisation, although no MHC molecules
were found. We hypothesise that many of these pro teins
are crucial to revascularisation, cell migration, and
diff erentiation in tissue-engineered organs and repre-
sent a major diff erence from synthetic scaff olds.
Therefore, proteomic analysis might be a valuable
addition to release criteria for biological scaff olds.
TGFβ was added to the scaff old to induce chondrocytic
diff erentiation, G-CSF to boost autologous MSC recruit-
ment, and hrEPO to increase angiogenesis. G-CSF is
used to mobilise bone marrow progenitor production
before haemopoietic cell transplantation.22 Although
some studies report a benefi cial eff ect of G-CSF on MSC
mobilisation,12,23 others suggest the opposite eff ect.17
Identifi cation of the contribution of G-CSF to the survival
and function of the graft in one patient in the short and
long term is not possible. However, we hypothesise that
Figure 5: Follow-up scans
(A) CT axial scan and (B) coronal scan done 12 months after surgery show the tracheal graft (arrows) surrounded by
omental fat (*). The lumen of the graft is narrow (6 mm) and its wall is thick (3–4 mm). Growth in length of the
graft was not seen on serial images, possibly because growth in height of the child was not matched by lengthening
of the chest. (C) A lung scan (ventilation-perfusion) at 18 months showed normal bilateral ventilation (the left lung
is contributing 45% to the total ventilation and the right lung 55%). There is a slight reduction in perfusion in the
left lung (receiving 37% of the right heart output) compared with the right lung (63%).
www.thelancet.com Published online July 26, 2012 http://dx.doi.org/10.1016/S0140-6736(12)60737-5
system ic application of G-CSF increased leucocyte
counts in week 1 and contributed to NET accumulation
in the trachea in the fi rst 6 weeks after surgery.23,24
hrEPO is used clinically to support erythropoiesis in
patients with cancer and renal disease.25 Pretreatment
with erythropoietin might improve the survival of cells
within tissue where angiogenesis is not yet adequate to
fully support respiration, by a mechanism mediated by
nitric oxide and vascular endothelial growth factor.26
Angiogenesis, measured by appearance, contact bleed-
ing, and laser doppler fl uxmetry, was equally fast in the
previously reported adult patient5 as in the child in the
present study. Despite the substantial increase in graft
mass in the child, we can only speculate about the added
angiogenic eff ect of hrEPO. Findings are further
confounded by the use of an omental fl ap, because the
purpose of it is to provide an improved vascular bed for
the graft. More research into angiogenic mechanisms in
re cellularised regenerative constructs is needed.
We hypothesised that TGFβ, a key signal for
chondrocytic diff erentiation of MSCs,27 would enable
repopulation of the preserved scaff old cartilage niche and
provide adequate biomech anical support in the long
term. However, TGFβ is also a powerful promoter of
myofi broblasts and scar tissue28 and restricts epithelial
cell survival and migration,29 both of which are undesirable
actions during the regeneration of tissue-engineered
trachea. The absence of rigidity in the proximal trachea
suggests that TGFβ did not support adequate cartilage
regeneration throughout the graft, although the length of
the graft, presence of PDO stents, or the absence of a
preoperative chondrocytic diff eren tiation step in the
process5 might also have been responsible.
PDO stents30,31 have been used in six lung transplant
patients who needed multiple insertions,31 which was
also the case with the child in this study. All six patients
were free of stenosis at a median of 24 months
(range 7–44). Recent experience in children with airway
stenosis is similar.32 The PDO stents were quick to apply
and provided circumferential support for 8 weeks, but
the absence of vertical elasticity was a problem and they
might have contributed to NET formation.
Analysis suggested that the problematic tracheal
exudate in this study was DNA NETs.14,15 The macroscopic
appearance of NETs is poorly described in man. Their
perceived role is to prevent bacterial colonisation and
dissemination, but formation can cause tissue damage.15
We hypothesise that neutrophil recruitment induced by
the graft and stent plus G-CSF treatment were causative,
and that NET resolution parallels the development of new
The epithelium was patchy by 2 months, although stents
caused discontinuity. The presence of viable, ciliated
epithelial cells was confi rmed on cytology at 1 year, when
mucosal continuity was noted throughout. Epithelialisation
occurred later than in the previous adult case,5 where
mucosal coverage was achieved at 1 month and mucociliary
clearance by 6 months.5 The need for early mucosal
coverage and mucociliary clearance for airway grafts in
patients, many of whom have compromised bronchial or
lung function, or both, means that research into
mechanisms of regeneration of the respiratory mucosa is
crucial, as is identifi cation of key stem or progenitor cells
and migration and diff erentiation factors.
This report should be compared with other published
case reports of tissue-engineered airways, described briefl y
earlier5,6,13 and reviewed in more detail elsewhere.3
Substantial areas for improvement in outcomes were
identifi ed by this experience of a stem-cell-based, paediatric
tracheal replacement; specifi cally,
biomechanical strength throughout the graft and speedy,
effi cient restoration of the mucosa. The response of
children to implants will probably diff er from that of adults
in important ways, including the need to accommodate
growth. Urgent research is needed to convert one-off ,
compassionate-use suc cesses, such as the one described in
this study, into more widely applicable clinical treatments
for the thousands of children with tracheal stenosis and
the need for
MJE, SS, and others did the transplant surgery. Postoperative endoscopies
and general medical care were done by MJE, SS, AF, DV, LC, CJ, and
MAB. Radiology, radiologically-guided procedures, and stent placements
were done by DR, CM, DV, and MAB. Good Manufacturing Practice cell
and cytokine preparation was done by ES, CC, and MWL. Analysis of
cytology was done by CB and SJ; histology by CB, CC, TC, and AMS; and
proteomics by CB, NJB, and JJH. Advice on clinical use of cytokines was
provided by AB. MJE, MWL, and MAB designed the protocol. Further data
collection was done by PDC, CB, and DV. MJE, PDC, ES, MWL, and MAB
did the literature search, data interpretation, and writing of the report.
Confl icts of interest
We declare that we have no confl icts of interest.
This work was supported by Great Ormond Street Hospital NHS Trust, The
Royal Free Hampstead NHS Trust, and University College Hospital NHS
Foundation Trust (all London, England), and by a grant
(pd 239- 28/04/2009, delib. GRT 1210/08) from the Region of Tuscany
(Italy) entitled “Clinical laboratory for complex thoracic respiratory and
vascular diseases and alternatives to pulmonary transplantation”. Both
Great Ormond Street Hospital and University College Hospital receive
Panel: Research in context
We searched PubMed for all publications, including clinical trials, meta-analyses, and
reviews, with the terms “graft” and “short-term” or “long-term”. However, we did not
restrict our searches to only papers that included the phrase “stem cells” and identifi ed
only two similar case reports, both in adults and with 6 and 5 months’ follow-up
respectively.5,6 Although several conventional treatments are available for the treatment
of congenital tracheal stenosis, no proven treatments exist for patients with end-stage
disease.1,16 Evidence from studies in animals suggests that stem-cell-based tissue
engineered tracheal implants could be useful as part of new treatment strategies for
incurable tracheal stenosis or malacia in children.
This study describes a stem-cell-based organ transplant in a child and is the fi rst in either
adults or children to report long-term follow-up (2 years).
Articles Download full-text
www.thelancet.com Published online July 26, 2012 http://dx.doi.org/10.1016/S0140-6736(12)60737-5 7
translational research funding from the UK Department of Health’s
National Institute for Health Research Biomedical Research Centres
scheme (MJE, PDC, SJ, and MAB). MJE is Director of the Service for
Severe Tracheal Disease in Children, funded by the National Health Service
National Commissioning Mechanisms. SJ is a recipient of a Wellcome
Trust Senior Fellowship in Clinical Science. Some laboratory work was
supported by a Medical Research Council Translational Stem Cell Research
Committee grant to MB (G1001539) and a Great Ormond Street Hospital
Charity grant to PDC. We thank Caroline Doyle, whose superb
administrative skills were essential to the co-ordination of this procedure.
We also thank the anaesthetic, technical, paramedical, and nursing staff in
operating theatres, intensive care units, and wards at Great Ormond Street
Hospital for Children National Health Service Trust, as well as the senior
management of the Trust who approved the internal funding. We thank
staff at the Royal Victoria Hospital NHS Trust in Belfast who saved the
child’s life at fi rst presentation and cared for him on several occasions over
the past 11 years. All the staff of the Paul O’Gorman Laboratories for Cell
Therapy at the Royal Free Hospital Hampstead NHS Trust contributed to
cell, cytokine, and graft preparation, as did many members of the
Nanotechnology and Surgical Sciences Laboratories at the same institution.
We thank our patient’s parents, who were an essential and supportive part
of the team and decision-making processes. We also thank the Tuscany
Transplant Authority, the Thoracic Surgeons, General and Medical
Directors of the University Hospital Careggi in Florence (Italy), staff at the
UK’s Medicines and Healthcare Products Regulatory Agency (MHRA),
especially Ian Rees, who gave timely and free advice on regulatory aspects
of this case. The Chairman and members of the Clinical Ethics Committee
at Great Ormond Street Hospital constructively considered all aspects to the
case and helped with design of the parent’s information sheet. We fi nally
pay particular tribute to the courageous and inspiring young man himself.
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