Category of paper: Mini Review
Tissue-engineered constructs for urethral regeneration
Kuo-Liang Chen 1, 2, Hsi-Chin Wu 1, 2, Chao-Hsiang Chang 1, 2
1 Department of Urology, China Medical University Hospital, Taichung, 40402,
Taiwan;
2 China Medical University, Taichung, 40402, Taiwan
Running title: Tissue-engineered urethra
Corresponding author: Kuo-Liang Chen
e-mail: CKL_2001@YAHOO.COM
Mailing address: No. 2, Yuh-Der Road, Taichung, Taiwan 40447
Telephone number: 886-4-22052121 # 4440
Fax number: 886-4-22053425
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Abstracts
Those who have urethral injury, long distance of urethral stricture, hypospadias or
epispadias, need tissue for urethral repair. Tissue engineering is one of the solutions
for urethroplasty. Three components essential for tissue engineering are cells,
scaffolds, and bioactive factors. Several animal studies of tissue-engineered urethras
were conducted in the past which have been transferred to human clinical trials by
1999. Of these studies, the remarkable results and concepts were that the maximum
distance for normal tissue regeneration in tubularized urethral replacement with
unseeded matrices is 0.5 cm. Whilst autologous tissue-engineered tabularized
urethras were successful in clinical trial, this method could be a new alternative
treatment in urethral reconstruction.
Key words: Tissue engineering, urethroplasty, reconstructive surgery
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INTRODUCTION:
Those who have urethral injury, long distance of urethral stricture, hypospadias or
epispadias, need tissue for urethral repair. Skin graft, bullock’s urethra, vein, ureter,
appendix vermiformis, fascia of the thigh, bladder mucosa, tunica vaginalis,
peritoneum, rectal mucosa, buccal mucosa or prepuce, have been used for this
purpose.1 However, either the surgeons did not succeed, or the patients must suffer
from donor site surgery, even donor site complications. Tissue engineering is one of
the solutions for urethroplasty.
What are tissue engineering and regenerative medicines? The term “tissue
engineering” was introduced to medicine in 1987.1 S.F. Badylak and R.M. Nerem had
a vivid description of it in 2010 that “tissue engineering” involves the ex vivo creation
of replacement tissues intended for subsequent in vivo implantation.2 They also
indicated that “Regenerative medicine” emphasizes on tissue replacement with ex
vivo manufactured products which have evolved to include broad strategies to
induce both in vivo constructive remodeling of cell-based and cell-free scaffold
materials and true tissue regeneration.2
PRINCIPLES OF TISSUE-ENGINEERING:
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Three components essential for tissue engineering are cells, scaffolds, and bioactive
factors. A usual study design for tissue engineering is to seed cells into scaffolds with
or without bioactive factors which are the constructs for the experimental group. In
the control group, scaffolds without cells are used for comparison. All of them are
implanted to the optimal subjects for being retrieved and evaluated sometime in the
future.3 The cells for tissue engineering might be stem cells (for examples: fertilized
eggs, embryonic stem cells, parthenogenetic stem cells, induced pleuripotent stem
cells, adult stem cells, and so on), progenitor cells (for example: endothelial
progenitor cells, etc.), and differentiated cells (for examples: urothelial cells, smooth
muscle cells, squamous cells, and so on). The scaffolds for tissue engineering might
be classified as natural, synthetic; or absorbable, non-absorbable. Most scaffolds are
absorbable, and few non-absorbable. They can be synthetic polymers or nature ones.
The biomaterials for urethral tissue engineering in the past were all absorbable,
including synthetic polymers [aliphatic polyesters (PGA)] and natural collagens (small
intestine submucosa, bladder-derived acellular submucosa, acellular urethral
submucosa, foreskin acellular matrix, acellular arterial matrix, and so on). The
bioactive factors for tissue engineering might be growth factors, drugs, genes, gene
products, and bioreactors.
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MAXIMUM DISTANCE FOR NORMAL TISSUE REGENERATION:
What is the maximum distance for normal tissue regeneration? To answer this
question, Dorin and his colleagues designed a study using varying lengths (0.5, 1, 2, 3
cm) of tubular scaffolds without cells up to 4 weeks in a rabbit model in vivo.4 They
used acellular bladder submucosas as scaffolds. Follow-up urethrograms
demonstrated normal urethral calibers in the 0.5 cm group at all time points. The
evolution of a stricture was demonstrated in the other longer grafts by 4 weeks.
There were ingrowths of urothelial cells from the anastomotic sites in all grafts at 1
week. They concluded that 0.5 cm appeared to be the maximum defect distance
using acellular grafts that rely on the native cells for tissue regeneration.
ONLAY V.S. TUBULARIZED REPLACEMENT OF URETHRAS:
To regenerate urethra, onlay and tubularized replacement has been used in study
design.(Figure) Onlay replacement needs a healthy urethral bed which provides
healthy cells to migrate into the construct.5 The critical point is the width of the
construct instead of the length. We foresee the success of urethroplasty using
scaffolds without cells while the width of the construct is less than 0.5 cm. 4,6 The
width of the construct can be longer than 0.5 cm if scaffolds with cells are used.
Tubularized replacement is what we usually treat long distance of urethral stricture.
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It may be successful if the construct is a scaffold with enough cells. If only scaffolds
are used without cells, the maximal length for tubularized replacement is 0.5 cm in
the animal study as aforementioned.4
CELLULAR ORIGIN IN TUBULARIZED REPLACEMENT:
The examples of the most common study designs for tissue-engineered urethras are
like the studies of De Filippo’s or Fu’s.3,7 In Fu and his colleagues’ study, the scaffolds
were allogeneic bladder submucosas, and cells were autologous foreskin epidermal
cells. They compared tubular grafts in 1, 2, 6 months using bladder submucosa with
or without foreskin epidermal cells in a rabbit model in vivo. They concluded that
acellular bladder submucosa seeded with epidermal cells could be used for
tabularized urethral replacement without stricture. However, the unseeded
tubularized bladder submucosa led to poor recovery and strictures of the urethras.
They used cell-labeling techniques to find that BrdU-stained foreskin epidermal cells
were found at 1 and 2 months after grafting, but not at 6 months. Therefore, they
thought that the epithelial cells of the graft originated and subsequently proliferated
from implanted epidermal cells, instead of extensions from surrounding transitional
cells.
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BIOACTIVE FACTORS:
Gene therapy of the vascular endothelial growth factor (VEGF) has been used as the
bioactive factor for urethral tissue engineering. Guan and his colleagues compared
the rabbit’s grafts after subcutaneous implantation into nude mice for 4 weeks using
rabbit bladder urothelial cells seeded into decellularized rabbit artery matrix with or
without VEGF ex vivo.8 Their scaffolds were decellularized rabbit carotid artery
matrices, and cells were rabbit bladder urothelial cells which were transfected with
MSCV-VEGF165-GFP (murine stem cell virus [MSCV]; green fluorescent protein [GFP])
retrovirus in experimental group or MSCV-GFP retrovirus in control group. They
found that VEGF-modified cells significantly enhanced neovascularization and the
formation of a urethral layer compared to GFP-modified cells. These results
indicated that VEGF gene therapy might increase the blood supply in tissue
engineering for treatment of urethral damage or loss.
HUMAN CLINICAL TRIALS:
These animal studies of tissue-engineered urethras have been transferred to human
clinical trials by 1999. Atala et al reported a clinical trial using unseeded bladder
submucosa for onlay replacement in hypospadias patients with 75% successful rate
in 1999.9 Bhargava et al also presented a clinical trial using tissue-engineered buccal
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mucosa for onlay replacement in human with 60% successful rate in 2008. 10 One of
the most remarkable clinical trials for urethral regeneration which gave us important
results and concepts was Raya-Rivera and his colleagues’ study.11 Synthetic
tubularized polyglycolic acid: poly(lactide-coglycolide acid) scaffolds were used. Both
autologous bladder smooth muscle and urothelial cells from previous urinary
bladder biopsy were harvested and expanded. Urothelial cells were seeded onto the
luminal surface and muscle cells onto the outer surface of the tubular scaffolds. Five
boys who had urethral defects underwent urethral reconstruction with the
tissue-engineered tabularized urethras. They remain functional in a clinical setting
for up to 6 years. To the best of our knowledge, this is the first successful clinical trial
in tabularized tissue-engineered urethral replacement.
CONCLUSIONS:
In conclusions, the maximum distance for normal tissue regeneration in tubularized
urethral replacement with unseeded matrices is 0.5 cm. Whilst autologous
tissue-engineered tabularized urethras were successful in clinical trial, this method
could be a new alternative treatment in urethral reconstruction.
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ACKNOWLEDGEMENTS:
This work was supported by a grant from the China Medical University Hospital
(DMR-95-053), Taichung, Taiwan.
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Figure legends
Figure. Onlay and tubularized replacement has been used in tissue-engineered
urethral regeneration.
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