Tissue engineering in reconstructive urology—The current status and critical insights to set future directions-critical review

Abstract

Advanced techniques of reconstructive urology are gradually reaching their limits in terms of their ability to restore urinary tract function and patients’ quality of life. A tissue engineering-based approach to urinary tract reconstruction, utilizing cells and biomaterials, offers an opportunity to overcome current limitations. Although tissue engineering studies have been heralding the imminent introduction of this method into clinics for over a decade, tissue engineering is only marginally applied. In this review, we discuss the role of tissue engineering in reconstructive urology and try to answer the question of why such a promising technology has not proven its clinical usability so far.

Full text

Tissue engineering in
reconstructive urology—The
current status and critical insights
to set future directions-critical
review
Karolina Ławkowska
1
*, Clemens Rosenbaum
2
*, Piotr Petrasz
3
*,
Luis Kluth
5
*, Krzysztof Koper
4
*, Tomasz Drewa
1
*,
Marta Pokrywczynska
1
*, Jan Adamowicz
1
* and
the Trauma and Reconstructive Urology Working Party
of the European Association of Urology Young Academic
Urologists
†
1
Department of Regenerative Medicine, Collegium Medicum, Nicolaus Copernicus University,
Bydgoszcz, Poland,
2
Department of Urology Asklepios Klinik Barmbek Germany, Urologist in
Hamburg, Hamburg, Germany,
3
Department of Urology Voivodeship Hospital Gorzów Wielkopolski,
Gorzów Wielkopolski, Poland,
4
Department of Clinical Oncology and Nursing, Collegium Medicum,
Nicolaus Copernicus University, Curie-Skłodowskiej 9, Bydgoszcz, Poland,
5
Department of Urology,
University Medical Center Frankfurt, Frankfurt am Main, Germany
Advanced techniques of reconstructive urology are gradually reaching their
limits in terms of their ability to restore urinary tract function and patients’quality
of life. A tissue engineering-based approach to urinary tract reconstruction,
utilizing cells and biomaterials, offers an opportunity to overcome current
limitations. Although tissue engineering studies have been heralding the
imminent introduction of this method into clinics for over a decade, tissue
engineering is only marginally applied. In this review, we discuss the role of
tissue engineering in reconstructive urology and try to answer the question of
why such a promising technology has not proven its clinical usability so far.
KEYWORDS
tissue engineering, reconstructive urology, regenerative medicine, urology,
biomaterials
1 Introduction
Tissue engineering has evoked hopes over the last few decades for new therapies aimed at
replacing an injured or resected urethra, urinary bladder, or ureter. To offer this possibility,
different biomaterials combined with cells were applied to create an artificial wall of the
urinary tract, restoring function (Adamowicz et al., 2013). Advanced techniques of
reconstructive urology supported by modern surgical tools are reaching their limits in
terms of their ability to restore urinary tract function and a patient’s quality of life. Tissue
engineering has been considered the ideal strategy to push reconstructive urology to the next
OPEN ACCESS
EDITED BY
Bryan Brown,
University of Pittsburgh, United States
REVIEWED BY
Prashanth Ravishankar,
Namida Lab, Inc., United States
Reetta Sartoneva,
Tampere University, Finland
*CORRESPONDENCE
Karolina Ławkowska,
karolinalawkowskaa@gmail.com
Clemens Rosenbaum,
rosenbaumclemens@gmail.com
Piotr Petrasz,
Petraszpiotr@gmail.com
Krzysztof Koper,
krzyskoper@gmail.com
Luis Kluth,
luiskluth@me.com
Tomasz Drewa,
tadrewa@gmail.com
Marta Pokrywczynska,
marta.pokrywczynska@interia.pl
Jan Adamowicz,
adamowicz.jz@gmail.com
†
Members of the consortium are listed at
the end of the article
SPECIALTY SECTION
This article was submitted to Tissue
Engineering and Regenerative Medicine,
a section of the journal
Frontiers in Bioengineering and
Biotechnology
RECEIVED 09 September 2022
ACCEPTED 13 December 2022
PUBLISHED 01 March 2023
CITATION
Ławkowska K, Rosenbaum C, Petrasz P,
Kluth L, Koper K, Drewa T,
Pokrywczynska M, Adamowicz J and
the Trauma and Reconstructive Urology
Working Party of the European
Association of Urology Young Academic
Urologists (2023), Tissue engineering in
reconstructive urology—The current
status and critical insights to set future
directions-critical review.
Front. Bioeng. Biotechnol. 10:1040987.
doi: 10.3389/fbioe.2022.1040987
COPYRIGHT
Copyright © 2023 Ławkowska, Rosenbaum,
Petrasz, Kluth, Koper, Drewa,
Pokrywczynska, Adamowicz and the
Trauma and Reconstructive Urology
Working Party of the European Association
of Urology Young Academic Urologists. This
is an open-access article distributed under
the terms of the Creative Commons
Attribution License (CC BY). The use,
distribution or reproduction in other forums
is permitted, provided the original author(s)
and the copyright owner(s) are credited and
that the original publication in this journal is
cited, in accordance with accepted
academic practice. No use, distribution or
reproduction is permitted which does not
comply with these terms.
Frontiers in Bioengineering and Biotechnology frontiersin.org01
TYPE Review
PUBLISHED 01 March 2023
DOI 10.3389/fbioe.2022.1040987

level, where urologists would utilize cell-seeded biomaterials and
stem cells in daily practice (Adamowicz et al., 2019a). The approach
intended to create urological grafts, i.e., whole substitutes or tissues
that can be implanted, regenerated, or permanently replace the
urethra, urinary bladder, or ureter. Despite plenty of valuable
research data revealing the biology of stem cells, the behavior of
implanted adult stem cells, and the remodeling of biomaterial grafts
within urinary tracts, tissue engineering is nowadays marginally
influencing urological management. The tissue engineering-
oriented sessions, so numerous in recent years, gradually
disappeared from the scientific programs at essential urological
meetings. This may be interpreted as a sign of skepticism about the
relevance of tissue engineering for urological therapy. On the other
hand, researchers responsible for the development of tissue
engineering led to urologists’awakening through the publication
of results suggestive of groundbreaking outcomes. From the
clinician’s perspective, the simplification of research models and
the dominating positive interpretation of results made from tissue
engineering achievements in urology produce few valuable reports.
This review aims to summarize and critically evaluate the role
of tissue engineering in reconstructive urology and to provide
informative data presenting the current status rather than
focusing on remaining problems or glorifying achievements.
2 Current challenges in
reconstructive surgery–Urological
surgeon’s perspective
The current challenges in reconstructive urology consist of
three major points. The first and most crucial challenge is to
achieve the best oncologic results possible. The second challenge
is to achieve the best functional results. The third driver in
reconstructive surgery should be the goal of lowering morbidity
at the explantation site.
New reconstructive material must fulfill these requirements.
Due to the currently used reconstructive materials, it is no
wonder that tissue-engineered materials or materials that do
not need to be harvested, for example, amniotic membranes, have
been investigated throughout the last century (Kaleli and Ansell,
1984). The primary driver of these efforts is the limited functional
results and the side effects of tissue harvesting nowadays.
Autologous tissues are used, i.e., intestinal segments for upper
tract and bladder reconstruction, and skin or oral mucosa for
urethral reconstruction (Xiong et al., 2020).
In bladder reconstruction, intestinal segments guarantee
excellent oncological results. Recurrence-free survival in patients
treated with radical cystectomy for bladder cancer ranges between
60% and 68% at five-year follow-up. Of those, local recurrences
account for 30%–54% (Mari et al., 2018). Pathologic stage, lymph
node invasion, the extent of lymphadenectomy, multifocality, and
prostatic involvement were found to be independent predictors of
pelvic recurrence, whereas the type of urinary diversion was not
(Umbreitetal.,2010). In general, recurrence in the intestinal segment
used to create a neobladder or ileal conduit is infrequent. However,
the price for reasonable oncologic results is high. Radical cystectomies
with urinary diversion come with an early complication rate of
almost 100%, a 25% likelihood of readmission to the hospital, and a
5% risk of perioperative death (Vetterlein et al., 2019). Most of these
side effects are related to resecting the intestinal segment.
Gastrointestinal-related complications represent a significant
portion of short- and long-term complications. Within the first
30 days postoperatively, almost 20% develop gastrointestinal
problems. Further on, metabolic complications related to the use
of the intestinal segment remain a long-term problem. Metabolic
acidosis and chronic renal failure lower the life expectancy and
quality of life of patients. Summed up, radical cystectomy and urinary
diversion represent surgery-related high-risk complications. Almost
all patients experience postoperative complications, and most of
them can be explained as explantation site-related (Umbreitetal.,
2010). Further functional results are also unsatisfactory. In the long
term, complications that need to be operated on, such as stenosis of
the ileal conduit or the ileal ureter anastomosis, are described in 12%–
24% of cases (Lee et al., 2003)(Hautmann et al., 2011). This
underlines the need for novel materials to be used for urinary
diversion (Kloskowski et al., 2015a).
In urethral reconstruction, the oncologic outcome does not
play a role, whereas functional results are of significant interest.
Functional results depend on different stricture-related factors,
such as bulbar stricture location or shorter strictures. If both facts
come together, no substitution material should be used (Morey
et al., 2014). Excision and primary anastomosis of the urethra are
indicated and guarantee excellent results (Chapple et al., 2014).
For penile or longer and more complex strictures, substitution for
urethroplasty is indicated. Nowadays, buccal mucosa graft is the
most commonly used substitution material. This substitution
material has ruled out others like penile skin due to higher
success rates. Still, success rates of buccal mucosa graft
urethroplasties are lower than those of urethroplasties by
excision and primary anastomosis. Success rates of buccal
mucosa graft urethroplasties range between 70% and 87%
(Vetterlein et al., 2018). When taking into consideration that
these results include complex and lengthy strictures, the results
can be seen as very good. Nevertheless, the side effects of harvest
site grafting are not negligible. Efforts have been made to improve
the management of the harvest site (Soave et al., 2018). However,
oral complaints are common, especially in patients who require
longer grafts due to longer strictures (Rosenbaum et al., 2016).
Oral complaints consist of pain, bleeding, swelling, numbness,
alteration of salivation and taste, and also impairment of mouth
opening, smiling, whistling, diet, and speech. These complaints
can reduce the quality of life for patients. Therefore, alternative
substitution material has been sought and is currently being
tested in clinical trials. A tissue-engineered oral mucosal graft
(MukoCell
®
) has been used for the typical indication of buccal
mucosa grafts and showed comparable success rates of 84%
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(Barbagli et al., 2018). Even though the application of this tissue-
engineered substitution material is described as being more
complicated than the application of buccal mucosa grafts,
functional results are very encouraging.
3 Why can we offer so little to our
patients?
Whenever a new concept of medical therapy becomes
available, it needs to be validated in clinical trials. Despite
research efforts, tissue engineering is struggling to introduce
reliable therapeutic options to clinics. Currently, only in the
field of urethral reconstructive surgery have clinical trials
involving tissue engineering shown relevant results, justifying
the continuation of research (Kanematsu, 2018). Despite the
media attention, research, and community interest, tissue
engineering therapies did not reach the mainstream
application. They are limited by their inability to effectively
recapitulate the complex cellular, structural, and mechanical
environment of native tissues when transitioning from in vitro
to in vivo applications (Figures 1,2)(O’Donnell et al., 2019).
3.1 Questionable effectiveness in clinical
trials
3.1.1 Urinary bladder
The landmark study of Atala et al. (2006) aimed to augment
the bladder in spina bifida patients with a collagen scaffold
FIGURE 1
Overview of tissue engineering strategies in reconstructive urology. (A) Cell types used for the development of implantable grafts. (1) Urothelial
cells; progenitors of urothelial cells derived from urine. (2) Smooth muscle cells derived from the bladder detrusor; smooth muscle cells progenitors
derived from mesenchymal cells. (3) Mesenchymal cells derived from bone marrow or adipose tissue, (B) Biomaterials used in tissue-engineered
urinary tract reconstruction. (1) Natural biomaterials, et cetera, amniotic membrane, small intestinal submucosa SIS (2) Decellularized scaffolds,
et cetera. Acellular matrix of the bladder (BAM) (3) Polymer biomaterial, et cetera, collagen, gelatin, alginate, cellulose, and chitin (4) Scaffolds with
incorporated bioactive components, et cetera, growth factors, smart biomaterials, (C) A tissue-engin eered approach to experimental urinary tract
reconstruction. (1) patch grafts aimed to replace urinary bladder wall in partial cystectomy model (2) patch grafts or tubular grafts for whole full-
thickness urethral reconstruction (3) artificial urinary conduits (4) patch grafts or tubular grafts for ureteral reconstruction.
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FIGURE 2
Tissue-engineered reconstruction of the urinary tract wall (A) A normal urinary tract wall, composed of a stratified urothelial layer (1) with
interstitial cells beneath forming a syncytial-like layer (2) layered arrangement of the detrusor muscle layer (3) complex interstitial neuronal network
(4), (B) Urinary tract wall reconstruction with cell-seeded grafts (1), (C) Early (<3 months) regeneration results showed partial restoration of proper
structure. All layers exhibited extensive structural and histological disturbances. Hypertrophic urothelium (1) with an incomplete external layer.
No one of the studies focused on the regeneration of regulatory intestinal cells or the neuronal network. It is likely, however, that regeneration of
these components does not occur. Extensive progressive fibrotic reaction due to ongoing inflammation and ischemia within the graft. Smooth
muscle layer regrowth is irregularly arranged and gradually loses its native spatial configuration (2), (D) Seeded cells (1) are predominantly
phagocytized (2) shortly after reconstruction by activated macrophages. The surveillance time of cells seeded on the biomaterial graft after
implantation is not certain, but it is estimated at several weeks. In light of this data, seeded cells provide a temporary boost for limited natural
regeneration mechanisms by supplying the environment with bioactive compon ents, (E) With the persistence of inflammation, the tissue remodeling
with the reconstructed urinary tract wall is oriented toward fibrosis , (F) The final reconstruction outcomes are not satisfying. Proper restoration of the
urothelial layer is due to the high intrinsic regeneration potential of the epithelium. Nevertheless, it is unknown whether the complex regulatory
function of urothelium is preserved. The biomaterial underwent efficient degradation and was replaced by collagen-rich scar tissue (2), covering
most of the reconstructed area.
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seeded with autologous cells. Indeed, the concept demonstrated
new frontiers for patients and promised implementation of this
solution in clinics. The bladder dome-shaped collagen scaffold
presented here, pre-seeded with autologous urothelial and
smooth muscle cells for augmentation, seemed to be a viable
solution. Following 46 months of follow-up, including a series of
urodynamic tests, the bladder function was good, with reliable
results. Thanks to this seemingly successful study, the attention
of urologists and investors was turned toward the tissue
engineering industry. Unfortunately, the study was not
continued, and the urological community was left with
unanswered questions, mainly regarding the durability of this
management and long-term complications. The study by Atala
et al. was a spark to start Tengion, a company that aimed to
fabricate commercially available electrospun biomaterial
scaffolds for universal urinary tract wall replacements. Neo-
Urinary Conduit and Neo-Bladder patches were evaluated
during phase I and phase II clinical trials, respectively
(ClinicalTrials, 2004)(Joseph et al., 2014). Unfortunately,
none of these products met expectations and successfully
completed the trials, and the reasons for failure were sparingly
discussed to define objectives for therapy improvement.
Disclosure of Tengion graft behavior and remodeling in vivo
after implantation to patients would be particularly valuable to
set future research directions. Similarly, the results of an artificial
urinary conduit study were not made publicly available. Tengion
biomaterial most likely failed to prevent fibrotic reaction and
scarring, resulting in gradual loss of initial elasticity and
compliance necessary for integration with the urinary tract.
Insufficient angiogenesis within the graft likely led to hypoxia-
related fibrosis. Since 2014, no registered study has planned to
evaluate new biomaterials or cell-seeded grafts for tissue-
engineered urinary diversion.
3.1.2 Urethra
The field of urethral reconstruction, contrary to the
suspicious nature of this paragraph, is so far from being the
most solid argument for tissue engineering supporters. There are
eight reports available, involving 180 patients who underwent
urethra reconstruction procedures using tissue-engineered grafts
(Barbagli et al., 2018)(Romagnoli et al., 1990)(Romagnoli et al.,
1993)(Bhargava et al., 2008)(Raya-Rivera et al., 2011)(Lazzeri
et al., 2014)(Ram-Liebig et al., 2017)(Osman et al., 2014). The
histological structure of the urethra is less complicated than the
bladder, and there is not a complex functional background.
Nevertheless, in terms of urethral reconstruction, the major
challenge concerns fibrosis within the graft’s lumen, averaging
8mm–9 mm or less, which is responsible for stricture recurrence
(Mangera et al., 2010).
In recent years, MukoCell
®
, a personalized tissue-engineered
autologous graft, has gained much attention from the
reconstructive urology community due to several
accomplished clinical studies. MukoCell
®
is a laboratory-
grown graft from cells of the oral mucosa used in the
treatment of urethral stenosis. (Barbagli et al., 2018)(Lazzeri
et al., 2014)(Ram-Liebig et al., 2017). Reported results
underlined that the application of MukoCell
®
for urethroplasty
guaranteed similar success rates to the native buccal mucosa. It
must be admitted that the harvesting of a patient’s buccal mucosa
epithelial cells during an off-patient clinic procedure to create a
transplantable graft is the quintessence of tissue engineering
management. Nevertheless, the enthusiasm should be
tempered to allow a reliable assessment of this technology.
The major question mark arises due to the homogeneity of
the available reports, which are continuously derived from the
same centers involved simultaneously in MukoCell
®
commercialization. This product was not evaluated in large-
scale trials by independent research teams. Moreover, there is
limited available data transparently demonstrating the
MukoCell
®
preparation method and graft safety (one poster)
(Lazzeri et al., 2014). A relatively short 12-month follow-up used
in all MukoCell
®
trials raises doubts for reconstructive surgeons
in terms of therapy efficiency and superiority over standard
management. In regards to the pathophysiology of wound
healing after biomaterial implantation, this period is not
enough to document graft resistance to inflammatory or
fibrotic narrowing (Anderson et al., 2008). Delayed response
to implanted biomaterials lasts up to 24 months after the initial
procedure, and during all this time, slowly progressive scarring
occurs (Morris et al., 2017).
Current urethroplasty techniques based on buccal mucosa
are effective treatment modalities, so why does current urology
need alternative materials for reconstruction? One of the reasons
is the fast depletion of treatment methods for challenging cases,
recurrent stenosis, and pediatric hypospadias. These patients are
in real need of important therapeutic advances in segmental
urethral replacement. El Kassaby et al. (2008) demonstrated in a
randomized trial that the use of tissue-engineered human bladder
acellular matrix (BAM) was a viable option for complex anterior
urethral repair (El Kassaby et al., 2008). The leading concept of
this study was to create an off-the-shelf biomaterial, an acellular
product intended to become an alternative for buccal mucosa.
Thirty patients underwent urethroplasty with BAM due to
stricture lengths ranging from 2 cm to 18 cm and were
followed for 36 months. The regeneration of the urethral wall
was believed to start from native mucosa, whereas BAM was
planned to generate an excellent environment for neo-tissue
formation. The authors did admit, however, the lower success
rate in the BAM group, especially in patients who had previously
undergone interventions. This was an important observation that
documented the inferiority of the acellular strategy and mostly
depended on the urethral mucosa epithelial cells’ability to
populate the scaffold and reconstitute the regrown consistent
layer. Following that, the same center published an observational
study with five boys evaluating grafts made of tabularized poly
(glycolic acid) (PGA) seeded with autologous urothelial and
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smooth muscle cells (Raya-Rivera et al., 2011). Tissue-engineered
urethras, as described by the authors, were used for segmental
urethroplasty, which is the most demanding technique due to the
high failure rate in urethral surgery. In this setting, the uroflow
analysis showed unobstructed urine flow up to 72 months after
surgery. The costs of the treatment were not revealed. Despite the
great success, the clinical value of research based on a few cases is
rather low due to the inability to prove the superiority of the
novel, more expensive method over standard management.
Moreover, a description of the applied methodology would be
difficult to comply with in other centers willing to test this option.
The unspoken issue was the potential impact of a patient’s young
age on therapeutic outcome. Human tissue’s ability to regenerate
declines with age due to the loss of stem/progenitor cell function
(Sousounis et al., 2014). Therefore, the outcomes of the methods
involving individual regeneration potential might be anticipated
to be worse in adults.
3.1.3 Urinary incontinence
Tissue engineering efforts have concentrated on therapy for
stress urinary incontinence (SUI) in women for many years.
The experimental restoration of a damaged urethral sphincter
has been carried out by cell transplantation of autologous
myocytes, muscle-derived stem cells, and adipose tissue-
derived stem cells (Pokrywczynska et al., 2016). Although
similar cell populations are applied, methodologies and data
acquisition are heterogeneous, making a comparison of results
and choosing an adequate technique difficult. Despite these
inconsistencies, available clinical trials showed that cell-based
therapy had a high success rate in SUI treatment (Cornu et al.,
2014)(Kuismanen et al., 2014). Most of the available trials
presented short-term benefits regardless of the material used,
including placebo saline injections. A thorough analysis of the
data indicated that cell-based therapy did not turn out to be
effective in clinical practice and should not be recommended to
patients. Most of the available trials reported a short-term
benefit corresponding to the effectiveness of placebo saline
injections. Endpoints were based on subjective parameters,
usually non-validated life-quality questioners. Interestingly,
none of the studies documented using the cough test as one
of the tools for evaluating SUI after therapy. As is commonly
demonstrated, an increased urethral pressure profile must be
interpreted with caution due to the uncertain diagnostic
resolution of this method. The issue hardly ever deliberated
in the studies’“discussion”paragraph is the demarcation
between improvement mediated by the bulking effect and
recovery of sphincter function due to alleged induced
regeneration (Vinarov et al., 2018).
3.1.4 Ureter
To our knowledge, there is not any study evaluating the tissue
engineering approach for ureter replacement/reconstruction in a
clinical trial.
3.2 Costs of tissue engineering research
The global tissue engineering market is categorized into
therapeutic products, tools, banks, and services. The
manufacturer price of tissue-engineered products ranges
between US$ $18,950 and US$ 93,432 on average (Schneider
et al., 2010). The major segments of tissue engineering products
include cell therapy, gene therapy, and tissue replacement. It is
estimated that the global tissue engineering market will exceed
US$ 94.7 billion in the near future, with a CAGR of 23%
(MarketWatch, 1997). Recognizing this potential, the National
Institutes of Health (NIH) of the United States invested an
estimated $940 million in regenerative medicine research in
2018 alone (National Institutes of Health (NIH), 2016).
Nevertheless, the optimistic data does not reflect the
significance of tissue engineering therapies in clinical practice.
At this point, it must also be underlined that MukoCell
®
is the
only tissue-engineered product for the urologic patient. One of
the important factors hampering the translation of experimental
tissue engineering therapies is the high cost of treatment in
comparison to the potential results. All studies evaluating
tissue engineering therapies for urethra or urinary
incontinence had a power of less than 0.8 (Vickers and
Sjoberg, 2015). As a consequence, underpowered studies
cannot convince medical care authorities to fund and widely
implement this approach in clinics. Nowadays, considering the
lack of research evidence, the question arises whether tissue
engineering in urology is economically viable. In our opinion,
this technology should be at the initial stage of development, and
the focus should be on the confirmation of its effectiveness rather
than delivering case reports. Another priority is to reduce efforts
and concentrate on the pharmacoeconomic aspects of tissue-
engineered procedures. Investigators should critically evaluate
what effects their research will have on their future market and
whether an established company would welcome a new product.
Startup companies have options as well. During the initial
investigation, small markets may not seem appealing to
entrepreneurs (Bayon et al., 2015). However, treatment with
unorthodox, technologically advanced therapies makes it more
feasible to enter the smaller markets first. The best example is
the success of MukoCell
®
in reconstructive surgery of the
urethra, rather than being a niche field of tissue engineering
application. Apart from its effectiveness, it is used in clinical
practice daily, and supporters of this method are experts in the
field.
The cost of funding and discovery of tissue engineering
products is primarily supported by small-to medium-sized
companies in collaboration with university units receiving
government or private research grants. Big pharma companies
are gradually increasing their investments in tissue engineering,
including urology, but it is still in its early stages, and the amount
of funding available has thus far been crowded out by their near-
term revenue priorities (Statnews, 2019). If tissue engineering is
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to revolutionize medicine, particularly urology, the
disproportionately distributed funding sources must be
rearranged. Careful consideration should be given to new
funding models and tax incentives that will attract new
sources of capital for interdisciplinary research groups
combining biotechnologists, doctors, and biomaterial experts
(Adamowicz et al., 2017a).
3.3 Law procedures
Cutting-edge tissue engineering therapies that combine
living cell transplantation with biomaterials are among the
most complicated in terms of clinical trials, regulations, and
the field of medicine. Tissue Engineered Medical Products
(TEMPs) intended for tissue repair, and replacement are
FIGURE 3
Comprehensive overview of tissue-engineered product certification in the EU and the United States.
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qualified as Cell-based Medicinal Products (CBMPs) or
Advanced Therapy Products (ATPs) (EMEA/CHMP, 2006)
(Johnson et al., 2011). In the USA and Europe, the approval
criteria for TEPs are regulated by the FDA and the European
Medicines Agency (EMA), respectively (Figure 3)(U.S. Food and
Drug Administration, 2019)(European Medicines Agency,
1995). In contrast to the national regulatory framework, a
different policy is applied to the regulation of cell therapy
products to be marketed in the countries of the EU.
Accordingly, there is one centralized procedure across the
countries of the EU. Analogously, in the United States, the
Center for Biologics Evaluation and Research (CBER)
regulates cellular and biomaterial-based therapies as a part of
the FDA (Center for Biologics Evaluation and Research (CBER),
2022). Good Manufacturing Practice (GMP) guidelines establish
quality control standards for the manufacturing of ATMPs and
are international, comprehensive, and mandatory to follow
(Greenberg-Worisek et al., 2018). To date, only a few TEMPs,
mostly from the field of oncologic hematology have been
approved by the FDA and EMA (O’Donnell et al., 2019). It
also reflects the unbalanced readiness of different branches of
medicine to apply these therapies in clinical practice. In contrast
to hematological departments, urological departments lack
interdisciplinary equipment for in vitro cellular manipulation.
The only solution is to outsource the manufacturing process of
TEMPs and deliver them “ready to use”. MukoCell
®
successfully
implemented this strategy. A significant disadvantage of the
outsourcing strategy, if widely used, is the possibility of a loss
of integrative supervision by clinicians throughout therapy and
the risk of quality issues. Another option is to establish universal
biotechnological units within organizational structures of leading
healthcare providers to administer, manage, and serve as a local
advisory board for TEMPs. Moreover, the complexity and
fragility of TEMPs with regard to their vitality necessitates the
education of clinical personnel to obtain a background in stem
cell biology and biomaterial science.
The development of TEMPs demands the integration of stem
cells, biomaterials, and chemicals, making it exceptionally
challenging for market authorization and commercialization.
Researchers often adopt the wrong strategy when they plan to
spin out a product from their laboratory. Specifically, they start to
approach regulatory issues after the novel product is at the end of
its development phase. While they should plan a regulatory path
at the beginning of their project, this model of research
management is already encouraged by authorizing agencies.
The EMA provides an expert panel to categorize and select
the best regulatory pathway for novel TEMPs (TEMPs, 1995).
(TEMPs, 1995)(TEMPs, 1995)(TEMPs, 1995)For instance,
tissue-engineered Xenograft tissues are qualified as device
biologics by the EMA instead of TEMPs. As a result, they go
through a simplified certification process (European Medicines
Agency, 2011). Choosing components according to regulatory
guidelines at the inception of research is an underestimated
factor that has a crucial impact on authorization procedures
during preclinical and clinical trials.
Potentially, a wide range of TEMPs dedicated to
reconstructive urology will emerge as science evolves.
Realistically, clinicians, including urologists, are not trained to
overcome the complex aspects of clinical effectiveness and safety,
along with the regulatory and ethical issues of TEMPs. This
situation may raise uncertainty and cause the reluctant
introduction of innovative therapies into clinical practice. To
increase the likelihood of clinical success, it is high time to
gradually familiarize clinicians with the regulatory bases of
therapies that are based on genes, tissues, or cells. The
education of clinicians will positively influence their
involvement in preclinical trials. By implementing
developments in TEMP made recently by medical authorities,
investigators can reduce the gap between the bench and the
patient.
4 Achievements–Critical evaluation
4.1 Urothelial layer regeneration
Regeneration of the urothelial layer of the reconstructed
urethra, urinary bladder, or ureter determines the
reconstitution of the barrier against urine. Table 1 shows the
achievements in urothelial layer regeneration. Against the dogma
of reconstructive urology that urine acts as a source of nutrients
for regenerating the urothelial layer, it has a devastating effect on
exposed cells without the protection of the umbrella cell layer
(Adamowicz et al., 2012). Difficulties in eliminating urine contact
with the regenerating area are one of the reasons for the enhanced
fibrotic response. Urothelium, like all epithelia, has an intrinsic
tendency to spontaneously regrow from the surrounding tissue
and to reform a stratified layer (Kloskowski et al., 2017). Future
tissue engineering therapies will require technology to obtain a
sufficient number of urothelial cells (UCs) in vitro, ready for
transplantation or graft creation. UCs can be isolated from the
native urothelium, requiring open or endoscopic surgery for
tissue sampling (Kloskowski et al., 2014). There is no
clinically validated protocol determining how much urothelial
tissue is needed or from which urinary tract region the tissue
should be harvested. In the case of invasive procedures, such
protocols must be formulated to be obligatorily followed. In
October 2018, the clinical trial “Urothelium Tissue Engineering
Using Biopsies from Transurethral Resection of Prostate”was
launched, and its results might serve as a base to define guidelines
optimizing the creation of autologous tissue-engineered
urothelium (J.-L. L. Nicolas Berte, 2018).
An alternative solution was proposed earlier by Nagele et al.
(2008) who showed that human UC cultures could be efficiently
established from bladder washings. Stratified cultures and
detached sheets stained 100% positive for pan-cytokeratin and
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CK20, indicating differentiation into superficial cells.
Notwithstanding this fact, Uroplakin III expression, which is a
specific marker for umbrella cells, was not observed. Cell sheet
viability was confirmed by rapid cell outgrowth in explant
cultures. An important advantage of this study was the
exclusion of contentious p63-positive mesenchymal cells.
Therefore, UCs obtained in vitro in this setting originated
from precursor cells or activated mature UCs. This study
raised many questions regarding the efficiency of the
presented method and the quality of urothelial cells used to
establish primary cultures.
In most of the diseases requiring urinary tract
reconstruction, autologous urothelial cells are not suitable
for application (Drewa et al., 2012). The cardinal
contradiction is urothelial cancer and the related risk of
panurothelial disease. Presently, there is no available method
to identify and separate cancerous cells. Additionally, even in
the case of benign conditions needing reconstruction,
autologous cells might be compromised. For instance, the
urothelium derived from obstructed urinary tracts or after
chronic infections showed a lower survival rate and potential
to replicate (Lee et al., 2016). When healthy urothelium is not
available, stem cells or even dedifferentiated mature cells may
be used as an alternative cell source.
USCs are currently being explored as a natural, accessible
source for UC generation to be used in tissue engineering. USCs
TABLE 1 Achievements in urothelial layer regeneration.
Study In vitro/In vivo
study
Used cell type Achievement
Nagele et al.,
(2008)
in vitro Human UCs Human UCs isolated from bladder washings were used to make urothelial sheets
Lu et al., (2019) in vitro Pluripotent stem cells identified in
urine
Identifying pluripotent stem cells in urine that might also belong to mesenchymal
stromal cells - like progenitor cells that reside within the bladder wall
Wan et al.,
(2018)
in vitro Urine-derived stem cells (USC) Obtaining a differentiated epithelium of the urinary tract from USC.
Tian et al.,
(2010)
in vitro Bone marrow mesenchymal stem
cells (BMSCs)
Urothelium-like cells derived from human BMSCs can be used as an alternative source
of cells for urinary bladder reconstruction
Inoue et al.,
(2019)
in vitro in vivo UCs First successive transplant of urothelial cells into the bladder lumen
Suzuki et al.,
(2019)
in vitro Pluripotent stem cells (hiPSCs) Established a protocol for the directed differentiation of hiPSCs into stratified bladder
urothelium
TABLE 2 Achievements in the regeneration of the smooth muscle layer.
Study In vitro/In vivo
study
Used cell type Achievement
Opitz et al., (2007) in vitro Vascular SMCs Determination of phenotypic plasticity of vascular-derived SMC which allows
oscillation between proliferative and differentiated phenotype depending on
pressure stress conditions
Pokrywczynska et al.,
(2019)
in vivo Adipose tissue-derived mesenchymal
stromal cells (ASCs)
ASCs create an environment rich in morphogenetic signals corresponding to
an early organogenesis environment when the mesenchymal forms an early
smooth muscle layer
Liu et al., (2017) in vivo USCs The use of a porous SIS scaffold seeded with USCs allows for the regeneration of
the urethra
Mirzaei et al., (2019) in vitro Human iPSCs Human iPSCs seeded on poly (lactic-co-glycolic acid) (PLGA) resulted in
acquiring a smooth muscle
Hoogenkamp et al.,
(2016)
in vitro UCs, SMCs Development of a method of obtaining collagen scaffold of the size of the entire
urinary bladder
Ardeshirylajimi et al.,
(2018)
in vitro MSCs The scaffold releasing TGF-βand seeded with MSCs allows for bladder
regeneration
Opitz et al., (2007) demonstrated the phenotypic plasticity of vascular-derived SMC, and their ability to oscillate between a proliferative and differentiated phenotype in response to pressure
stress conditions. This phenomenon might be an essential clue in efforts to regenerate human detrusor muscles, as it shows the necessity of stimulating regenerating tissue through
physiological pressure stress. This relationship must be kept in mind because, in urology, there is a tendency to preserve long-lasting decompression of healing regions by catheterization.
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TABLE 3 Achievements in the regeneration of bladder innervation.
Study In vitro/In vivo
study
Used cell type Achievement
Madduri et al.,
(2009)
in vitro Dorsal root ganglion (DRG) Glial cell line-derived neurotrophic factor (GDNF) significantly influences
axonal elongation, and nerve growth factor (NGF) induces extensive axonal
branching
Kikuno et al.,
(2009)
in vivo -NGF may support the regeneration of a functional bladder formed from BAM
Nitta et al., (2010) in vivo Skeletal muscle-derived multipotent Sk-34
and Sk-DN stem cells
The transplant allowed for significant functional recovery (80%) thanks to the
incorporation of the transplanted cells into the damaged peripheral nerves and
blood vessels
Adamowicz et al.,
(2011)
in vitro Schwann cells Protocol for the effective isolation of Schwann cells from a pre-degenerating
peripheral nerve
TABLE 4 Application of biomaterials in urethral reconstruction.
Study Biomaterial In vitro/In
vivo study
Model Outcome
Dorin et al.,
(2008)
Acellular collagen matrix in vivo rabbit It was established that .5 cm is the maximum defect distance that can
support proper tissue formation using acellular tubularized grafts
Orabi et al.,
(2013)
BAM in vivo canine The use of cell-seeded tubularized urethral scaffolds allows for the
repair of defects up to 6–7 cm long
Jia et al., (2015) Collagen scaffold modified with collagen
binding domain (CBD) VEGF
in vivo canine The use of CBD-VEGF allowed for a much thicker epithelial cell layer
compared to the collagen group
Pinnagoda et al.,
(2016)
Acellular double-layered collagen scaffolds in vivo rabbit After nine months of research, they observed significant changes in
acellular double-layered collagen scaffolds, obtaining a structure
similar to the normal urethral tissue
Shakeri et al.,
(2009)
Amniotic membrane in vivo rabbit Urethral reconstruction was successful in all 20 operated rabbits, with
no inflammation or tissue loss
Güneşet al.,
(2017)
Buccal mucosa and amniotic membrane in vivo rabbit The combination of buccal mucosa and amniotic membrane for
ventral onlay penile urethroplasty contributed to better tissue healing
Koziak et al.,
(2007)
Amniotic membrane in vivo human Regeneration of extensive ureteral defects without serious
complications
TABLE 5 Application of biomaterials in the reconstruction of the urinary bladder.
Study Biomaterial Biomaterial fabrication method In vitro/In
vivo study
Model Outcome
Horst et al., (2017) PLGA on a BAM A hybrid microfibrous scaffold was obtained
by direct electrospinning of PLGA on a BAM
in vivo rat The seeded scaffolding ensured layered
regeneration of the bladder walls in vivo
Pokrywczynska
et al., (2018)
BAM BAM was obtained by a multistep detergent
washing procedure
in vivo pig The tissue-engineered bladder worked
normally. Stem cells additionally supported
the regeneration of the urinary bladder
Jiang et al., (2016) PLGA nanoparticle-
modified BAM
BAM was incorporated with VEGF and
bFGF-loaded PLGA nanoparticles and
mixed with a hydrophilic gel
in vivo rabbit The scaffolding developed proved to be an
effective method for achieving long-term
sustained release of VEGF and bFGF.
Zhou et al., (2013) BAM BAM was incorporated with platelet-derived
growth factor-BB and VEGF.
in vivo rabbit BAM combined with PDGF-BB and VEGF
significantly improves muscle contractility
and angiogenesis
Adamowicz et al.,
(2020)
AM and graphene Graphene layers were transferred without
modifying the AM surface
in vitro - Composite made of AM, graphene, and
seeded with smooth muscle cells was capable
of induced contraction in vitro
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are a heterogenous cell population without a clear isolation
methodology, and further segregation is required to
distinguish the most useful subpopulations. There are also
concerns regarding available isolation protocols,
i.e., repeatability and efficiency (Kloskowski et al., 2015b). In
general, USCs are described predominantly as pluripotent stem
cells supposedly derived from renal epithelium due to high gene
expression for kidney cortex markers (Pavathuparambil Abdul
Manaph et al., 2018). According to a recent study by Lu et al.
(2019) pluripotent stem cells identified in urine might also belong
to a newly described mesenchymal stromal cell-like progenitor
cell that resides within the bladder wall. Wan et al. (2018)
reported that urothelial-conditioned medium combined with
dynamic flow culture-induced human USC differentiation into
UC with a developed uroplakin protective barrier. Differentiated
USCs expressed urothelial-specific transcripts and proteins
(Uroplakin III and Ia), epithelial cell markers (CK20 and
AE1/AE3), and tight junction markers (ZO-1, ZO-2,
E-cadherin, and Cingulin). Also, obtained UC grown on
collagen matrix spontaneously formed a multilayer structure
corresponding to a healthy urothelial layer.
Various types of mesenchymal stem cells (MSC), including
bone marrow-derived or adipose tissue MSC, showed a capacity
to induce urothelial cell differentiation (Xiong et al., 2015). Tian
et al. (2010) postulated that epidermal growth factor, platelet-
derived growth factor-BB, transforming growth factor-beta1,
and vascular endothelial growth factor were crucial to initiate
and maintain the urothelial differentiation pathway.
Biotechnology is nowadays taking the first steps to develop
scientific know-how in terms of controlled in vitro
differentiation of adult cells into different cell lineages,
including the urothelium. Inoue et al. recently demonstrated
a method to generate human urothelial cells from dermal
fibroblasts by transducing genes for four transcription
factors, FOXA1, TP63, MYCL, and KLF4 (FTLK) (Inoue
et al., 2019). Innovative gene engineering resulted in the
creation of a cell population expressing urothelium-specific
markers and capable of forming an impermeable barrier.
These cells were also evaluated in vivo after being
transplanted into the bladder of mice with Interstitial
Cystitis (IC), and histological analysis revealed a significant
improvement in the quality of the urothelial layer.
TABLE 6 Application of biomaterials in the reconstruction of the ureter.
Study Biomaterial Biomaterial fabrication
method
In vitro/In
vivo
study
Model Outcome
Smith et al.,
(2002)
Porcine SIS A porcine SIS allograft was performed in vivo pig The SIS transplant caused the regrowth of
the ureters
Xu et al.,
(2012)
PLLA The PLLA was dissolved in chloroform,
cast into .8 mm-thick polymer films, and
then evaporated at 25°C–28°C with a 20%
relative humidity
in vivo rat The scaffold created allowed for the
proper proliferation of cells and the
creation of vascular networks
Shi et al.,
(2012)
PLA and collagen Chloroform was used to dissolve PLA to
create polymer films. The films were
spiral-wrapped around a glass rod. The
scaffold’s exterior surface was then
covered with a layer of mesh made either
entirely of PLA or PLA and collagen
in vitro,
in vivo
mouse Human adipose-derived stem cells
(hADSCs) have been differentiated into a
urothelial lineage by alternating the
microenvironment with urothelial cells.
The scaffold is compatible with cell
survival and proliferation
Koch et al.,
(2015)
Extracellular matrix crosslinked with
carbodiimide (CDI), genipin (GP),
glutaraldehyde (GA), or
glutaraldehyde (BP)
The decellularized ureters were then
crosslinked with different agents, such as
GP, CDI, GA, and BP.
in vitro,
in vivo
rat Carbodiimide crosslinked scaffolds
showed multilayer formation of smooth
muscle cells
Zhang
et al.,
(2012)
Silastic tubes Six female beagles had silicone tubes
inserted into their peritoneal cavities.
The tubes were removed after three
weeks, and the tubular tissue that
covered them was gently elongated
in vivo canine A two-month follow-up showed that the
neo-ureter demonstrated normal ureteral
architecture. The multilayered
urothelium was surrounded by smooth
muscle bundles
Liao et al.,
(2013)
BAM The bladder of a rabbit was removed and
decellularized
in vivo rabbit At 8 and 16 weeks after implantation, the
scaffold was characterized by multilayer
urothelium and organized bundles of
smooth muscles
Zhao et al.,
(2019)
Vessel extracellular matrix (VECM) Seeded VECM was tubularized and
wrapped by two layers of a rabbit
omentum for vascularization
in vivo rabbit Histological evaluation showed a layered
structure of the ureter with a multilayered
urothelium over the organized, smooth
muscle tissue
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The discovery of iPSCs opens new perspectives for
personalized regenerative medicine, as these cells may act as
an unlimited source of autologous engineered tissue. iPSCs are a
type of pluripotent stem cell that can be generated directly from a
somatic cell. iPSCs have generated a lot of interest in tissue
engineering because they can multiply indefinitely and also
transform into all types of cells in the body. Suzuki et al.
developed a protocol for the directed differentiation of hiPSCs
into stratified bladder urothelium through the definitive
posterior endoderm and caudal hindgut by recapitulating
embryogenesis by using high inhibitory doses for the enzyme
GSK-3 (Suzuki et al., 2019). This method was the first to obtain
terminally differentiated UCs expressing uroplakins from iPSCs.
4.2 Regeneration of the smooth muscle
layer
The development of technology for constructing tissue-
engineered muscle layers is required for the proper function
of the reconstructed urethra, urinary bladder, and ureter. These
hollow organs have active and passive diameter tension that is
controlled by a smooth muscle layer tone (Ajalloueian et al.,
2018). Smooth muscle from the bladder and ureter displays
patterns of spontaneous contractile activity determining the
storage and voiding phases of the micturition cycle (Brading,
2006). Interestingly, even the urethral muscle layer is involved in
humans in the generation of voiding pressure. Detrusor smooth
muscle cells experience a sevenfold length change and
continually preserve the ability to contract and generate urine
outflow over this broad length range (Tuna et al., 2012). The
anisotropy and heterogeneity of the mechanical characterization
of the human urinary bladder are related to its complicated
geometry, structure, and functions. Unfortunately, tissue-
engineered smooth muscle layers with a similar function have
yet to be replicated. Table 2 shows the achievements in
regeneration of the smooth muscle layer.
The restoration of the smooth muscle layer for tissue
engineering purposes might be accomplished by induced
regeneration combined with the adaptive cytoskeletal plasticity
of spontaneously regenerating cells. This mechanism is dominant
in cases where part of the native urinary tract wall acts as a
regeneration primer and is a source of muscle precursor cells (Liu
et al., 2019). Nevertheless, the self-regenerating capability of the
smooth muscle layer is minimal, therefore boosting endogenous
regenerative potency is necessary (Drewa et al., 2008).
MSC transplantations were documented to stimulate the
regeneration of tissue-engineered urinary bladders by
activating the hedgehog signaling pathway. Pokrywczynska
et al. (2019) showed, using a rat model, that MSC created an
environment rich in morphogenetic signals corresponding to an
early organogenesis environment when the mesenchyme forms
an early smooth muscle layer. In this scenario, MSC shaped the
paracrine framework to support the regrowth of the muscle layer,
guided by activated muscle cells sprouting from the entire region.
Adult smooth muscle cells retain the ability to form a
subpopulation of highly proliferative precursor cells. Yang
et al.formulated a similar conclusion using mesenchymal
USCs in a rabbit urethroplasty model to trigger regeneration
within the SIS graft. Part of the PKH67 labeled USCs were
capable of differentiation into cell lineages expressing
urothelial, smooth muscle, endothelial, and interstitial cell
markers, proving the multipotency of this cell population.
Application of USC led to the development of a low-grade
inflammatory response with the extensive rebuilding of the
muscle layer expressing myosin and actin (Liu et al., 2017).
The majority of indications for urinary tract reconstruction
include cancer, which eliminates the possibility of applying
autologous cells harvested from urinary tracts due to safety
concerns. For this purpose, Mirzaei et al. (2019) investigated
human iPSCs as a stem cell source for the generation of bladder
smooth muscle cells. They reported that in vitro cultivation of
iPSCs seeded on PLGA resulted in acquiring a smooth muscle
phenotype confirmed by the expression of alpha-smooth muscle
actin (ASMA), smooth muscle 22 alpha (SM- 22a), calponin-1,
caldesmin1, and myosin heavy chain (MHC). It was discussed
that the nanofibrous scaffold resembled native bladder ECM and
provided adhesive signaling that enhanced differentiation. The
characteristics of the attached phenotype might indicate
immature smooth muscle cells or myofibroblasts. These cells
might not accomplish the development of an adult SMC
phenotype that determines complex function and hierarchal
organization. The phenotypic changes during the smooth
muscle differentiation pathway have actin and myosin appear
early in development, whereas caldesmon and calponin serve as
markers for the final smooth muscle differentiation stages
(Huber and Badylak, 2012). Yipeng et al. (2017) underlined
that the acquisition of a smooth muscle phenotype depended
on a 3D scaffold and could not be replicated in a 2D culture
environment. 3D-structure and the geometry of the extracellular
matrix modulated SMC behavior and the development of the
contractile apparatus. These interactions, which are currently
being gradually identified and described, are particularly crucial
for the restoration of the linear arrangement of regenerated
smooth muscle layers within urinary tracts.
Priority is given to maintaining control over the arrangement
of SMCs cultivated on scaffolds for graft preparation by guiding
them using a configured biomaterial ultrastructure. The rationale
behind it is that an uncontrolled increase in ECM anisotropy
might initiate cancerogenic transformation (Walker et al., 2018).
Hoogenkamp et al. (2016) presented a method to obtain a
collagen scaffold in the shape of the whole bladder with
integrated anastomotic sites for the ureters and urethra. An
integrative approach for the restoration of ECM continuity
from the ureters to the urethra is an idea worth further
development as it should improve function recovery by
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reconstituting the consolidated middle layer of urinary tracts.
The fabricated scaffolds’ultrastructure had a uniform wall
thickness and a unidirectional pore structure to facilitate the
cell’s migration and attachment. The polarized cross-section of
the scaffold wall with uniform wall thickness and a unidirectional
pore structure guided proliferating SMCs and allowed for the
restoration of layered architecture.
The smooth muscle layer regeneration in vivo is a time-
consuming process, while most of the regulating growth factors
and cytokines are rapidly localized and systemically eliminated
(Khosravi et al., 2007)(Lee et al., 2011). This is one of the reasons
why the initial satisfactory regenerating results diminish over
time after the initial procedure. Ardeshirylajimi et al. recently
demonstrated the scaffold’s ability to release TGF-βin a
controlled manner to direct MSC differentiation into smooth
muscle (Ardeshirylajimi et al., 2018). Additionally, the 3D
network of electrospun nanofibers enabled MSC parallel
alignment, which successfully enhanced the formation of
muscle bundles. A novel biomimetic scaffold provided
multidirectional stimulation of the cellular component to
induce the desired differentiation pathway. The sparingly
discussed issue is the influence of the short shelf-life of most
of the growth factors and cytokines used for the induced
regeneration of SMCs on the eventual market optimization of
bioactive biomaterial scaffolds (Suarato et al., 2018). Currently,
fabrication of a ready-to-use scaffold, e.g., a bladder patch
enriched with growth factors, would be challenging due to
difficulties in the development of a reliable method for
bioactive component preservation.
4.3 Regeneration of bladder innervation
It has been proven that tissue engineering methods enable the
regeneration of the urothelial epithelium, smooth muscle, and
blood vessels in the reconstructed bladder. The problem remains
to recreate the neural network as shown in Table 3. The proper
functioning of the bladder is based on the cooperation of all
layers of the bladder wall. This allows the holding and passing of
urine to be regulated.
To achieve this, it is important to innervate the tissue-
engineered bladder. The process consists of the following
steps: axonal outgrowth, neural survival, branching, and target
nerve reconnection (Sharma and Bikramjit, 2022). Bladder
reinnervation is a complex process, therefore many studies are
in the initial stages of research.
Madduri et al. (2009) showed that the combination of GDNF
and NGF has a positive effect on the recovery of injured
peripheral nerves. GDNF significantly influences axonal
elongation, and NGF induces extensive axonal branching.
Kikuno et al. (2009) however, showed that VEGF combined
with NGF allows for the formation of aggregated bundles of
smooth muscles and the regeneration of nerves and fibers.
An alternative solution is to use cell-based therapies. Nitta
et al. used skeletal muscle-derived multipotent stem cells, which
after transplantation differentiated into Schwann cells,
perineurial cells, vascular smooth muscle cells, pericytes, and
fibroblasts around the bladder. The applied method allowed for
the recovery of 80% of the bladder’s functionality (Nitta et al.,
2010). Another approach is to use isolated Schwann cells to
innervate the tissue-engineered bladder. Adamowicz et al. (2011)
developed a protocol for the effective isolation of Schwann cells
from pre-degenerating peripheral nerves, which can deliver the
required amount of cells for transplantation into a urinary
bladder graft.
4.4 General concerns related to cell-
based tissue engineering
The clinical application of cell-based therapies in tissue
engineering is indispensable, but there is also a significant
question that is hardly ever addressed. First of all, oncological
safety of potential therapy utilizing differentiated in vitro cells.
Every cell division has a small chance of introducing deleterious
mutations, and systemic mechanisms such as immune
recognition aimed to correct these alternations do not
function in in vitro culture (Närvä et al., 2010). Some reports
indicated that the tumorigenicity of stem cells had been predicted
to increase proportionally with the length of in vitro culturing
(Knoepfler, 2009).
In most cases, the final recommendation of the experimental
method to be applicable for differentiation into SMCs or UCs was
based on the successful identification of several essential markers.
It must be admitted that knowing the complexity of urothelium
or detrusor muscle function and cytoarchitecture, this reasoning
is an oversimplification. In fact, these cells might be considered a
rather immature cell population with an unstable phenotype.
Detected markers might only be a manifestation of uncontrolled
or partial activation of the differentiation pathway, and their
presence does not determine function like healthy bladder SMCs
or UCs.
Moreover, the incomplete differentiation process supported
by indiscriminately activated pathways might become a platform
for malignant transformation. Unfortunately, we are also doing
little to advance our understanding of cell differentiation.
Current research accentuates that the differentiation fate of
UCs or SMC precursors is dependent on paracrine
stimulation and biomechanical signaling derived from the
scaffold structure. The next step is going to be genetic
engineering aimed at triggering the desired phenotype. Zhao
et al. obtained SMC expressing a mature contractile phenotype
from USCs using both miR-199a-5p and TGF-β1(Zhao et al.,
2019). The results of the study showed that SMCs converted by
miR-199a-5p and TGF-β1 had significantly better contractile
activity than TGF-β1 alone.
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Another problem is the survivability of transplanted cells
within the regenerating environment. There is a rising number of
reports indicating that transplanted cells, either alone or as part
of tissue-engineered grafts combining biomaterials, are
eliminated mainly by macrophages within several weeks
(Arutyunyan et al., 2015)(Yang et al., 2008). In this situation,
implanted cells, i.e., MSC, act as a temporary booster, modifying
the regenerative environment but without long-lasting effects
(Pokrywczynska et al., 2018). Considering this, the ability to
rebuild neo-tissue that is integrated with host urinary tracts by
implanting mature cells is to be questioned. Alternatively,
according to some authors, the putative regenerating effect of
transplanted stem cells depends on infiltrating immune cells
(Abnave and Ghigo, 2019).
5 Biomaterials
The extracellular matrix of the urinary tract wall is a three-
dimensional network composed of multi-domain
macromolecules such as collagen, elastin, glycosaminoglycans
(GAGs), and cell-binding glycoprotein (Binette and Binette,
1991). Successful tissue-engineered-based replacement of the
urethra, urinary bladder, or ureter requires the development
and fabrication of a biomaterial scaffold acting as a
supporting frame for growing tissue.
5.1 Urethra
Acellular biomaterial scaffolds are desired for implantation
as they do not require the expensive and time-consuming process
of cell seeding. From the perspective of commercialization of
tissue engineering therapies, the acellular approach is more
accessible due to the convenience of use in urology
departments that are not equipped with the infrastructure
necessary for cell culturing. It is generally believed that cell-
free biomaterials are suitable for small defects only, but to our
knowledge, there is no study comparing cell-seeded and acellular
scaffolds for urethroplasty outside of a control group
(Versteegden et al., 2017).
In the pioneering study of Dorin et al. (2008) the authors
used acellular bladder submucosa for tabularized urethroplasty at
varying lengths in a rabbit model. They reported breakthrough
observations for further research focusing on urinary tract wall
replacement, including the urethra. Bridging grafts showed
ingrowth and healthy regeneration of the urethral wall only at
the anastomotic edges. At the same time, increased collagen
deposition and fibrosis toward the center occurred. Orabi et al.
(2013) demonstrated in a preclinical study evaluating tabularized
collagen scaffolds for extended urethral defects using a canine
model, that cell seeding is necessary to counteract the fibrotic
reaction and stricture formation.
These observations led to the evolution of regenerating
biomaterials into the design of bioactive matrices.
Accordingly, Jia et al. (2015) applied collagen membranes
linked with VEGF for the repair of 5 cm-long anterior urethra
defects using a canine model. The study’s concept concentrated
on the potential ability of VEGF to improve neo-angiogenesis
and related blood supply within the implanted biomaterial. The
authors concluded that collagen scaffolds enriched with VEGF
promoted urethral tissue regeneration and improved the
function of the neo-urethra. Non-etheless, the attached
histological data documenting the regrowth of the urethral
wall demonstrated a highly disordered network of neo-vessels
and smooth muscle cell bundles that deviated significantly from
normal muscle architecture. It is very likely that after a follow-up
longer than six months, the stricture would be rebuilt due to the
gradual increase of local ischemia and a lack of spatial resistance
usually mediated by the resting tone of muscle layers.
Pinnagoda et al. (2016) delivered data from the longest
reported follow-up after partial completion of urethral
replacement with a tissue-engineered acellular graft. In this
study, the rabbit urethra was reconstructed with a double-
layer collagen scaffold expected to support regeneration and
simultaneously prevent the graft from collapsing under the
pressure generated by forming a scar. After nine months, the
histological analysis revealed a well-regenerated urethral wall
with stratified epithelium and an abundance of muscle cells. This
promising result should be approached cautiously because the
collagen scaffold was approximately 1.3 mm thick. In this case,
the regenerating area could be passively perfused by neighboring
tissue, preventing the development of ischemia. In general,
1mm–2 mm is a limit value for efficient oxygen exchange and
diffusion derived from the local blood supply (Moon and West,
2008). In terms of greater distances, the hypoxic zone promotes a
fibrotic response. For a clinical application, a collagen scaffold
approximately 1 mm thick does not provide efficient mechanical
endurance for urethral surgery; the estimated young modulus
was approximately seven kPa, which corresponds to human liver
tissue. On the other hand, the mechanical properties of scaffolds
created from liquid collagen are challenging to balance due to the
tendency during collagen solution dehydration to form rigid
structures with a high young modulus (Meyer, 2019).
Nowadays, biomaterial science still struggles to replicate the
high biocompatibility of naturally derived biomaterials and
implement them as a fabricated scaffold for tissue engineering.
AMA is a promising biomaterial with an abundance of unique
properties (low immunogenicity, promotion of epithelization,
anti-inflammatory properties, angiogenic and antiangiogenic
properties, antifibrotic properties, antimicrobial properties,
and anticancer properties) (Adamowicz et al., 2019b). Despite
excellent application potential, it remains a relatively unknown
biomaterial for the urological community.
Ophthalmologists, on the other hand, routinely apply AM as
a biological wound dressing for the treatment of corneal injury
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(Dua et al., 2004). AM grafts were utilized for urethral wall
replacement in several studies using small animal models
(Shakeri et al., 2009)(Güneşet al., 2017). All of them
underline excellent AM properties and the ability to induce
local regeneration. Nevertheless, the research data derived
from these reports has low translational potential because of
insufficient research group numbers, essential diagnostic tools,
and the heterogenic model of urethral injury. Admittedly, two
short reports show the feasibility of using AM for urethroplasty
in humans (Koziak et al., 2007)(Koziak et al., 2004). Although
these papers presented a novel method, the leading concept of the
studies (conducted on a few patients) was instead to demonstrate
extravagance surgery rather than to change the current
management dependent on buccal mucosa.
An electrospinning technique provided the ability to produce
biomaterials with nanoscale properties for tissue engineering.
Given the importance of intercellular interactions between the
biomaterial and the ingrowing tissue, electrospinning allows for
the fabrication of 3D scaffolds arranged in a complex fibrous
porous matrix similar to a healthy ECM (Davis et al., 2018). It is
especially important for the regeneration of multilayer
hierarchically organized structures, such as the urinary tract
wall, as this parallel spatial architecture allows the
regenerating tissue to maintain its orientation. Moreover,
electrospinning allowed for the creation of small-diameter
tubes with high uniaxial mechanical resistance appropriate for
urethral surgery, as demonstrated by Sánchez-Pech et al. (2019)
The significant advantage of their work is that it includes
extensive biomechanical analysis, which is usually only
provided marginally. Composite tubular scaffolds created from
PCL and PLGA exhibited a high elastic modulus (19 MPa), ideal
to withstand bursting pressure within the human urethra.
Additionally, a low strain-to-break value should guarantee a
proper surface for stable anchoring and fixing sutures, which
would be necessary to convince a surgeon to use them.
Notwithstanding, the optimal biomechanical features of PCL-
based electrospun scaffolds have a high hydrophobicity and
hence create an environment that inhibits cell attachment and
growth. This is a seldom-reported problem that needs to be
addressed by modification of the scaffold’s surface. Alternatively,
the high adaptability of electrospinning technology makes it
possible to utilize electrospun nanofibers as a skeleton for a
naturally derived biomaterial with low mechanical resistance, for
instance, AM (Adamowicz et al., 2016).
To overcome the mentioned problem, the application of
several low-cost modes such as centrifugal jet spinning and
immersive rotary jet spinning combined with hydrogels might
solve the problem of an environment that impedes cell
attachment and growth (Ravishankar et al., 2021), (Gonzalez
et al., 2017).
Khang et al. (2017) report a procedure that allows for
engineering biphasic Janus-type polymeric nanofiber networks
via the centrifugal jet spinning technique that provides unique
structural support and biological activity and has many
applications in tissue engineering approaches, such as where
there might be a need for different properties on either side of the
scaffold, such as environment resistance on one side and
biocompatibility or potential therapeutic properties on the
other side.
Sharma et al. (2022) used Contact-Active Layer-by-Layer
Grafted TPU/PDMS and were able to validate TPU/PDMS
blends as an antiencrustation and antibacterial platform for
next-generation urological biomaterials with physiological
relevancy for functionality. These layer-by-layer grafted blends
displayed significant grafting stability and antibacterial efficacy
against common uropathogens. The application of biomaterials
in tissue engineering in urethral reconstruction has been
described in Table 4.
5.2 Urinary bladder
The urinary bladder’s proper function is dependent on its
ability to continually repeat loading and unloading cycles
corresponding to urine storage and voiding (Yamanishi et al.,
2011). This behavior model needs biomaterial for tissue
engineering purposes that can withstand dynamic pressure
changes during frequent mechanical loading and unloading
(Adamowicz et al., 2017b). The bulk of studies introducing
new biomaterials for urinary bladder replacement was
published several years ago. Nowadays, sporadically appearing
in articles covering this field, the topic has a repetitive character.
In the field of urinary bladder experimental reconstruction,
the polymer materials PLA, PGA, and PLGA were the most
influential contributors to the creation of biodegradable cellular
scaffolds (Serrano-Aroca et al., 2018). All these biomaterials have
FDA approval for clinical usage, and their degradation rates can
be changed based on their molecular weights and compositions
(FDA’s Regulatory Science Program for Generic PLA/PLGA-
Based Drug Products, 2016).
Non-etheless, the major disadvantage of biodegradable
copolymers such as PLA, PLGA, and PGA is evident rigidity
and non-linear elasticity in comparison to the native bladder wall
(Ajalloueian et al., 2018). When using these biomaterials, micro-
environmental mechanical stress may affect regenerating tissue,
inducing scarring by activating stretch-induced activation of
TGF-β1(Yong et al., 2015). Baker et al. (2009) showed a
correlation between biomaterials’Young’s modulus and a
scaffold’s ability to support urothelial layer formation in vitro.
Horst et al. (2017) proposed to enrich a PLGA scaffold with
polyester urethane to improve elasticity and thus adjust it to a
highly compliant urinary bladder wall. Indeed, the obtained
biocomposite exhibited extraordinary passive elasticity, but
careful analysis of cystograms demonstrated a stepped
increase in intravesical pressure. This indicated the dominance
of mechanical properties related to PLGA that were particularly
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visible eight weeks after bladder reconstruction. It was likely a
consequence of the unstable mechanical properties of
electrospun polyester urethane during uncontrolled
degradation in the urinary tract environment. In order to
design scaffolds for urinary bladder wall replacement, the
degradation period needs to be adjusted to the time-lapse of
tissue regeneration to prevent structural failure of the implant
and graft rupture. Research data concerning changes in
mechanical parameters and structural integrity during the
degradation of electrospun scaffolds for soft tissue
regeneration is minimal.
Besides difficulties in the adaptation of mechanical
parameters, the tissue-engineered-based reconstruction of the
urinary bladder demands the fabrication of biomaterial scaffolds
in sizes applicable for human use. In most of the studies, authors
introduce different cell matrices utilizing small animal models
using biomaterial samples without a standardized or repeatable
fabrication protocol (Pokrywczynska et al., 2014). We can take a
chance and say that in most cases, biomaterials tested on small
animal models would be very difficult or even impossible to
manufacture due to the size needed for translational animal or
human trials. It is a reason why scaffolds made from
decellularized extracellular matrices became popular in the
field of bladder tissue engineering (Pokrywczynska et al., 2015).
BAM is derived from the bladder after a decellularization
procedure conducted according to established protocols that
enable the preservation of the native ECM architecture.
Hence, BAM is composed of a complex collagen network
enriched with fibronectin, elastin, and plenty of GAGs and
growth factors (Song et al., 2013). It should be emphasized
that a particular arrangement of fibril proteins presented
within BAM cannot be artificially replicated by current
technology. Also, BAM retains an intact basement membrane,
supporting rapid re-epithelialization. The rationale for BAM
matrix fabrication was the assumption that its bioactivity
profile would enhance regeneration by stimulating and
guiding growing cells naturally, restoring the urothelial and
detrusor muscle layers. The recent study of Pokrywczynska
et al. (2018) proved that BAM undoubtedly created a
favorable regenerative environment, but it also exhibited the
same limitations as all known biomaterials tested for bladder
augmentation so far. First of all, the active regeneration evaluated
as the restoration of a healthy layered bladder structure was
observed only within BAM regions firmly attached to the native
bladder wall. The closer to the center of the graft, the more
abundant the scar tissue was and, analogously, the decrease in the
density of neovessels. Similar to available reports, regeneration of
the bladder wall did not occur evenly. Instead, a reconstructed
part of the bladder could be divided into three regions. The part
of the graft that borders native tissue with well-regrown
urothelial and smooth muscles. The transient part where the
regeneration quality is heterogenic, due to the gradually
increasing content of fibrotic tissue disrupting the newly
forming urothelial and muscle layers. Finally, the center of the
graft is overgrown with thick stellar scars responsible for an
average graft shrinkage of 50%. As the mechanism of scar
development is strictly linked to an insufficient blood supply
and related local ischemia, the counteraction might be inducing
angiogenesis. For instance, Jiang et al. (2016) fabricated BAM
modified with VEGF-loaded PLGA NP. The study was
conducted using a rabbit model. The VEGF quantification
results demonstrated that the modified BAM achieved long-
term sustained VEGF release in vivo. The contractile rate of
the acellular matrix in the experimental group was significantly
lower than in the control group. The functional evaluation of
isolated stripes in vitro revealed that bladders reconstructed with
VEGF-supplemented grafts were more responsive and exhibited
the ability to undergo rapid cyclic contraction and relaxation.
Incorporating additional substances into biomaterials such as
BAM is a common strategy used to obtain a specific effect and
control the regeneration process.
On the other hand, we still do not have enough knowledge to
modulate such a complicated process intentionally. Zhou et al.
(2013) announced that the co-administration of PDGF and
VEGF with BAM significantly improved muscle contractility
and angiogenesis. There was, however, no control group with
one of the growth factors to check its role autonomously. Before
the introduction of regular supplementation of the regenerative
environment with bioactive substances, there are several
questions to answer. Firstly, at what point during the healing
process does the particular growth factor act? Secondly, what
concentration is required? Thirdly, what is the exposure time to
obtain the appropriate effect?.
Interestingly, despite different published strategies to use
growth factors to supply the regenerative environment, no
research groups have discussed potential risks. For example,
VEGF, which is the most often used to improve regeneration
outcomes, is also involved in cancerogenesis (Takahashi and
Shibuya, 2005). VEGF is one of the potent factors inducing MET
in mesodermally derived neoplasms. The healing environment,
especially when artificially created within biomaterials, is
susceptible in the long term to cancerogenic transformation.
The presence of cells at various stages of differentiation and
chronic inflammation per se creates a favorable environment for
tumor transformation. If we additionally add potent bioactive
substances without a deep background understanding of their
action, we may trigger cancerogenic transformation.
Biomaterials of natural origin, such as the aforementioned
BAM, are often used in tissue engineering due to their very good
mimicry of the natural environment for cell growth, providing
adhesive substances, cell binding sites, and compatibility with the
tissues surrounding the regenerated organ or tissue (Dalamagkas
et al., 2016).
Aside from their many benefits, biomaterials have some
drawbacks, such as the limited ability to modify them and the
heterogeneity of the scaffold structure in terms of chemical purity
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(Schmidt and Leach, 2003). In addition, most natural polymers
used in tissue engineering have limited and batch-dependent
mechanical properties, which often make it impossible to
unequivocally assess the effectiveness of the method used.
Synthetic biomaterials can also be distinguished, the great
advantage of which is the possibility of modifying their structure
during the production stage. The second most important thing is
the production of identical scaffolding on a large scale, which will
make it possible to reproduce the same results (Dalamagkas et al.,
2016).
Sivaraman et al. (2017) highlighted the advantages and
disadvantages of natural and artificial biomaterials using the
example of a hydrogel, and how their combination can affect
the properties of the hydrogel itself. They used composite
hydrogel of Tetronic 1107-acrylate with ECM moieties like
collagen and hyaluronic acid seeded with bladder smooth
muscle cells, which provided a viable environment for bladder
smooth muscle cells to survive and reconstruct the scaffold. In
comparison with the control, acellular hydrogel, the mechanical
properties, stiffness, and strength of the cellular composite
hydrogels were significantly greater. The team reports that
culturing the construct for longer periods after bladder
smooth muscle cell seeding and adding growth factors to the
composite hydrogel system would further aid in accurately
mimicking the mechanical properties of the bladder wall.
Furthermore, Sivaraman et al. provide evidence that Poly
(Carbonate-Urethane)-Urea scaffolds possess high porosity
and cytocompatibility, indicating their ability to support
bladder cell growth and the formation of viable tissue. The
researchers also demonstrated that the Poly (Carbonate-
Urethane)-Urea scaffolds are suitable for the bladder’s
primary functions of maintaining low pressure during storage
and withstanding high pressure during voiding (Sharma et al.,
2021).
Promising synthetic biomaterials also include graphene and
graphene-based nanomaterials, which, due to their structure,
have many unique properties and are especially useful in the
innervation of organs reconstructed using tissue engineering
techniques. The mechanical strength of most polymers is
characterized by declining strength when used under
physiological conditions. This unique, two-dimensional
material, thanks to its organized structure, is characterized by
high strength and at the same time is stretchable and elastic
(Ławkowska et al., 2022).
Additionally, as reported in the literature, nanomaterials
based on graphene can improve Young’s modulus and
increase the compressive and tensile strength of the scaffold
itself (Tiwari et al., 2020), (Cheng et al., 2018).
The most breakthrough application of graphene in
reconstructive urology, however, is its use for the replacement
of the neuronal network of the tissue-engineered urinary bladder.
The folded structure and adapted porosity of the biomaterial
not only provide mechanical support for cell proliferation but
also replace the extracellular matrix, inducing the appropriate
growth and elongation of axons (Adamowicz et al., 2020)
(Ławkowska et al., 2022).
A major problem with the reconstruction of the urinary
bladder is the occurrence of inflammation. Here, an additional
advantage of graphene is its antibacterial effect, which may
additionally support the process of reconstruction of tissues
and organs of the urinary system (Liu et al., 2011).
However, additional studies should be carried out to
unequivocally assess the cytotoxicity of graphene-based
nanomaterials and to standardize graphene production
methods so that the highest quality graphene can be used in
the research. Biomateirals of natural and synthethic origin in the
reconstruction of the urinary bladder have been mentioned in
Table 5.
5.3 Ureter
Various types of biomaterials were introduced in ureteral
reconstruction as shown in Table 6. Most studies focused on
acellular, tabularized SIS grafts using a porcine model (Liatsikos
et al., 2001)(Smith et al., 2002). The translational potential of
these reports was low due to the different methodologies used
and the short follow-ups. Studies conducted using a porcine
model also did not address clinical needs as model ureter
injuries could be effectively treated with available surgical
techniques. The primary aim for tissue engineering research
should be to develop a graft suitable for ultimately bridging the
long ureter gap. In addition, there is a need to reconstitute
peristaltic motion, to prevent incrustation, and provide long-
term patency of the reconstructed hollow region (Adamowicz
et al., 2019a).
Several studies evaluated tissue-engineered grafts made from
PLLA on small rodent models. The quality of this report was low
and must be critically classified due to the low number of animal
groups, lack of controls, and several weeks of follow-up (Xu et al.,
2012)(Shi et al., 2012).
The biocompatibility of the introduced biomaterials was not
enough to prevent a fibrotic reaction and stricture formation. To
overcome these problems, Koch et al. developed a methodology
for manufacturing decellularized ureters obtained from pigs
(Koch et al., 2015). Proper decellularization techniques enable
the use of xenogeneic ECM grafts without the risk of acute
rejection. The decellularized ureter was described as a thin,
flexible biomaterial without resistance to collapse. This is the
reason why the authors applied different crosslinking agents
(carbodiimide, glutaraldehyde, and genipin) to improve its
mechanical characteristics. Carbodiimide-crosslinked scaffolds
increased infiltrating 3T3-cells and smooth muscle cells’
multilayer formation. Nevertheless, the prepared grafts were
not evaluated in vivo. Evaluation of biomaterial
biocompatibility by implanting biomaterials into rat
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subcutaneous tissue is a commonly used method to analyze
immune reactivity and recellularization capacity. This strategy
has many pitfalls that are not considered during the
interpretation of results. The striking differences are in the
healing response because rat skin and subcutaneous tissue
have little in common with their human counterparts
(Dorsett-Martin, 2004). In rats, loose-skinned healing occurs
by wound contraction, which is the primary mechanism of
wound healing instead of scarring followed by epithelization
in humans. Additionally, rats can convert L-glucono-gamma-
lactone to vitamin C within subcutaneous tissue, which acts as a
powerful antioxidant and scar-prevention factor (Weber et al.,
2019). As a result, implanted tested biomaterials appeared not to
be susceptible to scarring, but in fact, they would trigger a
different response in humans. Sprouting and elongation of a
new vessel within the scaffold are dependent on a three-
dimensional biomaterial architecture. Highly porous
biomaterials provide a better surface to support the formation
of a branched vascular network. In terms of ureter
reconstruction, a scaffold’s ability to sustain angiogenesis
should be considered the most decisive parameter, critical
for a successful outcome. The ureter’s blood supply is
primarily composed of a highly dichotomized vascular
network that penetrates the ureter’s wall superficially. In
this situation, the scaffold must have a microporous
structure ready for the ingrowth of these small caliber
vessels. In the case of ureter tissue-engineered-based
reconstruction, the strategy of utilizing pre-implantation
before repairing a ureteral defect was tested. Zhang et al.
(2012) exploited pre-implantation to create a tubular
scaffold from the fibrous capsule, which was formed in the
peritoneal cavity. Liao et al. (2013) used omental pre-
implantation as an in vivo bioreactor to increase
neovascularization in the construct.
Although preimplantation is a tempting idea because it
theoretically enables the creation of a functional vascular
network. This method was only tested in animal settings.
Zhao et al. (2019) recently demonstrated the most
sophisticated approach for ureteral reconstruction so far. They
used VECM, which is naturally rich in VEGF, for rabbit ureter
reconstruction. The tubular graft underwent three weeks of
maturation within the omentum to develop a branched
vascular network. At two months post-ureter reconstruction,
the histological evaluation showed a layered structure of the
ureter with a multilayered urothelium over the organized,
smooth muscle tissue.
In the case of the ureter, local hypoxia associated with
insufficient angiogenesis would be revealed after a shorter
period when compared to the urethra and the bladder. A
narrow ureteric lumen and a tendency to collapse due to
higher constant higher abdominal pressure make tissue-
engineered ureter particularly prone to scarring. It is one of
the reasons why no clinical trial has been conducted yet.
Conclusion
It is undeniable that reconstructive urology needs advanced
therapeutic modalities utilizing recent biotechnological advances to
improve current treatment effectiveness. Tissue engineering
focusing on manipulation with cells and biomaterials ideally fits
the modern reconstructive urology development pathway.
Nevertheless, due to inefficient translational research, a
misunderstanding of clinical needs, a tendency toward overly
enthusiastic result interpretation, and sometimes a questioning
attitude among clinicians toward tissue engineering applied
outside the experimental field, this method is marginally used in
urology. Despite decades of research, tissue engineering is still in the
early stages of revolutionizing urology. Only a forward-looking
approach to tissue engineering research and reliable study reports
with repeatable outcomes will accelerate this process.
Members of the Trauma and
Reconstructive Urology Working
Party of the European Association of
Urology Young Academic Urologists
Jan Adamowicz, Enrique Fes Ascanio, Andrea Cocci, Mikołaj
Frankiewicz, Campos Juanatey, Guglielmo Mantica, Clemens M.
Rosenbaum, Wesley Verla, Malte W. Vetterlein, Marjan Waterloos
Author contributions
Conceptualization, KŁand JA; formal analysis, KŁ;
writing—original draft preparation, KŁand JA;
writing—review and editing, CR, PP, KK, LK, TD, JA, and
MP; visualization, KŁ; supervision, JA; project administration,
JA; funding acquisition, JA All authors have read and agreed to
the published version of the manuscript.
Conflict of interest
The authors declare that the research was conducted in the
absence of any commercial or financial relationships that could
be construed as a potential conflict of interest.
Publisher’s note
All claims expressed in this article are solely those of the
authors and do not necessarily represent those of their affiliated
organizations, or those of the publisher, the editors and the
reviewers. Any product that may be evaluated in this article, or
claim that may be made by its manufacturer, is not guaranteed or
endorsed by the publisher.
Frontiers in Bioengineering and Biotechnology frontiersin.org18
Ławkowska et al. 10.3389/fbioe.2022.1040987

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Glossary
AM Amniotic membrane
ASMA Alpha-smooth muscle actin
ATPs Advanced Therapy Products
BAM Bladder acellular matrix
BMSCs Bone marrow mesenchymal stem cells
BP Glutaraldehyde
CBD Collagen binding domain
CBER Center for Biologics Evaluation and Research
CBMPs Cell-based Medicinal Products
CDI Carbodiimide
DRG Dorsal root ganglion
ECM Extracellular Matrix
EMA European Medicines Agency
FDA Food and Drug Administration
GAGs Glycosaminoglycans
GDNF Glial cell line-derived neurotrophic factor
GMP Good Manufacturing Practice
GP Genipin
hiPSCs Pluripotent stem cells
IC Interstitial Cystitis
iPSCs Induced pluripotent stem cells
MHC Myosin heavy chain
MSC Mesenchymal stem cells
NGF Nerve growth factor
NIH National Institutes of Health
PGA Poly(glycolic acid)
PLA Polylactic acid
PLGA Poly(lactic-co-glycolic acid)
PLLA Spiral poly (L-lactic acid
SIS Small intestinal submucosa
SM- 22a Smooth muscle 22 alpha
SMC Smooth muscle cells
SUI Stress urinary incontinence
TEMPs Tissue Engineered Medical Products
UCs Urothelial cells
USC Urine-derived stem cells
VECM Vessel extracellular matrix
VEGF Vascular endothelial growth factor
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