Tissue Engineering and Stem Cell Therapy in Pediatric Urology.

The rapidly expanding field of tissue engineering along with stem cell therapy has a promising future in pediatric urological conditions. The initial struggle seemed difficult in renal regeneration but a functional biounit has been developed. Urine excretion has been demonstrated successfully from stem cell-generated embryonic kidneys. Three-dimensional (3D) stem cell-derived organoids are the new paradigm in research. Techniques to regenerate bladder tissue have reached the clinic, and the urethra is close behind. 3D bioprinted urethras would soon be available. Artificial germ cells produced from mouse pluripotent stem cells have been shown to give rise to live progeny. Myoblast and fibroblast therapy has been safely and effectively used for urinary incontinence. Stress urinary incontinence has been clinically treated with muscle-derived stem cells. Skeletal muscle-derived stem cells have been shown to get converted into smooth muscle cells when implanted into the corpora cavernosa in animal models. This review encompasses the various experimental and clinical developments in this field that can benefit pediatric urological conditions with the contemporary developments in the field. Copyright:

INTRODUCTION

Scientific and technological advances have been growing at a fast pace in the field of stem cell biology and tissue engineering research. Ever since the fact was acknowledged that stem cells can be used to cure a number of diseases, the scientists, basic and clinical researchers have been leaving no stone unturned to find the possible methods and indications where this technology can be used for the benefit of mankind. Stem cells are undifferentiated, unspecialized cells with undetermined function which under the influence of a definite signal can divide and become a specialized cell with a defined function. There are various conditions in pediatric urology where the surgeon feels that the tissue is inadequate due to congenital absence, disease or injury and hopes for some bioengineered tissues to fill in the gaps. Stem cells with potential to transform into healthy cells and repair damaged cells may prove beneficial in various congenital malformations.[1]

In urology, the limitations of tissue engineering include reduced proliferative capacity and low quality in vitro cultures of urothelial cells and the fact that tissue engineering cannot be used to treat malignancy. Allogenic tissue/organ replacement surgery has been associated with rejection or need for immunosuppression. Hence, autologous cells have been explored to avoid these issues. Accordingly, with recent advances, the need for tissue engineering and stem cell biology to go hand in hand is recognized where bioengineered tissue is generated using autologous stem cells for clinical applications. This review outlines the developmental and technological advances in this field that can benefit pediatric urological conditions and portrays the future scientific directions to improve the outcomes.

ORIGIN OF STEM CELLS

Mouse embryonic stem cells in laboratory were first discovered in 1981. Seventeen years later, in 1998, Human embryonic stem cells were discovered simultaneously from blastocyst by James Thomson at University of Wisconsin-Madison and from primordial germ cells by John Gearhart at Johns Hopkins University. There are four types of stem cell populations according to Hierarchy [Figure 1]:

Figure 1

Various types of Stem cells according to hierarchy

Totipotent: capable of forming a complete human being, e.g., zygote

Pluripotent not capable of a complete human being, but almost all the tissues of a human being e.g., germ cell

Multipotent capable of forming many cell types, for example, hematopoietic stem cells

Unipotent capable of forming a single cell type, for example, skin, nerve, progenitor cells.

SOURCES OF STEM CELLS

For a pediatric surgeon to understand the science and technology of tissue engineering at a molecular level, it is important to understand the various cell types and their sources. The commonly available sources of stem cells include

Embryonic stem cells: These can be derived from excess human embryos and aborted fetuses. The limitations include ethical issues, restricted resource, incompatibility, and risk of tumor generation

Adult stem cells: These cells are derived from tissues that develop from all three embryonic germ layers. For example, brain (particularly hippocampus), bone marrow, peripheral blood, blood vessels, skeletal muscle, epithelia of the skin and digestive system, cornea, tooth dental pulp, retina, and liver, pancreas. The sources of stem cells for utility in pediatric urology are hematopoietic, mesenchymal and spermatogonial stem cells

Umbilical cord blood stem cells. These are multipotent stem cells, similar to adult stem cells. The advantages include easy availability, immaturity, reduced rejection

Amniotic fluid stem cells. These were discovered recently in 2003. They are pluripotent. The advantages include the possible differentiation into all three germ layers with low immunogenicity and high anti-inflammatory action

Placental stem cells. These are multipotent adult stem cells. They possess the phenotypic plasticity of many cells with immunomodulatory properties

Induced pluripotent stem cells. These stem cells are derived from patient's tissue and induced into pluripotency. The most common and successful method of inducing pluripotency is through viral vectors, which questions the associated dangers in clinical treatments[2]

Urine-derived stem cells. These are derived from the parietal cells or podocytes within glomerulus in kidney and can be isolated from voided urine.[3]

CELL BASED OR REGENERATIVE THERAPY

Stem cell therapy may be either cell-based therapy that may be used to provide cells for reparative tissue such as the kidney or regenerative or reparative medicine that would be needed to replenish lost tissue like the ureter or the urethra. The authors have used the spare preputial skin in cases of circumcism and urethroplasty for research work.[45]

ROLE OF TISSUE ENGINEERING AND STEM CELLS IN UROLOGY

Pediatric urological conditions amenable to cell-based or regenerative therapy are depicted in Table 1.

Table 1

Conditions in Pediatric Urology amenable to cell based therapy

	Tissue	Condition
1.	Renal	Renal damage due to Pelviureteric junction Obstruction
		Renal Dysplasia
		Multicystic kidney Disease
		Vesicoureteric Reflux
		Bilateral Wilms Tumour post treatment
2	Ureter	Ureteral Atresia
		Ureteric Injury
3	Bladder	Exstrophy bladder
		Posterior urethral valve bladder
		Neurogenic bladder
		Myogenic bladder
		Small bladder
		Rhabdomyosarcoma bladder
		Radiation cystitis
4	Urethra	Hypospadias
		Epispadias
		Urethral Stricture
		Traumatic loss
5	Sphincter muscle tissue	Incontinence
		Exstrophy epispadias complex
6	Gonadal tissue	Disorders of sexual differentiation
		Undescended testis
7	Cavernosal tissue	Disorders of sexual differentiation
		Epispadias
		Traumatic loss
		Aphallia

The various tissues that may be derived using tissue engineering and stem cell therapy include [Figure 2]:

Figure 2

Urological tissues that may be derived using tissue engineering and stem cell therapy include (a) Renal tissue and urinary tract tissues like ureter, bladder, urethra (b) male genital tract such as testicular tissue, gametes, and cavernosal tissue, (c) female genital tract such as ovarian tissue and healthy endometrial tissue

Renal tissue

Urinary tract tissues: ureter, bladder, urethra

Sphincter muscle tissue

Gonadal tissue and Gametes

Cavernosal tissue.

RENAL TISSUE

The kidney is the most important organ in urology. The number of renal transplants is increasing every day and the dearth of donor organs continues in countries where the cadaveric donor program is low. Although a large of scientists are working on the role of stem cells in renal regeneration, the challenge is grave as there are numerous different types of cells that make up a functional nephron. The use of renal progenitor cells to generate complex renal structures has been attempted.[6] Adult bone marrow stem cells have been shown to differentiate into renal tubular epithelial cells and associated stromal cells.[7] The main aim of cell-based therapy with supplementary cells in the repair and regeneration of a damaged kidney to accelerate the natural healing process through cellular supplementation by rooting them in the natural healing process.[8] Renal cell repair and regeneration following acute renal failure follows a program of de-differentiation, migration and proliferation, and restoration of differentiated function. Accelerating and augmenting this process through cellular supplementation is beneficial.[9] Gupta et al. isolated a unique population of cells that behaved similar to a renal stem cell from rat kidneys that exhibited plasticity demonstrated by the ability of the cells to be induced to express endothelial, hepatocyte, and neural markers by reverse transcriptase-PCR and immunohistochemistry.[10] These cells could differentiate into renal tubules when injected under the capsule of an uninjured kidney or intra-arterially after renal ischemia-reperfusion injury. The authors have identified renal stem cells in a mouse model of Obstructive renopathy that can be targeted to preserve renal function.[11] The authors have clinically used bone marrow-derived mononuclear cells in a single case of bilateral multicystic kidney disease and demonstrated improvement in renal function parameters and formation of immature tubules though the baby succumbed at 17 months age.[12] Urine excretion has been demonstrated successfully from stem cell-generated embryonic kidneys.[13] Methods have been established to generate kidney organoids from human pluripotent stem cells. These organoids consist of cells with the characteristics of podocytes, proximal tubules, loops of Henle, and distal convoluted tubules in a contiguous arrangement resembling nephrons in vivo as well as interstitial cells.

URINARY TRACT TISSUES

The urinary tract tissues are mainly composed of two cell types, i.e., epithelial and mesenchymal. It is a great challenge to obtain both differentiated smooth muscle and urothelial cells from stem cells. Ureteral grafts have been created in experimental models. Tissue-engineered tubular grafts have been constructed by seeding bone marrow mesenchymal stem cells and smooth muscle cells into a bladder acellular matrix for ureteral reconstruction.[14] There has been a lot of ongoing research in tissue engineering for bladder reconstruction.[315] Bladder reconstruction with tissue engineering technology is possible through the use of normal autologous bladder cells seeded on biodegradable scaffolds.[16] The various types of stem cells used in preclinical animal models to repair or regenerate bladder tissue include pluripotent stem cells such as embryonic stem cells, induced pluripotent stem cells, multi-potent mesenchymal stem cells, bone marrow-derived mesenchymal stromal cells, adipose-derived stem cells, hair follicle stem cells, umbilical MSCs, urothelial stem cells and most recently, urine-derived stem cells employing either trans-differentiation, or paracrine effects to stimulate endogenous cells participating in tissue.[17181920212223242526] The authors have used autologous bone marrow-derived mononuclear cells in cases of meningomyelocele and found improvement in the neurogenic bladders.[1] The authors had layered autologous bone marrow aspirate above the bladder closure during a redo repair of Exstrophy bladder. The child attained urinary continence that is persistent at 7 years’ follow-up.

The urethra has been the most studied organ in tissue engineering and stem cell research. Atala was the first one to use tissue-engineered urethra[27] Tissue-engineered grafts comprising of epithelial cells seeded on scaffolds have been explored for urethroplasty.[28] Epithelial cells from urinary bladder, urethra, and buccal mucosa have been used in tissue-engineered grafts for urethroplasty and urethral reconstructive surgeries. Raya-Rivera et al. created tissue-engineered autologous tubularized urethras in five boys with urethral defects.[29] They isolated stem cells from bladder tissue, seeded them onto a synthetic tubular meshwork and created tissue-engineered tubularized urethras that were then used for urethral reconstruction. They reported a 100% success over a median follow-up of 71 months.[29] Unfortunately, these results have not been replicated by others. Attempts to produce tissue-engineered oral mucosa as a substitute for the patient's natural buccal mucosa have yielded disappointing results.[30]

Treatment of urethral stricture using tissue-engineered epithelial cell graft has evoked keen interest among urologists because of the ample availability of tissue-engineered graft and minimal donor site morbidity.[31] In vitro studies have demonstrated that epithelial cell-conditioned medium from various confluent epithelial cell cultures inhibits urethral stricture fibroblasts’ proliferation and migration.[32] Urethral reconstruction with autologous urine-derived stem cells seeded in three-dimensional porous small intestinal submucosa in a rabbit model has been reported.[33] Urine-derived stem cells easily isolated from voided urine can be extensively expanded to high numbers in vitro, and efficiently give rise to urothelial cells and smooth muscle cells in vitro and vivo and can be utilized as a promising cell source for tissue-engineered urethras.[33]

SPHINCTER MUSCLE TISSUE

Research on stress incontinence in both sexes has been one of the focal areas of research.[34] A triple injection of muscle, neuron, and endothelial progenitor cells derived from human amniotic fluid stem cells has shown promising results in mice with urethral sphincter damage.[35] The common sources for clinical use include skeletal-muscle-derived progenitor cells or muscle-derived stem cells, myoblasts and fibroblasts. The advantage of muscle-derived stem cells is that these will fuse to form postmitotic multinucleated myotubes and thus would limit persistent expansion and risk of obstruction that may occur with other cell sources such as fibroblasts. Carr et al. treated eight women with stress urinary incontinence with muscle-derived stem cells and reported a total continence rate of 70% at 1 year follow-up.[36] The use of myoblast and fibroblast therapy has been described as safe and effective treatment for postprostatectomy incontinence at 1-year follow-up.[37]

GONADAL TISSUE AND GAMETES

The various types of stem cells that have been used to produce sperm cells include Spermatogonial stem cell, embryonic stem cell, and bone marrow stem cell.

Spermatogonial stem cell was first described by Brinster et al. in transgenic mice in 1994.[38] They isolated stem cells from testes of donor male mice that repopulated sterile testes when injected into seminiferous tubules. The donor cell spermatogenesis in recipient testes showed normal morphological characteristics and produced mature spermatozoa. These unipotent stem cell lines are found in male testis. The spermatogonial stem cells can be harvested and cryopreserved and subsequently transplanted back to the patient during his reproductive age. This procedure can be especially beneficial in young children receiving chemotherapy or radiotherapy. The spermatogonial stem cell is still in experimental stage with limited research into efficacy of approach. The spermatogonial stem cells have also been shown to differentiate into renal cells, though further clinical research is needed to fully explore potential therapeutic strategies.[39]

In 2003, Hubner et al. reported that oocyte-like cells could be produced from mouse embryonic stem cells in vitro.[40] The artificial germ cells produced from mouse pluripotent stem cells proved to be functional as they were capable to differentiate into spermatozoa and oocytes that can give rise to live progeny.[41] However, these still showed some notable differences from their in vivo counterparts and culture conditions suitable to complete oogenesis and spermatogenesis in vitro have not been established yet.[41]

The ability to derive male germ cells from bone marrow stem cells reveals novel aspects of germ cell development and opens the possibilities for use of these cells in reproductive medicine. In mice, adult bone marrow cells, in a favorable testicular environment have been shown to differentiate into somatic and germ cell lineages.[42] The resident neighboring cells in the recipient testis may control site-appropriate stem cell differentiation.[42] This clinically relevant finding raises the possibility for treatment of male infertility and testosterone deficiency through the therapeutic use of stem cells. In humans, no live offspring have been documented so far and transdifferentiation is limited to the stage of spermatogonium only male germline stem cells maintain spermatogenesis in the postnatal human testis. Drusenheimer et al. showed that a small population of bone marrow cells can transdifferentiate to male germ cell-like cells.[43] These preliminary findings provided direct evidence that human bone marrow cells can differentiate to putative male germ cells, and thus, bone marrow is a potential source of male germ cells that could sustain sperm production.[43]

CAVERNOSAL TISSUE

The cell sources that have been explored for cavernosal tissue are bone marrow stem cells, adipose tissue-derived stem cells, and neural crest stem cells. They may be useful in cases with poorly developed phallus in proximal hypospadias especially when associated with disorders of sexual differentiation. They are also useful in erectile dysfunction where there is damage to penile cavernous smooth muscle cells and sinus endothelial cells.[4445]

Human neural crest stem cells transplanted into the rat penile corpus cavernosum differentiated into endothelial cells or smooth muscle cells, as shown by their expression of cell type-specific markers for the cell types.[44] Skeletal muscle-derived stem cells have been shown to get converted into smooth muscle cells when implanted into the corpora cavernosa in rat models.[45]

CONTEMPORARY DEVELOPMENTS AND NEW TECHNOLOGY

To overcome the challenges in tissue engineering and stem cell therapy, scientists having been exploring new approaches and targeting the cells with different approaches. These include

Improving angiogenesis by combination of stem cell approaches

It has been observed that the central area of the tissue engineered grafts have low nutrient and oxygen supply that is responsible for incomplete regeneration and graft failure.[46] This is specially noticed in the structurally larger grafts used for bladder reconstruction. Scientists have thus explored the use of a combination of the two stem cell types such as mesenchymal stem cell and hematopoietic stem progenitor cells which has shown to improve vascularization of the entire graft, including the center area.[47] In addition, adding prevascularization strategies such as generation of preformed microvascular networks in grafts before implantation to the combined stem cell approaches is also beneficial for implantation of large grafts.[48]

Utilizing induced pluripotent stem cells

Induced pluripotent stem cells (iPSCs) are prime candidates for tissue engineering strategies. They can be generated directly from adult tissue by converting adult cells to stem cells using specific genes that encode transcription factors. These can further be differentiated into urothelial and smooth muscle cells.[4950] The advantage of iPSCs is that though they share similar plasticity to embryonic stem cell, they can be easily obtained.

Exploring potential of urine-derived stem cells

The discovery of urine-derived stem cells has opened a new field for further research.[325] In vitro research with addition of urine-derived stem cells to small intestinal submucosa graft has shown better regeneration regarding urethral caliber, speed of urothelial regeneration, content of smooth muscle, and vessel density compared to an acellular graft with less inflammatory cell infiltration and fibrosis.[33]

Identification of progenitor cells in urothelium

Resident multipotent progenitor cells have been identified in the urinary bladder urothelium in the basal cells and intermediate cells following various types of experimental bladder injury. Exploring the potential of indigenous progenitor cell sources are needed for further growth in the field of regeneration.[51] These include cells such as keratin 5-positive basal cells and UP3-positive intermediate cells.[515253]

Understanding dynamic reciprocity and regulating remodeling

Understanding the molecular signaling pathways and dynamic crosstalk between the urothelium and stroma in bladder regeneration is vital for further growth in the field of regeneration.[51] Dynamic reciprocity refers to the interactions between urothelium and stroma and bi-directional interactions between tissue-engineered grafts and the host tissue environment.[54] These are vital as they influence the outcomes in tissue remodeling. The various elements of remodeling include host cell adhesion and migration, differentiation, immune response such as macrophage reaction, differentiation, apoptosis, and formation of new extracellular matrix. Two types of macrophages – pro-inflammatory M1 macrophages and anti-inflammatory macrophages of M2 type – play distinct roles in tissue repair during wound healing following injury of various tissues.[5155] Both types of macrophages are needed in a dynamic sequence during tissue repair. It has been shown that pore size and fiber diameter of scaffolds alter macrophage activation, leaning to an increased presence of either M1 or M2 phenotypes.[5657] A coordinated sequence of M1 over M2 dominance in an early period of repair seems to promote angiogenesis, while M2 over M1 dominance in later stages seem to induce functional tissue formation and diminish scar tissue formation.[5859]

Creating smart scaffolds

Smart scaffolds are the latest thrill in the field of tissue engineering. These specifically designed scaffolds release trophic factors in a set way and have been used to optimize host reactions toward the scaffold.[51] The timely release of bioactive substances such as interleukins-4, 13, interferon-regulatory factors, and pharmacologic modulators of retinoic acid-related peroxisome proliferator-activated receptor signaling may help to modulate the macrophages M1 and M2 involved in tissue regeneration.[606162] Retinoic acid-dependent pathways are important for the final stages of urothelial differentiation.[52] Although growth factors are present in acellular natural derived grafts, it has been seen that they do not yield satisfactory outcomes. Modifying such growth factors such as fibroblast growth factor-2 and insulin-like growth factor-1 fusion protein have been shown to promote vascularization, smooth muscle cell repair and ingrowth, and urodynamic parameters.[6364] Administration of a mixture of growth factors such as platelet-derived growth factor and vascular endothelial growth factor; nerve growth factor and vascular endothelial growth factor has yielded better histological outcome and functional bladder contractility; bladder capacity and compliance, respectively.[6566]

Utilizing nanotechnology

Nanotechnology involves the use of nanoparticles that are very small in size in the range of 0.1–100 nm. Nanoparticles are also unique in the fact that they show completely novel physicochemical properties from their bulk counterpart. The promise of nanoparticles for diagnostic and therapeutic applications has been widely explored. In stem cell research, nanoparticles can enable targeted and controlled stem cell delivery. Thus, use of nanoparticles helps to further harness the potential of stem cells for biotherapeutic applications as these small particles can be used to deliver the stem cells to the site of action that may be even intracellular. Thus, effective amalgamation of nanotechnology and stem cell-offers immense benefits.[6768] Various aspects of nanotechnology in stem cell research like stem cell visualization and imaging, genetic modifications and reprogramming by gene delivery systems and creating stem cell niches need attention.[67] Various nanocarrier systems, such as carbon nanotubes, quantum dots, nanofibers, nanoparticles, nanodiamonds, nanoparticle scaffold, etc., can be utilized for the delivery of stem cells inside the body [Figure 3]. Highly uniform nanoparticles widely used in medicine include colloidal gold nanoparticles, silver nanoparticles, silica nanoparticles, titania nanoparticles, inorganic fluorescent nanoparticles (quantum dots and upconversion nanocrystals), as well as biodegradable polymer nanoparticles. Carbon dots (C-dots) are generally small carbon nanoparticles (<10 nm in size) with good solubility high stability, good conductivity, low toxicity and strong luminescence, c-dots have found wide applications including imaging, biosensor, drug delivery, and tissue and implant engineering. They are highly uniform and monodisperse without agglomeration and aggregation. Fluorescent carbon quantum dots can have a wide particle size range for broad applications. Graphene and graphene oxide with different layers and sizes can also be used for stem cell delivery in larger quantities. The carbon nanotube can be of various types; single-walled, double-walled and multi-walled structures with different size and surface modifications. Nanotechnology-assisted adipose-derived stem cell therapy has been found beneficial in an experimental rodent model for erectile dysfunction of cavernous nerve injury.[69] With nanoparticles, beneficial effects with a lower dose of stem cells were seen thus reducing the chances of any inadvertent side effects of stem cell therapy.[69] The mechanism of erectile function improvement was proposed to be related to the regeneration of the smooth muscle, endothelium, and nerve tissues.

Figure 3

Numerous nanoparticles that can be used for stem cell delivery

CHALLENGES IN CLINICAL APPLICATIONS OF Tu ENGINEERING AND STEM CELL THERAPY IN PEDIATRIC UROLOGY

The attempts to form an artificial bladder date back to 1950s when nonbiodegradable synthetic materials such as polytetrafluoroethylene, silicone, rubber, polyvinyl, and polypropylene were tried but were found to rapidly encrust with prolonged urinary contact.[70] In addition, these materials were susceptible to bacterial colonization and foreign body reactions.[71] While basic stem cell and tissue engineering research is expanding rapidly, translation of the basic research to clinical practice has been at an exceedingly slow pace. Till date, there have been only limited applications of tissue engineering and stem cell therapy in pediatric urology. Most of the research has been mainly in the experimental domain. The main reasons for the same have been ethical concerns regarding the harvest and use of embryonic stem cells, lack of sufficient differentiation protocols and risk of senescence, and epigenetic modifications in adult stem cells.

Although the possibility of renal regeneration is exciting in children with poorly functioning kidneys due to congenital anomalies, and autologous bone marrow mononuclear cells are relatively safe and ethical, there are certain anticipated hurdles.[9]

Cell-based therapy may be more effective only in the initial few months of life when the kidney still has potential to develop

The bone marrow in patients with chronic renal injury is usually hypoplastic. One may have to resort to Erythropoietin injections to stimulate the bone marrow

Repeated doses of bone marrow-derived cells may be needed.

We need to implement tissue engineering with cell-based therapy individually for each case. Embryonic stem cells seem to be promising as far as plausibility of generation of many different cell types is concerned. However, the ethical concerns for the use of human embryonic stem cells are equally significant as the use of the embryo for the medicinal purpose is not approved ethically.[72] The existent national guidelines for stem cell research should be strictly adhered to depending on the jurisdiction where the research is carried out.[73] Autologous sources are advantageous as there is no need for immunosuppression for cellular therapies derived from them. However, autologous cells may be less desirable when dealing with genetic diseases because the cells may possess the same genetic predisposition to disease.[72]

FUTURE DIRECTIONS

Potential impact of advances in stem cell technology on prospective cell-based therapeutic approaches for pediatric urology is enormous. A thorough understanding of basic stem cell biology and protocols to guide their differentiation toward specific cell fates in vitro, is needed. There is probably a practical reason to explore the utility of stem cell research in urology as the organs comprising the lower urinary tract are easily accessible by endoscopy with minimal morbidity.[74]

As embryonic stem cell research is restricted by ethical issues, scientists have explored other cell sources, including progenitor and stem cells derived from adult tissues, amniotic fluid, placenta and urine. In addition, newer methods of generating stem cells are being developed including somatic cell nuclear transfer, in which the nucleus of an adult somatic cell is placed into an oocyte, and reprogramed to induce stem-cell-like behavior. Such techniques are now being used in tissue engineering. Induced pluripotent stem cells have the potential to establish a new area of immunocompatible, on-demand renal transplantation. Successful integration of these cells as autograft therapies will require the demonstration of safety and efficacy equal or superior to the existing gold standards of kidney allograft transplantation and dialysis.[75]

Techniques to regenerate bladder tissue have reached the clinic, and exciting progress is being made regeneration of the kidney and urethra.[7677] An ideal engineered bladder should be composed of biocompatible material. It should be able to sustain mechanical forces for bladder filling and emptying. Engineered urothelial layer should contain tight junctions-effective barrier between bladder mesenchyme and urine. It should provide environmental cues that support neovascular in growth and not rejection by host immune system.

Further work is needed to ensure proper muscle alignment and adequate neovascularization. Appropriate seeding techniques need to be designed that will enable us to replicate anatomically correct and physiologically functional engineered bladders, promote angiogenesis, and neural regeneration to create the ideal, dynamic urinary reservoir.

Cell therapy as a treatment for incontinence and infertility might soon become a reality.[76] Scientists and clinicians should be optimistic that regenerative medicine and tissue engineering will one day provide the mainstream treatment options for urologic disorders.[76] Therapeutic cloning and cellular reprogramming may provide a potentially limitless source of cells for tissue engineering applications.[77] Seeding cells efficiently and uniformly onto three-dimensional scaffolds is the key for engineering urological tissue with an ideal histological structure in vitro.[78] This allows cells to interact positively with biomaterials, resulting in successful reconstructive surgery.[78]

The three-dimensional (3D) bioprinting is a new technique to replace traditional tissue engineering.[79] Zhang et al. loaded bladder epithelial cells and smooth muscle cells in hydrogel and maintained sufficient viability and proliferation in the hydrogel.[79] The highly porous scaffold mimics a natural urethral base-membrane and facilitates contacts between the printed epithelial cells and smooth muscle cells on both sides of the scaffold and thus provides a strong foundation for future studies on 3D bioprinted urethra. Limitations with 3D bioprinting in its present format are decreased mechanical strength and poor tissue integration and biocompatibility of constructs when implanted in vivo.

Acellular extracellular matrices are useful for repairing small urethral defects and cell-seeded extracellular matrices can augment or replace defective tissue segments in the ureter and bladder.[80] Although further immense detailed research is the need of the hour to fully utilize the hidden potential of stem cell research and tissue engineering, there is cautious optimism that the field of tissue engineering will play an increasing role in the management of a spectrum of urological disease.[81]

CONCLUSIONS

To conclude, development of tissue engineering and stem-cell-based strategies for treatment of urological disorders is in its budding stage. It is vital to achieve a natural crosstalk between the transplanted stem cells and scaffold, host immune system and existing microenvironment. The delicate intricacies of the relationships between cell lines, extracellular matrices scaffolds and the host's physiological environment need to be fully elucidated to allow a rampant progress in tissue-engineering in urology from bench to bedside. Further basic and translational research is necessary to fully elucidate the possibilities and verify the optimal use of transplanted and residential stem cells along with biodegradable scaffolds made by tissue engineering.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.