Pflugers Arch - Eur J Physiol DOI 10.1007/s00424-014-1504-4

INVITED REVIEW

On the road to bioartificial organs X. Ren & H. C. Ott

Received: 21 February 2014 / Accepted: 18 March 2014 # Springer-Verlag Berlin Heidelberg 2014

Abstract Biological organs are highly orchestrated systems with well-coordinated positioning, grouping, and interaction of different cell types within their specialized extracellular environment. Bioartificial organs are intended to be functional replacements of native organs generated through bioengineering techniques and hold the potential to alleviate donor organ shortage for transplantation. The development, production, and evaluation of such bioartificial organs require synergistic efforts of biology, material science, engineering, and medicine. In this review, we highlight the emerging platforms enabling structured assembly of multiple cell types into functional grafts and discuss recent advances and challenges in the development of bioartificial organs, including cell sources, in vitro organ culture, in vivo evaluation, and clinical considerations. Keywords Bioartificial organs . Decellularization . Bioprinting . Organoid . Tissue engineering . Organ engineering

Current strategies for bioartificial organ engineering In order to generate functional replacements of native organs, two major endpoints need to be reached: the correct constituent cell types have to be generated, and a physiologic spatial X. Ren : H. C. Ott Center for Regenerative Medicine, Massachusetts General Hospital, Boston, MA, USA X. Ren : H. C. Ott Harvard Medical School, Boston, MA, USA H. C. Ott (*) Division of Thoracic Surgery, Department of Surgery, Massachusetts General Hospital, 185 Cambridge Street, CPZN 4800, Boston, MA 02114, USA e-mail: [email protected] H. C. Ott Harvard Stem Cell Institute, Boston, MA, USA

and functional organization has to be established, both of which are highly unique to specific organs [55]. Several technology platforms emerging in recent years provide promising starting points and at least the theoretical translational potential of transplantable bioartificial organs. These platforms include whole-organ decellularization and regeneration based on the utilization of organ’s native extracellular scaffold reflecting its unique geometry [8], three-dimensional organ printing based on computer-aided construction of matrix scaffold and targeted positioning of desired cell types [18, 50], and organoid engineering highlighted by the self-organization of different cell types into specific structures (Fig. 1) [38, 83]. Whole-organ decellularization and recellularization The native extracellular matrix (ECM) is produced and maintained by the resident cells of each tissue or organ, which reflects the unique cellular organization and constitutes the microenvironment for each cell type to survive and function [7]. Whole-organ decellularization by detergent perfusion through native blood vessels removes all the cellular components leaving the organ’s ECM scaffold more or less intact. Importantly, distinct compartments within decellularized organs are well-preserved, which enables the delivery of different cell types to their physiologic locations. Since the initial introduction in heart engineering [60], this approach has been applied to other organs, including lung [59, 63], liver [86], kidney [75], and intestine [85]. Proof-of-principle regeneration of bioartificial organs based on these scaffolds in rodent models has been achieved by seeding vascular endothelial cells and organ-specific cell types (cardiomyocytes for heart, corresponding epithelial cells for lung, liver, and kidney). Some of the resulting bioartificial organs were transplanted into animal models and mimicked functions of their native counterparts, such as gas exchange in lung, metabolic processing in liver, and urine production in kidney [59, 63, 86]. This regenerative approach has recently been upscaled to large animal and human scale [9, 11, 21, 56, 79, 93].

Pflugers Arch - Eur J Physiol Fig. 1 Current strategies for bioartificial organ engineering. a Whole-organ decellularization and regeneration, b threedimensional organ printing, and c organoid engineering

Most organ decellularization protocols are detergentbased, although enzymatic and physical methods have been evaluated [16]. Several detergents have been explored for the purpose of decellularization, including sodium dodecyl sulfate (SDS), sodium deoxycholate (SDC), and 3-[(3-cholamidopropyl)dimethylammonio]-1propanesulfonate (CHAPS). Many protocols allow the generation of acellular scaffolds; consensus has not been reached regarding the optimal approach/reagent for decellularization to maximize the preservation of ECM and removal of cellular components for each respective

organ [21, 62, 88]. Although selective cell delivery into different compartments within decellularized organ scaffold has been well-demonstrated (e.g., vascular vs. airway compartment in lung), the achievement of homogenous cell distribution within each compartment remains challenging, resulting in patchy tissue formation inside whole-organ scaffolds. Furthermore, targeted seeding of different locations within a compartment (e.g., proximal vs. distal airway in lung, proximal vs. distal tubule in kidney) has proven to be challenging. While there is evidence for cell-specific niches preserved within the

Pflugers Arch - Eur J Physiol

matrix, as seen in glomerular epithelium in kidney regeneration [75], further strategies to enable site-specific delivery and engraftment have to be developed. Bioprinting Many tissue-engineering approaches are based on scaffolds, on which cells adhere, survive, and function to enable the formation of living tissue. Instead of utilizing native organ scaffolds by stripping off all cells, bioprinting generates threedimensional (3D) scaffolds de novo through layer-by-layer additive robotic fabrication. The design files are generated by identifying key architectural structures of target organs and decomposing these exterior and interior features into series of parallel slices. These slices then serve as templates for additive fabrication [18, 51]. Indirect bioprinting involves generation of patterned scaffolds through 3D printing and seeding cells onto the printed scaffolds. For example, cartilage tissues were regenerated by delivering chondrocytes onto 3D-printed PEG/PBT scaffold, which was proven to be superior to that regenerated based on PEG/PBT scaffold manufactured by conventional leaching method [27]. Direct bioprinting of cells incorporates scaffold materials and cells at the same time and has been explored to engineer small-size cartilage and bone tissues [17, 20]. Cartilage implants printed using poly(ethylene glycol) dimethacrylate together with human chondrocytes integrated into surrounding native cartilage and showed evidence of cell-based extracellular matrix modification [17]. Compared to conventional scaffold manufacturing using molds, 3D scaffold printing through additive fabrication allows precise control of internal architecture with high reproducibility. This facilitates efficient screen and optimization of structural variables including scaffold porosity, pore size, and permeability, all of which are critical factors determining subsequent biological delivery and tissue regeneration [18, 27, 65]. In the case of indirect bioprinting for subsequent cell seeding, a larger range of fabrication technologies can be used, including contact-involving analog printing with a high spatial resolution of less than 1 μm [18]. A further advantage of this technology is the possibility of incorporation of bioactive molecules. Growth factors or cell adhesion molecules can be added during scaffold printing and delivered following predefined patterns or concentration gradients, to control the behavior of subsequently seeded cells [13, 35]. In direct cell printing, the entire manufacturing process needs to be biocompatible and free of cytotoxicity. Contactless inkjet printing is one of the commonly used methods for printing cells, which offers an attainable spatial resolution limit of 100 μm [18]. As we have discussed above, efficient and homogenous cell delivery into preexisting scaffolds remains challenging, especially for engineering solid tissues/organs such as bone and muscle. Direct cell printing

potentially overcomes this cell delivery challenge by incorporating cells and scaffold materials at the same time. The grafts generated by 3D bioprinting currently remain small and simple in structure [18]. To print grafts with more complex architecture and increased size closer to those of native organs, improvements have to be made to increase the resolution and scale of direct cell printing. Moreover, a deeper understanding of the cellular and ECM organization within target organs will lead to design files that mimic the hierarchical organization of native organs more closely. Organoid engineering Organoid engineering is an emerging technology that enables the engineering of mini-organs, or “organoids,” from differentiating embryonic stem cells (ESCs) or induced pluripotent stem cells (iPSCs) in 3D culture conditions. Through this approach, organ development has been mimicked for systems such as intestine [69, 68], pituitary [78], thyroid [3], retina [19, 54], and most recently cerebrum [38]. These studies highlight the self-assembly capability of differentiating cells in 3D culture. Vascularized liver organoids that mimicked many aspects of native liver’s anatomy and function were generated by cultivating human iPSC-derived hepatocytes with human endothelial and mesenchymal stem cells in 3D culture [83]. This further emphasizes the importance of stromal cells in organ construction and demonstrates the inherent capability of different cell types to coordinate their assembly during organ formation. Organoid engineering is generally believed to complement conventional 2D culture to further our understanding of organ development and to promote disease modeling for mechanism study and drug screen [30, 38, 68]. From a regenerative medicine point of view, these mini-organs reveal the striking self-assembly capability of dissociated cells forming organized structure when proper 3D culture conditions are provided. In order to generate transplantable bioengineered grafts from such organoids, several challenges need to be addressed. Scaling organoids from currently millimeters to a clinically relevant size has to be accomplished. Although organoids mimic many aspects of native organs, even for complex structures such as the brain [38], they are still relatively simple structures. In order to bring organoids to the complexity and hierarchy of mature organs, supporting scaffold or structural guidance may need to be introduced. During development, many organs assemble in a modular fashion with interconnected functional units, such as human lung having about 500 million alveoli and human kidney having about 500 thousand glomeruli [33, 58]. Constructing the functional units or a cluster of these units through selfassembly may be accomplished through organoid engineering. These units can then serve as building blocks for subsequent organ engineering approaches. For example, mini-tissue

Pflugers Arch - Eur J Physiol

spheroids with different cellular compositions and organizations have been used as building blocks for macrotissue formation during organ printing [51]. Knowledge acquired and protocols developed in organoid engineering will certainly benefit recellularization of decellularized organ scaffold and organ printing. Understanding the unique interaction between different cell types during self-assembly will help to develop strategies and culture conditions promoting organ regeneration.

Providing the right cells for organ engineering With the tremendous progress that has been made in stem cell biology and especially the development of iPSC technology, generating “personalized” bioartificial organs from patientderived cells has become a theoretical possibility. While primary adult-derived stem and progenitor cells and embryonic stem cells continue to be of scientific interest, we decided to focus the following discussion on topics specifically related to the use of iPSC-derived cells in organ engineering. Cellular differentiation One of the strategies currently used for directed differentiation of human ESCs and iPSCs is to recapitulate developmental programs using sequential administration of growth factor cocktails. This strategy has proven to be successful in generating various differentiated cell types of all the three germ layers. As an emerging trend, compound screens were performed and identified small molecules that can enhance or replace the action of growth factors during directed differentiation, resulting in improved iPSC differentiation towards endoderm [12], pancreatic progenitors [14], hepatocytes [71], endothelial cells [32], specific neuronal lineages [22], and many others. Growth factor-free protocols have been developed for cardiomyocyte differentiation with up to 98 % efficiency [43, 49]. The use of small molecules to replace growth factors during iPSC differentiation offers advantages including improved reproducibility, lowered risk of contamination, and reduced cost, which will eventually facilitate the generation of scalable sources of clinical-grade cells for tissue and organ engineering. There are several concerns associated with the use of iPSCderived cells for clinical applications. First, the conventional way of generating iPSCs involves virus-mediated transduction of reprogramming factors [81]. The safety concerns related to viral transduction were alleviated by the development of non-integrating and non-viral methods of expressing reprograming factors using synthetic mRNAs [90] and episomal vectors [94]. Most recently, mouse iPSCs were generated from somatic cells using only chemical compounds [28]. This technique could facilitate clinical application of iPSC

technology if applied to generate human iPSCs. Second, the differentiation competency of different iPSC lines can be highly variable [36, 52], which indicates that newly generated iPSC lines need to be carefully characterized regarding their pluripotency and differentiation potential. Lastly, human iPSC lines were found to have a high level of mutational load, the majority of which were enriched in genes mutated or having causative effects in cancers [23]. This raises a concern that some of these mutations could increase disease risk, especially cancer risk, when iPSC-derived cells/tissues are used in clinical applications. Thus, standardized genetic evaluation of iPSC lines is required to ensure safety before clinical use [23]. Functional characterization Different cell types and their spatial organization determine the identity of an organ and provide the basis for its function. Cells can be characterized both molecularly and functionally. Molecular characterization generally refers to the analysis of a combined expression of genes (mainly transcriptional factors or surface markers) and represents the initial characterization of cell lineages differentiated from iPSCs. Functional characterization on the other hand can be complicated. The functions of cells are determined not only by cell-autonomous programs (gene expression profiles) but also by their niche, which is composed of neighboring cell types, ECM, and physiological stimuli [66]. For example, the acquisition of polarization and ciliation and the establishment of proper barrier function in iPSC-derived airway epithelial cells require ex vivo air-liquid interface culture that mimics post-natal airway epithelial environment [91]. Although many of ESC/iPSC-derived cell types display molecular profiles similar to their in vivo counterparts, they are usually functionally immature [80]. Cardiomyocytes derived from ESCs exhibit fetal instead of adult ventricular characteristics [25, 34, 53]. It was recently shown that iPSCs derived from murine ventricular myocytes exhibited markedly higher propensity to further differentiate into cardiomyocytes with a ventricular phenotype [92]. This indicates that establishment of certain epigenetic signatures is required for further maturation of iPSC-derived cells. These signatures are currently not recapitulated during in vitro differentiation. It remains challenging to develop in vitro culture conditions that enable the crossing of epigenetic barriers to efficiently differentiate desired cell types from generic cell sources that are easy to obtain, such as skin fibroblasts. As the differentiation efficiency of various cell types improves, more attention is required to evaluate and promote the acquisition of mature functions. Cellular heterogeneity Cell types such as vascular endothelial cells and fibroblasts are important building blocks for almost all organs. Up to now,

Pflugers Arch - Eur J Physiol

generic endothelial cells and fibroblasts have been used for the purpose of tissue engineering. These generic cells include mature cells isolated from primary tissues (e.g., human umbilical vein endothelial cells) [37, 83], adult-derived progenitors (e.g., circulating endothelial progenitors or mesenchymal stem cells) [6, 83], and those differentiated from ESCs or iPSCs [67, 89]. Recent studies indicate that endothelial cells contain highly heterogeneous and specialized subpopulations with morphological and functional differences among different organs (e.g., non-fenestrated endothelium in lung capillary vs. discontinuous fenestrated endothelium in liver capillary) [1, 2, 5]. Microvasculature also displays remarkable differences within organs such as liver and kidney (e.g., glomerular capillary vs. descending vasa recta endothelium) [1]. We need to consider these functional differences when choosing endothelial cells for regenerating higher organ functions such as filtration or gas exchange. The molecular mechanism directing endothelial heterogeneity just starts to be understood. Microarray expression profiling has been performed comparing a large number of in vitro cultured endothelial cell lines and comparing microvascular endothelial cells freshly isolated from different organs. These studies reveal molecular signatures unique to the microvasculature of each organ [15, 57]. These molecular signatures seem to be acquired through the interaction between endothelial cells and their organspecific niches, as generic ESC-derived endothelial cells underwent in vivo tissue-specific maturation after engrafted in regenerating tissues [57]. Similarly, blood-brain barrier endothelial cells were generated through sequential neural and endothelial differentiation of human iPSCs [44], which further indicates that the in vivo endothelial maturation can be mimicked by in vitro co-culture of endothelial cells with specific neighboring cell types. These recent advances open up the possibility of generating tissuespecific endothelial cells, which can be used to construct morphologically and functionally specified vascular networks to achieve closer modeling of organ function through bioengineering. Similar heterogeneity is also present in stromal support cells such as fibroblasts. Fibroblasts with distinct marker expression are associated with specific tissues or neighboring cell types. Taking the lung as an example, fibroblasts expressing NG2 proteoglycan are mainly associated with lung capillaries and are usually referred as pericytes [64]. On the other hand, fibroblasts expressing platelet-derived growth factor receptor alpha (PDGFRA) but not NG2 were able to recapitulate the alveolar type 2 stem cell niche supporting stem cell growth and differentiation [10]. These studies imply that different fibroblast subpopulations may need to be recruited for optimal construction of lung capillaries and airway structures.

In vitro expansion Regeneration of human-sized organs requires a strikingly large number of cells. For example, human lung is predicted to contain about 220 billion cells [76]. Although human iPSCs theoretically represent an unlimited source of cells, purification is generally required during or after iPSC differentiation to enrich the desired cell types. Purification is usually done by fluorescence-activated cell sorting (FACS) or magneticactivated cell sorting (MACS). Both techniques are not suited for purification of the large number of mature cells necessary for human organ regeneration. Ideally, the purification step would be performed at an earlier differentiation stage, with the resulting lineage-committed cells to be expanded to the desired number. However, once certain developmental milestones are achieved, expansion of the resulting cell types during long-term in vitro culture without senescence becomes challenging, risking potential loss of identity or functional competency. In recent years, several attempts have been made to improve long-term expansion of ESC/iPSCderived cell types by introducing assisting molecules, matrices, and feeder cells. Through a compound screen, TGF-β inhibitor was identified as a potent small molecule additive to promote the expansion ESC-derived endothelial cells after FACS purification [32]. ESC/iPSCderived hepatoblast-like cells can be extensively expanded and functionally maintained by using selected ECM coating materials [82]. Furthermore, the proliferation and self-renewal of isolated pancreatic progenitors can be achieved when co-cultured with matching mesenchymal cells [74]. It is likely that these approaches or their combination will make it possible to expand desired iPSC-derived cell lineages at multiple differentiation stages, which can then be tested for optimal performance in organ engineering. One of the questions we are currently trying to answer is the identification of the ideal seeding stage, when the transition from two-dimensional expansion to three-dimensional organ culture can be made. Matching cell differentiation stage to environment will be key in order to achieve cell engraftment and formation of functional tissues [83, 87].

Organ assembly and maturation Once scaffolds have been repopulated with regenerative cells, in vitro culture conditions of the resulting 3D multicellular constructs become key drivers of functional tissue formation and maturation. Proper in vitro culture conditions need to (1) support cell metabolism, (2) promote cell-cell and cellscaffold interaction, and (3) promote the acquisition of desired tissue/organ function.

Pflugers Arch - Eur J Physiol

Vascularization Proper metabolism of tissue implants with a volume more than 2 to 3 mm3 cannot be supported by passive diffusion and requires a preexistent perfusable vascular network [47, 72]. Therefore, regeneration of functional vasculature within tissue-engineered organs to support efficient oxygen and medium perfusion is a prerequisite for maintaining cell metabolism and survival during long-term in vitro culture. This has been one of the main focuses of organ engineering based on all the platforms we have discussed. Wholeorgan decellularization by perfusing detergent through the main vessel supplying the organ preserves the acellular vascular bed with high integrity of underlying basement membranes [60, 86]. Vascular regeneration was therefore attempted by seeding the decellularized scaffold with endothelial cells through the main artery, resulting in re-endothelialization of the previous vascular track, and recapitulating the complexity of organ-specific vascular hierarchy [59, 60, 63, 75]. In 3D bioprinting, patterned vascular networks were generated by seeding endothelial cells around a three-dimensional filament network of carbohydrate glass. After removal of the template carbohydrate, the engineered vascular network enabled blood perfusion under high-pressure pulsatile flow, supporting the survival of other cell types seeded in the perivascular space [48]. In the case of organoid-based approaches, all the constituent cell types, including endothelial cells, are mixed together in ECM gels. In liver organoid engineering, vascular networks formed by endothelial self-assembly integrated well with the perivascular hepatocytes and mesenchymal cells, giving rise to functional tissues enabling blood perfusion and metabolite secretion [83]. Stimulating tissue formation In an ideal experiment, selective cell delivery positions specific cells in close vicinity to each other and thereby mimics native tissue organization. Subsequent tissue formation involves massive cellular remodeling setting up proper connections between cells and between cells and scaffold. Taking vascular regeneration as an example, endothelial cells seeded into decellularized organ scaffold start as a cord of cells. Vessel formation involves vascular lumen formation and establishment of apicobasal polarity. This process somewhat mimics vasculogenesis in vivo [26, 77]. Several approaches are being developed to facilitate tissue formation in bioengineered grafts, including introducing assisting stromal cells and scaffold functionalization with growth factors. Mesenchymal cells or fibroblasts are the most commonly used stromal cells for tissue engineering. Interestingly, the presence of fibroblasts or mesenchymal stem cells significantly improved long-term survival of endothelial cells in vascular grafts and promoted perfusable vessel formation [6, 37, 46,

67, 89]. Similarly, co-seeded fibroblasts facilitated skeletal muscle tissue formation from primary myoblasts by maintaining myotube viability, mediating biomaterial degradation, and promoting muscle self-assembly [41]. In liver organoid engineering, mesenchymal stem cells were essential for 3D tissue formation of co-seeded hepatocytes and endothelial cells [83]. Protocols are being established to isolate and expand clinicalgrade mesenchymal stem cells from human tissues for the purpose of cell therapy [70]. Mesenchymal stem cells differentiated from iPSCs can also be an alternative source of stromal cells [42]. Growth factors constitute a rich source of extracellular signals directing cell specification and tissue formation during animal development. It is noted that a significant portion of growth factors are bonded to ECM, and this ECM interaction allows the control of growth factor presentation in a temporal and spatial fashion [39]. Therefore, improving organ scaffolds by immobilizing growth factors offers the potential to facilitate tissue formation. Soluble growth factors are usually internalized after binding to their receptors. Compared to soluble factors, scaffold-immobilized growth factors cannot be internalized, which leads to more sustained activation of receptors and downstream signaling pathways [31]. Growth factors can be immobilized through biopolymetric gels, such as hydrogel, in which growth factors conjugated to matrix moieties are incorporated into the gel during matrix polymerization [39, 73]. This approach can be readily incorporated into bioprinting and organoid engineering, because in both systems, biopolymetric gels are conventionally used as cell carriers and supporting scaffolds. Another approach is direct chemical cross-linking, which is used to immobilize growth factors onto preexisting synthetic or natural scaffolds. Various growth factors have been immobilized through direct chemical cross-linking to promote angiogenesis, bone repair, dermal wound healing, and stem cell differentiation [31, 45]. However, challenges remain when applying direct chemical cross-linking to functionalizing decellularized whole-organ scaffolds. The non-selectivity of chemical reactions usually induces massive cross-linking of ECM, leading to significant change in scaffold properties, especially mechanical properties. This will be detrimental to the engineering of organs such as heart and lung, in which the tissue elasticity is essential for organ function. Biomimetic culture conditions Once cells are differentiated, positioned, and integrated into the physiologic context of multicellular tissue, functional stimulation promotes organ maturation during the later stages of in vivo development. Therefore, incorporating biomimetic culture conditions mimicking native physiological stimuli should facilitate the maturation of bioartificial organs in vitro. Electrical and mechanical stimulations have been

Pflugers Arch - Eur J Physiol

incorporated into the engineering of heart tissues on various scales, including monolayer cell sheet, 3D tissue, and whole heart regenerated from decellularized scaffolds [24, 60, 84]. These conditions improve the maintenance of differentiated cell phenotype and promote the assembly of synchronously contractile cardiac tissues [24, 84]. Organ regeneration based on decellularized whole-organ matrix makes it possible to apply more physiological stimulations during in vitro organ culture, due to the anatomical features with preserved mechanical strength. Various bioreactors have been developed for in vitro culture of bioengineered whole heart, lung, kidney, and liver [60, 59, 75, 86], enabling organ-specific stimulations and functional measurements. The lack of mechanical strength remains a challenge for organ engineering based on bioprinting and organoid, which limits the types and strength of mechanical stimulations that can be applied. Nonetheless, it was recently shown that biomimetic stimulations can be performed even on microfluidic devices that mimic the essential composition of an organ [29].

Moving into clinical application Preclinical evaluation in animal models The ultimate goal of organ engineering is to implant bioartificial constructs to augment or replace lost tissue and organ function [8, 25]. From a regulatory perspective, bioartificial organs are biologic devices and need to undergo vigorous characterization and safety examination as governed by the Food and Drug Administration (FDA) [4, 40]. In order to guarantee patient safety, it will be critical to test the in vivo performance of bioartificial organs in animal models of human disease before moving into clinical application. Small tissue-engineered grafts can be implanted into the cranial window of immunodeficient rodent models, in which graft survival, blood perfusion, and metabolic functions can be studied. This approach has been applied to various tissueengineered vascular grafts and liver organoids [37, 83]. Vascular grafts without in vitro culture became perfused with recipient’s blood within 2 weeks after implantation [89]. Liver organoids, after in vitro culture for 4 to 6 days, connected with host vasculatures within 48 h after implantation, allowing various metabolic functions to be measured in vivo [83]. Another approach is to implant tissue-engineered graft as a patch attaching to preexisting organs, which has been wellexplored in heart tissue engineering. Heart tissue was engineered using neonatal rat heart cells and ECM gel reconstituted in circular mold, which improved systolic and diastolic function in infarcted rat heart after implantation as patch [95]. The preservation of almost all the surgically relevant structures in decellularized organs could allow for orthotopic

transplantation after in vitro regeneration. Paired organs such as lung and kidney further make it possible to have one side of the organ to be replaced with the regenerated counterpart, which allows the recipient to survive while the bioartificial organ is to be tested in vivo. Single lungs regenerated from decellularized scaffold have been transplanted orthotopically by connecting pulmonary artery, pulmonary vein, and bronchus. The resulting transplanted bioartificial lungs were perfused with recipient’s circulation, ventilated through recipient’s airway, and participated in gas exchange in vivo for hours [59, 63]. Similarly, tissue-engineered kidneys were transplanted in an orthotopic position made available by experimental left nephrectomy and were able to produce urine [75]. Alternatively, ectopic transplantation can be performed, in which the regenerated organ is not positioned in its normal anatomical position. For example, liver grafts engineered from decellularized scaffold were transplanted by connecting to recipient’s renal vein and artery [86]. These studies highlighted the advantage of physiologic graft size and the anatomic features within the tissue-engineered grafts [75]. These implantation/transplantation models represent the preliminary proof of principle of bioartificial grafts generated in vitro providing measurable function in vivo. Most of these tissue-engineered constructs did not last for prolonged periods of time in vivo [59, 63, 86]. Optimizations on the following aspects are required to improve their long-term in vivo performance. First, graft vascularization needs to reach high endothelial coverage and proper barrier function to avoid blood clotting or hemorrhage. This is especially critical in models where engineered grafts will be connected to the host’s circulation by surgical anastomosis and receive blood perfusion immediately after transplantation [59, 63]. Second, the length and degree of in vitro maturation that is necessary before successful long-term transplantation has to be defined [59]. Lastly, immunosuppression protocols need to be developed to optimize the long-term performance of transplanted grafts. Athymic rodent models with compromised T cells have been used for transplantation of organs engineered using human cells [59]. However, it has been suggested that other components of the immune system, such as macrophages, may also be involved in the response against tissueengineered organ scaffold after implantation [61].

Regulatory considerations Bioartificial organs represent complex products with a combination of cells, scaffolds, and other factors. This complexity necessitates the establishment of procedures during product development, manufacturing, and characterization to ensure safety and effectiveness [40]. Recognition and identification of the regulatory pathways and understanding of the requirements at clinical phases will help to structure the early

Pflugers Arch - Eur J Physiol

development of bioartificial organs as potential regenerative medicine products [4]. Cells are regulated by FDA using the tissue rules, which emphasize the prevention of infectious disease transmission. To comply with this requirement, procedures need to be established to control cell sources (such as donor eligibility), manufacturing facilities and protocols, and cell characterization (such as biological phenotype and sterility) [40]. Differentiated cell types derived from human iPSCs become prevalent for the purpose of regenerative medicine. As we have discussed in the previous sections, these cells need to be well-characterized regarding their identity, purity, and functional maturity before being incorporated into tissueengineered products [4]. Any pluripotent cells need to be removed due to their teratogenicity. When multiple cell types are to be combined for organ regeneration, it needs to be a known mixture, which means that all the participating cell types need to be pure and well-characterized before being combined. Scaffold can be derived from synthetic materials (such as polymeric materials), from extracted biomaterials (such as proteins and peptides), and from decellularization of human or animal tissues/organs. Similar to the aforementioned tissue rules regulating cell products, bioburden within the scaffolds needs to be strictly controlled to minimize infectious disease transmission. Moreover, the biocompatibility of scaffolds needs to be thoroughly investigated [40]. Complete removal of cytotoxic chemicals need to be demonstrated for materials derived from chemical synthesis or organ decellularization, because in both cases, reactive chemicals or detergents are heavily used during scaffold production. Moreover, scaffolds need to be fully characterized regarding their stability, physical, and mechanical properties. After cells and scaffolds are being well-characterized, novel properties will emerge after their combination. On the one hand, cells may behave differently after seeding onto 3D scaffold, which is different from their regular 2D culture environment. On the other hand, cells are constantly modifying the scaffold they are attaching to through ECM degradation and production. Therefore, the final tissue-engineered products require further characterization in vitro and in vivo to ensure desired quality [40]. The other challenge for cellscaffold composite products is the control of product consistency [40]. This necessitates the development of procedures that ensure consistent cell delivery, in vitro culture, and functional stimulations.

Conclusions and future directions The engineering of bioartificial organs aims to generate functional replacement of native organs and offers the hope to alleviate donor organ shortage. Several strategies were

successful in providing proof of principle that cells and scaffolds can be combined to form functional tissue. These strategies either mimic the organization of native organs by selective cell delivery (e.g., whole-organ decellularization and bioprinting) or create a microenvironment facilitating tissue formation by self-assembly of participating cells (e.g., organoid engineering). Optimization of culture conditions and application of biomimetic stimulations further allows tissue-engineered constructs to survive and mature in vitro, which can then be tested in vivo. In parallel, advances in stem cell biology continue to generate differentiated cell types with higher similarities to their native counterparts not only molecularly but also functionally. Bioartificial organ engineering needs to emphasize on the development of grafts capable of providing physiological functions. Here, we propose four levels of functional tests from cells to organs during the engineering process, taking lung regeneration as an example. The first level of functional test occurs at the level of cells and scaffolds before their combination (e.g., surfactant secretion of type 2 alveolar epithelial cells and proper elasticity of lung scaffold suited for ventilation). The second level is the functional test of different tissue units within each organ (e.g., perfusion and barrier function of lung vasculature and ventilation of the airway). The third level is the in vitro test of combined functional output of multiple constituent units within the regenerated organ (e.g., in vitro gas exchange of the regenerated lung). The final level is in vivo functional test of the regenerated organ after transplantation (e.g., blood perfusion and in vivo gas exchange of a regenerated lung after orthotopic transplantation). The last level is similar to regular physiological examination of native organs. However, care should be taken to ensure the functional tests on the initial three levels are physiologically relevant and representative. Identification of this stepwise hierarchy of functional tests will facilitate the definition of clear endpoints directing optimizations on each step during technology development and bring us closer the ultimate goal of generating functional organ replacements for clinical use.

References 1. Aird WC (2007) Phenotypic heterogeneity of the endothelium. Circ Res 100(2):174–190. doi:10.1161/01.RES.0000255690.03436.ae 2. Aird WC (2007) Phenotypic heterogeneity of the endothelium. Circ Res 100(2):158–173. doi:10.1161/01.RES.0000255691.76142.4a 3. Antonica F, Kasprzyk DF, Opitz R, Iacovino M, Liao X-H, Dumitrescu AM, Refetoff S, Peremans K, Manto M, Kyba M, Costagliola S (2012) Generation of functional thyroid from embryonic stem cells. Nature 491(7422):66–71. doi:10.1038/nature11525 4. Atala A, Kasper FK, Mikos AG (2012) Engineering complex tissues. Science Translational Medicine 4 (160):160rv112. doi:10.1126/ scitranslmed.3004890

Pflugers Arch - Eur J Physiol 5. Atkins GB, Jain MK, Hamik A (2011) Endothelial differentiation: molecular mechanisms of specification and heterogeneity. Arterioscler Thromb Vasc Biol 31(7):1476–1484. doi:10.1161/ atvbaha.111.228999 6. Au P, Tam J, Fukumura D, Jain RK (2008) Bone marrow-derived mesenchymal stem cells facilitate engineering of long-lasting functional vasculature. Blood 111(9):4551–4558. doi:10.1182/blood2007-10-118273 7. Badylak SF (2007) The extracellular matrix as a biologic scaffold material. Biomaterials 28(25):3587–3593. doi:10.1016/j. biomaterials.2007.04.043 8. Badylak SF, Taylor D, Uygun K (2011) Whole-organ tissue engineering: decellularization and recellularization of three-dimensional matrix scaffolds. Annu Rev Biomed Eng 13:27–53. doi:10.1146/ annurev-bioeng-071910-124743 9. Barakat O, Abbasi S, Rodriguez G, Rios J, Wood RP, Ozaki C, Holley LS, Gauthier PK (2012) Use of decellularized porcine liver for engineering humanized liver organ. J Surg Res 173(1):e11–e25. doi:10.1016/j.jss.2011.09.033 10. Barkauskas CE, Cronce MJ, Rackley CR, Bowie EJ, Keene DR, Stripp BR, Randell SH, Noble PW, Hogan BLM (2013) Type 2 alveolar cells are stem cells in adult lung. J Clin Invest 123(7): 3025–3036. doi:10.1172/JCI68782 11. Booth AJ, Hadley R, Cornett AM, Dreffs AA, Matthes SA, Tsui JL, Weiss K, Horowitz JC, Fiore VF, Barker TH, Moore BB, Martinez FJ, Niklason LE, White ES (2012) Acellular normal and fibrotic human lung matrices as a culture system for in vitro investigation. Am J Respir Crit Care Med 186(9):866–876. doi:10.1164/rccm. 201204-0754OC 12. Borowiak M, Maehr R, Chen S, Chen AE, Tang W, Fox JL, Schreiber SL, Melton DA (2009) Small molecules efficiently direct endodermal differentiation of mouse and human embryonic stem cells. Cell Stem Cell 4(4):348–358 13. Campbell PG, Miller ED, Fisher GW, Walker LM, Weiss LE (2005) Engineered spatial patterns of FGF-2 immobilized on fibrin direct cell organization. Biomaterials 26(33):6762–6770. doi:10.1016/j. biomaterials.2005.04.032 14. Chen S, Borowiak M, Fox JL, Maehr R, Osafune K, Davidow L, Lam K, Peng LF, Schreiber SL, Rubin LL, Melton D (2009) A small molecule that directs differentiation of human ESCs into the pancreatic lineage. Nat Chem Biol 5(4):258–265. doi:10.1038/nchembio.154 15. Chi J-T, Chang HY, Haraldsen G, Jahnsen FL, Troyanskaya OG, Chang DS, Wang Z, Rockson SG, van de Rijn M, Botstein D, Brown PO (2003) Endothelial cell diversity revealed by global expression profiling. Proc Natl Acad Sci 100(19):10623–10628. doi:10.1073/ pnas.1434429100 16. Crapo PM, Gilbert TW, Badylak SF (2011) An overview of tissue and whole organ decellularization processes. Biomaterials 32(12): 3233–3243. doi:10.1016/j.biomaterials.2011.01.057 17. Cui X, Breitenkamp K, Finn MG, Lotz M, D'Lima DD (2012) Direct human cartilage repair using three-dimensional bioprinting technology. Tissue Eng A 18(11–12):1304–1312. doi:10.1089/ten.TEA. 2011.0543 18. Derby B (2012) Printing and prototyping of tissues and scaffolds. Science 338(6109):921–926. doi:10.1126/science.1226340 19. Eiraku M, Takata N, Ishibashi H, Kawada M, Sakakura E, Okuda S, Sekiguchi K, Adachi T, Sasai Y (2011) Self-organizing optic-cup morphogenesis in three-dimensional culture. Nature 472(7341):51– 56. doi:10.1038/nature09941 20. Fedorovich NE, Alblas J, Hennink WE, Öner FC, Dhert WJA (2011) Organ printing: the future of bone regeneration? Trends Biotechnol 29(12):601–606. doi:10.1016/j.tibtech.2011.07.001 21. Gilpin SE, Guyette JP, Ren X, Gonzalez G, Xiong L, Song JJ, Vacanti J, Ott HC (2013) Up-scaling decellularization and whole organ culture for human lung regeneration. J Heart Lung Transplant 32(4):S69–S70. doi:10.1016/j.healun.2013.01.172

22. Gonzalez R, Garitaonandia I, Abramihina T, Wambua GK, Ostrowska A, Brock M, Noskov A, Boscolo FS, Craw JS, Laurent LC, Snyder EY, Semechkin RA (2013) Deriving dopaminergic neurons for clinical use. A practical approach. Sci Rep 3:1463. doi:10. 1038/srep01463 23. Gore A, Li Z, Fung H-L, Young JE, Agarwal S, Antosiewicz-Bourget J, Canto I, Giorgetti A, Israel MA, Kiskinis E, Lee J-H, Loh Y-H, Manos PD, Montserrat N, Panopoulos AD, Ruiz S, Wilbert ML, Yu J, Kirkness EF, Belmonte JCI, Rossi DJ, Thomson JA, Eggan K, Daley GQ, Goldstein LSB, Zhang K (2011) Somatic coding mutations in human induced pluripotent stem cells. Nature 471(7336):63–67. doi: 10.1038/nature09805 24. Govoni M, Muscari C, Guarnieri C, Giordano E (2013) Mechanostimulation protocols for cardiac tissue engineering. Biomed Res Int 2013:918640. doi:10.1155/2013/918640 25. Hansson EM, Lendahl U (2013) Regenerative medicine for the treatment of heart disease. J Intern Med 273(3):235–245. doi:10. 1111/joim.12033 26. Herbert SP, Stainier DYR (2011) Molecular control of endothelial cell behaviour during blood vessel morphogenesis. Nat Rev Mol Cell Biol 12(9):551–564. doi:10.1038/nrm3176 27. Hollister SJ (2005) Porous scaffold design for tissue engineering. Nat Mater 4(7):518–524 28. Hou P, Li Y, Zhang X, Liu C, Guan J, Li H, Zhao T, Ye J, Yang W, Liu K, Ge J, Xu J, Zhang Q, Zhao Y, Deng H (2013) Pluripotent stem cells induced from mouse somatic cells by small-molecule compounds. Science 341(6146):651–654. doi:10.1126/science.1239278 29. Huh D, Matthews BD, Mammoto A, Montoya-Zavala M, Hsin HY, Ingber DE (2010) Reconstituting organ-level lung functions on a chip. Science 328(5986):1662–1668. doi:10.1126/science.1188302 30. Hynds RE, Giangreco A (2013) Concise review: the relevance of human stem cell-derived organoid models for epithelial translational medicine. Stem Cells 31(3):417–422. doi:10.1002/stem.1290 31. Ito Y (2008) Covalently immobilized biosignal molecule materials for tissue engineering. Soft Matter 4(1):46–56. doi:10.1039/ B708359A 32. James DNH, Seandel M, Nolan D, Janovitz T, Tomishima M, Studer L, Lee G, Lyden D, Benezra R, Zaninovic N, Rosenwaks Z, Rabbany SY, Rafii S (2010) Expansion and maintenance of human embryonic stem cell-derived endothelial cells by TGFbeta inhibition is Id1 dependent. Nat Biotechnol 28(2):161–166. doi:10.1038/nbt.1605 33. Johnson KJ, Wreford NG, Hoy WE, Bertram JF (2000) Estimating total glomerular number in human kidneys with a physical disector/ smooth fractionator combination. Nephrology 5(3):A96–A96. doi: 10.1007/978-1-61779-851-1_30 34. Kehat I, Kenyagin-Karsenti D, Snir M, Segev H, Amit M, Gepstein A, Livne E, Binah O, Itskovitz-Eldor J, Gepstein L (2001) Human embryonic stem cells can differentiate into myocytes with structural and functional properties of cardiomyocytes. J Clin Invest 108(3): 407–414. doi:10.1172/JCI12131 35. Ker EDF, Nain AS, Weiss LE, Wang J, Suhan J, Amon CH, Campbell PG (2011) Bioprinting of growth factors onto aligned sub-micron fibrous scaffolds for simultaneous control of cell differentiation and alignment. Biomaterials 32(32):8097–8107. doi:10. 1016/j.biomaterials.2011.07.025 36. Koehler K, Tropel P, Theile J, Kondo T, Cummins T, Viville S, Hashino E (2011) Extended passaging increases the efficiency of neural differentiation from induced pluripotent stem cells. BMC Neurosci 12(1):1–14. doi:10.1186/1471-2202-12-82 37. Koike N, Fukumura D, Gralla O, Au P, Schechner JS, Jain RK (2004) Tissue engineering: creation of long-lasting blood vessels. Nature 428(6979):138–139. doi:10.1038/428138a 38. Lancaster MA, Renner M, Martin C-A, Wenzel D, Bicknell LS, Hurles ME, Homfray T, Penninger JM, Jackson AP, Knoblich JA (2013) Cerebral organoids model human brain development and microcephaly. Nature 501(7467):373–379. doi:10.1038/nature12517

Pflugers Arch - Eur J Physiol 39. Lee K, Silva EA, Mooney DJ (2011) Growth factor delivery-based tissue engineering: general approaches and a review of recent developments. J R Soc Interface 8(55):153–170. doi:10.1098/rsif.2010.0223 40. Lee MH, Arcidiacono JA, Bilek AM, Wille JJ, Hamill CA, Wonnacott KM, Wells MA, Oh SS (2009) Considerations for tissue-engineered and regenerative medicine product development prior to clinical trials in the United States. Tissue Eng B Rev 16(1): 41–54. doi:10.1089/ten.teb.2009.0449 41. Li M, Dickinson CE, Finkelstein EB, Neville CM, Sundback CA (2011) The role of fibroblasts in self-assembled skeletal muscle. Tissue Eng A 17(21–22):2641–2650. doi:10.1089/ten.TEA.2010.0700 42. Lian Q, Zhang Y, Zhang J, Zhang HK, Wu X, Zhang Y, Lam FF-Y, Kang S, Xia JC, Lai W-H, Au K-W, Chow YY, Siu C-W, Lee C-N, Tse H-F (2010) Functional mesenchymal stem cells derived from human induced pluripotent stem cells attenuate limb ischemia in mice. Circulation 121(9):1113–1123. doi:10.1161/circulationaha. 109.898312 43. Lian X, Hsiao C, Wilson G, Zhu K, Hazeltine LB, Azarin SM, Raval KK, Zhang J, Kamp TJ, Palecek SP (2012) Robust cardiomyocyte differentiation from human pluripotent stem cells via temporal modulation of canonical Wnt signaling. Proc Natl Acad Sci 109(27): E1848–E1857. doi:10.1073/pnas.1200250109 44. Lippmann ES, Azarin SM, Kay JE, Nessler RA, Wilson HK, AlAhmad A, Palecek SP, Shusta EV (2012) Derivation of blood-brain barrier endothelial cells from human pluripotent stem cells. Nat Biotech 30(8):783–791. doi:10.1038/nbt.2247 45. Masters KS (2011) Covalent growth factor immobilization strategies for tissue repair and regeneration. Macromol Biosci 11(9):1149– 1163. doi:10.1002/mabi.201000505 46. Melero-Martin JM, De Obaldia ME, Kang S-Y, Khan ZA, Yuan L, Oettgen P, Bischoff J (2008) Engineering robust and functional vascular networks in vivo with human adult and cord blood-derived progenitor cells. Circ Res 103(2):194–202. doi:10.1161/circresaha. 108.178590 47. Melero-Martin JM, Khan ZA, Picard A, Wu X, Paruchuri S, Bischoff J (2007) In vivo vasculogenic potential of human blood-derived endothelial progenitor cells. Blood 109(11):4761–4768. doi:10. 1182/blood-2006-12-062471 48. Miller JS, Stevens KR, Yang MT, Baker BM, Nguyen D-HT, Cohen DM, Toro E, Chen AA, Galie PA, Yu X, Chaturvedi R, Bhatia SN, Chen CS (2012) Rapid casting of patterned vascular networks for perfusable engineered three-dimensional tissues. Nat Mater 11(9): 768–774. doi:10.1038/nmat3357 49. Minami I, Yamada K, Otsuji TG, Yamamoto T, Shen Y, Otsuka S, Kadota S, Morone N, Barve M, Asai Y, Tenkova-Heuser T, Heuser JE, Uesugi M, Aiba K, Nakatsuji N (2012) A small molecule that promotes cardiac differentiation of human pluripotent stem cells under defined, cytokine- and xeno-free conditions. Cell Rep 2(5): 1448–1460 50. Mironov V, Boland T, Trusk T, Forgacs G, Markwald RR (2003) Organ printing: computer-aided jet-based 3D tissue engineering. Trends Biotechnol 21(4):157–161. doi:10.1016/S0167-7799(03) 00033-7 51. Mironov V, Visconti RP, Kasyanov V, Forgacs G, Drake CJ, Markwald RR (2009) Organ printing: tissue spheroids as building blocks. Biomaterials 30(12):2164–2174. doi:10.1016/j.biomaterials. 2008.12.084 52. Miura K, Okada Y, Aoi T, Okada A, Takahashi K, Okita K, Nakagawa M, Koyanagi M, Tanabe K, Ohnuki M, Ogawa D, Ikeda E, Okano H, Yamanaka S (2009) Variation in the safety of induced pluripotent stem cell lines. Nat Biotech 27(8):743–745. doi:10.1038/ nbt.1554 53. Mummery C, Ward-van Oostwaard D, Doevendans P, Spijker R, van den Brink S, Hassink R, van der Heyden M, Opthof T, Pera M, de la Riviere AB, Passier R, Tertoolen L (2003) Differentiation of human embryonic stem cells to cardiomyocytes: role of coculture with

54.

55.

56.

57.

58.

59.

60.

61.

62.

63.

64.

65.

66.

67.

68.

visceral endoderm-like cells. Circulation 107(21):2733–2740. doi: 10.1161/01.cir.0000068356.38592.68 Nakano T, Ando S, Takata N, Kawada M, Muguruma K, Sekiguchi K, Saito K, Yonemura S, Eiraku M, Sasai Y (2012) Self-formation of optic cups and storable stratified neural retina from human ESCs. Cell Stem Cell 10(6):771–785. doi:10.1016/j.stem.2012.05.009 Nelson CM, Bissell MJ (2006) Of extracellular matrix, scaffolds, and signaling: tissue architecture regulates development, homeostasis, and cancer. Annu Rev Cell Dev Biol 22(1):287–309. doi:10.1146/ annurev.cellbio.22.010305.104315 Nichols JE, Niles J, Riddle M, Vargas G, Schilagard T, Ma L, Edward K, Francesca SL, Sakamoto J, Vega S, Ogadegbe M, Mlcak R, Deyo D, Woodson L, McQuitty C, Lick S, Beckles D, Melo E, Cortiella J (2013) Production and assessment of decellularized pig and human lung scaffolds. Tissue Eng A 19(17–18):2045–2062. doi:10.1089/ ten.TEA.2012.0250 Nolan DJ, Ginsberg M, Israely E, Palikuqi B, Poulos MG, James D, Ding B-S, Schachterle W, Liu Y, Rosenwaks Z, Butler JM, Xiang J, Rafii A, Shido K, Rabbany SY, Elemento O, Rafii S (2013) Molecular signatures of tissue-specific microvascular endothelial cell heterogeneity in organ maintenance and regeneration. Dev Cell 26(2):204–219. doi:10.1016/j.devcel.2013.06.017 Ochs M, Nyengaard JR, Jung A, Knudsen L, Voigt M, Wahlers T, Richter J, Gundersen HJG (2004) The number of alveoli in the human lung. Am J Respir Crit Care Med 169(1):120–124. doi:10. 1164/rccm.200308-1107OC Ott HC, Clippinger B, Conrad C, Schuetz C, Pomerantseva I, Ikonomou L, Kotton D, Vacanti JP (2010) Regeneration and orthotopic transplantation of a bioartificial lung. Nat Med 16(8): 927–933. doi:10.1038/nm.2193 Ott HC, Matthiesen TS, Goh S-K, Black LD, Kren SM, Netoff TI, Taylor DA (2008) Perfusion-decellularized matrix: using nature’s platform to engineer a bioartificial heart. Nat Med 14(2):213–221. doi:10.1038/nm1684 Perniconi B, Costa A, Aulino P, Teodori L, Adamo S, Coletti D (2011) The pro-myogenic environment provided by whole organ scale acellular scaffolds from skeletal muscle. Biomaterials 32(31): 7870–7882. doi:10.1016/j.biomaterials.2011.07.016 Petersen TH, Calle EA, Colehour MB, Niklason LE (2012) Matrix composition and mechanics of decellularized lung scaffolds. Cells Tissues Organs 195(3):222–231 Petersen TH, Calle EA, Zhao L, Lee EJ, Gui L, Raredon MB, Gavrilov K, Yi T, Zhuang ZW, Breuer C, Herzog E, Niklason LE (2010) Tissue-engineered lungs for in vivo implantation. Science 329(5991):538–541. doi:10.1126/science.1189345 Rock JR, Barkauskas CE, Cronce MJ, Xue Y, Harris JR, Liang J, Noble PW, Hogan BLM (2011) Multiple stromal populations contribute to pulmonary fibrosis without evidence for epithelial to mesenchymal transition. Proc Natl Acad Sci 108(52):E1475–E1483. doi: 10.1073/pnas.1117988108 Sachlos E, Czernuszka J (2003) Making tissue engineering scaffolds work. Review: the application of solid freeform fabrication technology to the production of tissue engineering scaffolds. Eur Cell Mater 5:29–39 Saha K, Jaenisch R (2009) Technical challenges in using human induced pluripotent stem cells to model disease. Cell Stem Cell 5(6):584–595 Samuel R, Daheron L, Liao S, Vardam T, Kamoun WS, Batista A, Buecker C, Schäfer R, Han X, Au P, Scadden DT, Duda DG, Fukumura D, Jain RK (2013) Generation of functionally competent and durable engineered blood vessels from human induced pluripotent stem cells. Proc Natl Acad Sci 110(31):12774–12779. doi:10. 1073/pnas.1310675110 Sato T, Clevers H (2013) Growing self-organizing mini-guts from a single intestinal stem cell: mechanism and applications. Science 340(6137):1190–1194. doi:10.1126/science.1234852

Pflugers Arch - Eur J Physiol 69. Sato T, Vries RG, Snippert HJ, van de Wetering M, Barker N, Stange DE, van Es JH, Abo A, Kujala P, Peters PJ, Clevers H (2009) Single Lgr5 stem cells build crypt–villus structures in vitro without a mesenchymal niche. Nature 459(7244):262–265. doi:10.1038/ nature07935 70. Sensebé L (2008) Clinical grade production of mesenchymal stem cells. Biomed Mater Eng 18(1 Suppl):3–10 71. Shan J, Schwartz RE, Ross NT, Logan DJ, Thomas D, Duncan SA, North TE, Goessling W, Carpenter AE, Bhatia SN (2013) Identification of small molecules for human hepatocyte expansion and iPS differentiation. Nat Chem Biol 9(8):514–520. doi:10.1038/ nchembio.1270 72. Shieh S-J, Vacanti JP (2005) State-of-the-art tissue engineering: from tissue engineering to organ building. Surgery 137(1):1–7. doi:10. 1016/j.surg.2004.04.002 73. Silva AKA, Richard C, Bessodes M, Scherman D, Merten O-W (2008) Growth factor delivery approaches in hydrogels. Biomacromolecules 10(1):9–18. doi:10.1021/bm801103c 74. Sneddon JB, Borowiak M, Melton DA (2012) Self-renewal of embryonic-stem-cell-derived progenitors by organ-matched mesenchyme. Nature 491(7426):765–768. doi:10.1038/nature11463 75. Song JJ, Guyette JP, Gilpin SE, Gonzalez G, Vacanti JP, Ott HC (2013) Regeneration and experimental orthotopic transplantation of a bioengineered kidney. Nat Med 19(5):646–651. doi:10.1038/nm.3154 76. Stone K, Mercer R, Freeman B, Chang L, Crapo J (1992) Distribution of lung cell numbers and volumes between alveolar and nonalveolar tissue. Am Rev Respir Dis 146(2):454–456. doi: 10.1164/ajrccm/146.2.454 77. Strilić B, Kučera T, Eglinger J, Hughes MR, McNagny KM, Tsukita S, Dejana E, Ferrara N, Lammert E (2009) The molecular basis of vascular lumen formation in the developing mouse aorta. Dev Cell 17(4):505–515. doi:10.1016/j.devcel.2009.08.011 78. Suga H, Kadoshima T, Minaguchi M, Ohgushi M, Soen M, Nakano T, Takata N, Wataya T, Muguruma K, Miyoshi H, Yonemura S, Oiso Y, Sasai Y (2011) Self-formation of functional adenohypophysis in three-dimensional culture. Nature 480(7375):57–62. doi:10.1038/ nature10637 79. Sullivan DC, Mirmalek-Sani S-H, Deegan DB, Baptista PM, Aboushwareb T, Atala A, Yoo JJ (2012) Decellularization methods of porcine kidneys for whole organ engineering using a highthroughput system. Biomaterials 33(31):7756–7764. doi:10.1016/j. biomaterials.2012.07.023 80. Svendsen CN (2013) Back to the future: how human induced pluripotent stem cells will transform regenerative medicine. Hum Mol Genet 22(R1):R32–R38. doi:10.1093/hmg/ddt379 81. Takahashi K, Tanabe K, Ohnuki M, Narita M, Ichisaka T, Tomoda K, Yamanaka S (2007) Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell 131(5):861–872 82. Takayama K, Nagamoto Y, Mimura N, Tashiro K, Sakurai F, Tachibana M, Hayakawa T, Kawabata K, Mizuguchi H (2013) Long-term self-renewal of human ES/iPS-derived hepatoblast-like cells on human laminin 111-coated dishes. Stem Cell Rep 1(4):322– 335. doi:10.1016/j.stemcr.2013.08.006 83. Takebe T, Sekine K, Enomura M, Koike H, Kimura M, Ogaeri T, Zhang R-R, Ueno Y, Zheng Y-W, Koike N, Aoyama S, Adachi Y, Taniguchi H (2013) Vascularized and functional human liver from an iPSC-derived organ bud transplant. Nature 499(7459):481–484. doi: 10.1038/nature12271 84. Tandon N, Cannizzaro C, Chao P-HG, Maidhof R, Marsano A, Au HTH, Radisic M, Vunjak-Novakovic G (2009) Electrical stimulation

85.

86.

87.

88.

89.

90.

91.

92.

93.

94.

95.

systems for cardiac tissue engineering. Nat Protocol 4(2):155–173. doi:10.1038/nprot.2008.183 Totonelli G, Maghsoudlou P, Garriboli M, Riegler J, Orlando G, Burns AJ, Sebire NJ, Smith VV, Fishman JM, Ghionzoli M, Turmaine M, Birchall MA, Atala A, Soker S, Lythgoe MF, Seifalian A, Pierro A, Eaton S, De Coppi P (2012) A rat decellularized small bowel scaffold that preserves villus-crypt architecture for intestinal regeneration. Biomaterials 33(12):3401–3410. doi:10.1016/j.biomaterials.2012.01.012 Uygun BE, Soto-Gutierrez A, Yagi H, Izamis M-L, Guzzardi MA, Shulman C, Milwid J, Kobayashi N, Tilles A, Berthiaume F, Hertl M, Nahmias Y, Yarmush ML, Uygun K (2010) Organ reengineering through development of a transplantable recellularized liver graft using decellularized liver matrix. Nat Med 16(7):814–820. doi:10.1038/nm.2170 Vunjak-Novakovic G, Tandon N, Godier A, Maidhof R, Marsano A, Martens TP, Radisic M (2010) Challenges in cardiac tissue engineering. Tissue Eng B Rev 16(2):169–187. doi:10.1089/ten.teb.2009. 0352 Wallis JM, Borg ZD, Daly AB, Deng B, Ballif BA, Allen GB, Jaworski DM, Weiss DJ (2012) Comparative assessment of detergent-based protocols for mouse lung de-cellularization and recellularization. Tissue Eng C Methods 18(6):420–432. doi:10.1089/ ten.tec.2011.0567 Wang ZZ, Au P, Chen T, Shao Y, Daheron LM, Bai H, Arzigian M, Fukumura D, Jain RK, Scadden DT (2007) Endothelial cells derived from human embryonic stem cells form durable blood vessels in vivo. Nat Biotechnol 25(3):317–318. doi:10.1038/nbt1287 Warren L, Manos PD, Ahfeldt T, Loh Y-H, Li H, Lau F, Ebina W, Mandal PK, Smith ZD, Meissner A, Daley GQ, Brack AS, Collins JJ, Cowan C, Schlaeger TM, Rossi DJ (2010) Highly efficient reprogramming to pluripotency and directed differentiation of human cells with synthetic modified mRNA. Cell Stem Cell 7(5):618–630. doi:10.1016/j.stem.2010.08.012 Wong AP, Bear CE, Chin S, Pasceri P, Thompson TO, Huan L-J, Ratjen F, Ellis J, Rossant J (2012) Directed differentiation of human pluripotent stem cells into mature airway epithelia expressing functional CFTR protein. Nat Biotechnol 30(9):876–882. doi:10.1038/ nbt.2328 Xu H, Yi BA, Wu H, Bock C, Gu H, Lui KO, Park J-HC, Shao Y, Riley AK, Domian IJ, Hu E, Willette R, Lepore J, Meissner A, Wang Z, Chien KR (2012) Highly efficient derivation of ventricular cardiomyocytes from induced pluripotent stem cells with a distinct epigenetic signature. Cell Res 22(1):142–154. doi:10.1038/cr.2011.171 Yagi H, Fukumitsu K, Fukuda K, Kitago M, Shinoda M, Obara H, Itano O, Kawachi S, Tanabe M, Coudriet GM, Piganelli JD, Gilbert TW, Soto-Gutierrez A, Kitagawa Y (2013) Human-scale wholeorgan bioengineering for liver transplantation: a regenerative medicine approach. Cell Transplant 22(2):231–242. doi:10.3727/ 096368912X654939 Yu J, Hu K, Smuga-Otto K, Tian S, Stewart R, Slukvin II, Thomson JA (2009) Human induced pluripotent stem cells free of vector and transgene sequences. Science 324(5928):797–801. doi:10.1126/ science.1172482 Zimmermann W-H, Melnychenko I, Wasmeier G, Didie M, Naito H, Nixdorff U, Hess A, Budinsky L, Brune K, Michaelis B, Dhein S, Schwoerer A, Ehmke H, Eschenhagen T (2006) Engineered heart tissue grafts improve systolic and diastolic function in infarcted rat hearts. Nat Med 12(4):452–458. doi:10.1038/nm1394

On the road to bioartificial organs.

Biological organs are highly orchestrated systems with well-coordinated positioning, grouping, and interaction of different cell types within their sp...
2MB Sizes 0 Downloads 3 Views