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    Noninvasive in vivo 3D bioprinting. Chen Yuwen,Zhang Jiumeng,Liu Xuan,Wang Shuai,Tao Jie,Huang Yulan,Wu Wenbi,Li Yang,Zhou Kai,Wei Xiawei,Chen Shaochen,Li Xiang,Xu Xuewen,Cardon Ludwig,Qian Zhiyong,Gou Maling Science advances Three-dimensional (3D) printing technology has great potential in advancing clinical medicine. Currently, the in vivo application strategies for 3D-printed macroscale products are limited to surgical implantation or in situ 3D printing at the exposed trauma, both requiring exposure of the application site. Here, we show a digital near-infrared (NIR) photopolymerization (DNP)-based 3D printing technology that enables the noninvasive in vivo 3D bioprinting of tissue constructs. In this technology, the NIR is modulated into customized pattern by a digital micromirror device, and dynamically projected for spatially inducing the polymerization of monomer solutions. By ex vivo irradiation with the patterned NIR, the subcutaneously injected bioink can be noninvasively printed into customized tissue constructs in situ. Without surgery implantation, a personalized ear-like tissue constructs with chondrification and a muscle tissue repairable cell-laden conformal scaffold were obtained in vivo. This work provides a proof of concept of noninvasive in vivo 3D bioprinting. 10.1126/sciadv.aba7406
    The current state of tissue engineering in the management of hypospadias. Chan Yvonne Y,Bury Matthew I,Yura Emily M,Hofer Matthias D,Cheng Earl Y,Sharma Arun K Nature reviews. Urology Hypospadias is a congenital malformation resulting from the disruption of normal urethral formation with varying global prevalence. Hypospadias repair, especially that of proximal hypospadias (in which reconstruction of a long urethra is necessary), remains a surgical challenge despite more than two decades of surgical technique development and refinement. The lack of tissue substitutes with mechanical and biological properties similar to those of native urethra is a challenge for which the field of tissue engineering might offer promising solutions. However, the use of tissue-engineered constructs in preclinical studies is still hindered by complications such as strictures or fistulae, which have slowed progression to clinical application. Furthermore, the generation of uniform tubular constructs remains a challenge. Exciting advances in the application of nanotechnology and 3D bioprinting to urethral tissue engineering might present solutions to these issues. 10.1038/s41585-020-0281-4
    Encapsulated three-dimensional bioprinted structure seeded with urothelial cells: a new construction technique for tissue-engineered urinary tract patch. Jin Yi-Peng,Shi Chong,Wu Yuan-Yi,Sun Ji-Lei,Gao Jiang-Ping,Yang Yong Chinese medical journal BACKGROUND:Traditional tissue engineering methods to fabricate urinary tract patch have some drawbacks such as compromised cell viability and uneven cell distribution within scaffold. In this study, we combined three-dimensional (3D) bioprinting and tissue engineering method to form a tissue-engineered urinary tract patch, which could be employed for the application on Beagles urinary tract defect mode to verify its effectiveness on urinary tract reconstruction. METHODS:Human adipose-derived stem cells (hADSCs) were dropped into smooth muscle differentiation medium to generate induced microtissues (ID-MTs), flow cytometry was utilized to detect the positive percentage for CD44, CD105, CD45, and CD34 of hADSCs. Expression of vascular endothelial growth factor A (VEGFA) and tumor necrosis factor-stimulated gene-6 (TSG-6) in hADSCs and MTs were identified by Western blotting. Then the ID-MTs were employed for 3D bioprinting. The bioprinted structure was encapsulated by transplantation into the subcutaneous tissue of nude mice for 1 week. After retrieval of the encapsulated structure, hematoxylin and eosin and Masson's trichrome staining were performed to demonstrate the morphology and reveal collagen and smooth muscle fibers, integral optical density (IOD) and area of interest were calculated for further semi-quantitative analysis. Immunofluorescent double staining of CD31 and α-smooth muscle actin (α-SMA) were used to reveal vascularization of the encapsulated structure. Immunohistochemistry was performed to evaluate the expression of interleukin-2 (IL-2), α-SMA, and smoothelin of the MTs in the implanted structure. Afterward, the encapsulated structure was seeded with human urothelial cells. Immunofluorescent staining of cytokeratins AE1/AE3 was applied to inspect the morphology of seeded encapsulated structure. RESULTS:The semi-quantitative assay showed that the relative protein expression of VEGFA was 0.355 ± 0.038 in the hADSCs vs. 0.649 ± 0.150 in the MTs (t = 3.291, P = 0.030), while TSG-6 expression was 0.492 ± 0.092 in the hADSCs vs. 1.256 ± 0.401 in the MTs (t = 3.216, P = 0.032). The semi-quantitative analysis showed that the mean IOD of IL-2 in the MT group was 7.67 ± 1.26, while 12.6 ± 4.79 in the hADSCs group, but semi-quantitative analysis showed that there was no statistical significance in the difference between the two groups (t = 1.724, P = 0.16). The semi-quantitative analysis showed that IOD was 71.7 ± 14.2 in non-induced MTs (NI-MTs) vs. 35.7 ± 11.4 in ID-MTs for collagen fibers (t = 3.428, P = 0.027) and 12.8 ± 1.9 in NI-MTs vs. 30.6 ± 8.9 in ID-MTs for smooth muscle fibers (t = 3.369, P = 0.028); furthermore, the mean IOD was 0.0613 ± 0.0172 in ID-MTs vs. 0.0017 ± 0.0009 in NI-MTs for α-SMA (t = 5.994, P = 0.027), while 0.0355 ± 0.0128 in ID-MTs vs. 0.0035 ± 0.0022 in NI-MTs for smoothelin (t = 4.268, P = 0.013), which indicate that 3D bioprinted structure containing ID-MTs could mimic the smooth muscle layer of native urinary tract. After encapsulation of the urinary tract patch for additional cell adhesion, urothelial cells were seeded onto the encapsulated structures, and a monolayer urothelial cell was observed. CONCLUSION:Through 3D bioprinting and tissue engineering methods, we provided a promising way to fabricate tissue-engineered urinary tract patch for further investigation. 10.1097/CM9.0000000000000654
    [The application and prospects of three dimensional bioprinting in urinary system reconstruction]. Gong Lina,Jin Xi,Li Hong,Wang Kunjie Sheng wu yi xue gong cheng xue za zhi = Journal of biomedical engineering = Shengwu yixue gongchengxue zazhi Three dimensional (3D) bioprinting is a new biological tissue engineering technology in recent years. The development of 3D bioprinting is conducive to solving the current problems of clinical tissue and organ repairing. This article provides a review about the clinical and research status of 3D bioprinting and urinary system reconstruction. Furthermore, the feasibility and clinical value of 3D bioprinting in urinary system reconstruction will be also discussed. 10.7507/1001-5515.201912016
    Organ-on-chip models: Implications in drug discovery and clinical applications. Mittal Rahul,Woo Frank W,Castro Carlo S,Cohen Madeline A,Karanxha Joana,Mittal Jeenu,Chhibber Tanya,Jhaveri Vasanti M Journal of cellular physiology Before a lead compound goes through a clinical trial, preclinical studies utilize two-dimensional (2D) in vitro models and animal models to study the pharmacodynamics and pharmacokinetics of that lead compound. However, these current preclinical studies may not accurately represent the efficacy and safety of a lead compound in humans, as there has been a high failure rate of drugs that enter clinical trials. All of these failures and the associated costs demonstrate a need for more representative models of human organ systems for screening in the preclinical phase of drug development. In this study, we review the recent advances in in vitro modeling including three-dimensional (3D) organoids, 3D microfabrication, and 3D bioprinting for various organs including the heart, kidney, lung, gastrointestinal tract (intestine-gut-stomach), liver, placenta, adipose, retina, bone, and brain as well as multiorgan models. The availability of organ-on-chip models provides a wealth of opportunities to understand the pathogenesis of human diseases and provide a potentially better model to screen a drug, as these models utilize a dynamic 3D environment similar to the human body. Although there are limitations of organ-on-chip models, the emergence of new technologies have refined their capability for translational research as well as precision medicine. 10.1002/jcp.27729
    Research project: Charleston Bioengineered Kidney Project. Mironov Vladimir,Drake Christopher,Wen Xuejun Biotechnology journal The goal of Charleston Bioengineered Kidney Project is to engineer a functional living human kidney suitable for surgical implantation using principles of directed tissue self-assembly and tissue fusion. This is a multidisciplinary project which incorporates multiple innovative bioengineering technologies and expertise from a broad spectrum of disciplines. The conceptual framework, engineering principles, design, potential cell source as well as the first preliminary data demonstrating the feasibility of the proposed Charleston Bioengineered Kidney Project are outlined. The potential challenges are described. Finally, the experts' opinion about the proposed project is also presented. 10.1002/biot.200690107
    In vitro systems to study nephropharmacology: 2D versus 3D models. Sánchez-Romero Natalia,Schophuizen Carolien M S,Giménez Ignacio,Masereeuw Rosalinde European journal of pharmacology The conventional 2-dimensional (2D) cell culture is an invaluable tool in, amongst others, cell biology and experimental pharmacology. However, cells cultured in 2D, on the top of stiff plastic plates lose their phenotypical characteristics and fail in recreating the physiological environment found in vivo. This is a fundamental requirement when the goal of the study is to get a rigorous predictive response of human drug action and safety. Recent approaches in the field of renal cell biology are focused on the generation of 3D cell culture models due to the more bona fide features that they exhibit and the fact that they are more closely related to the observed physiological conditions, and better predict in vivo drug handling. In this review, we describe the currently available 3D in vitro models of the kidney, and some future directions for studying renal drug handling, disease modeling and kidney regeneration. 10.1016/j.ejphar.2016.07.010
    Autologous Cells for Kidney Bioengineering. Wilm Bettina,Tamburrini Riccardo,Orlando Giuseppe,Murray Patricia Current transplantation reports Worldwide, increasing numbers of patients are developing end-stage renal disease, and at present, the only treatment options are dialysis or kidney transplantation. Dialysis is associated with increased morbidity and mortality, poor life quality and high economic costs. Transplantation is by far the better option, but there are insufficient numbers of donor kidneys available. Therefore, there is an urgent need to explore alternative approaches. In this review, we discuss how this problem could potentially be addressed by using autologous cells and appropriate scaffolds to develop 'bioengineered' kidneys for transplantation. In particular, we will highlight recent breakthroughs in pluripotent stem cell biology that have led to the development of autologous renal progenitor cells capable of differentiating to all renal cell types and will discuss how these cells could be combined with appropriate scaffolds to develop a bioengineered kidney. 10.1007/s40472-016-0107-8
    Renal reabsorption in 3D vascularized proximal tubule models. Lin Neil Y C,Homan Kimberly A,Robinson Sanlin S,Kolesky David B,Duarte Nathan,Moisan Annie,Lewis Jennifer A Proceedings of the National Academy of Sciences of the United States of America Three-dimensional renal tissues that emulate the cellular composition, geometry, and function of native kidney tissue would enable fundamental studies of filtration and reabsorption. Here, we have created 3D vascularized proximal tubule models composed of adjacent conduits that are lined with confluent epithelium and endothelium, embedded in a permeable ECM, and independently addressed using a closed-loop perfusion system to investigate renal reabsorption. Our 3D kidney tissue allows for coculture of proximal tubule epithelium and vascular endothelium that exhibits active reabsorption via tubular-vascular exchange of solutes akin to native kidney tissue. Using this model, both albumin uptake and glucose reabsorption are quantified as a function of time. Epithelium-endothelium cross-talk is further studied by exposing proximal tubule cells to hyperglycemic conditions and monitoring endothelial cell dysfunction. This diseased state can be rescued by administering a glucose transport inhibitor. Our 3D kidney tissue provides a platform for in vitro studies of kidney function, disease modeling, and pharmacology. 10.1073/pnas.1815208116
    Patterned cell arrays and patterned co-cultures on polydopamine-modified poly(vinyl alcohol) hydrogels. Beckwith Kai M,Sikorski Pawel Biofabrication Live cell arrays are an emerging tool that expand traditional 2D in vitro cell culture, increasing experimental precision and throughput. A patterned cell system was developed by combining the cell-repellent properties of polyvinyl alcohol hydrogels with the cell adhesive properties of self-assembled films of dopamine (polydopamine). It was shown that polydopamine could be patterned onto spin-cast polyvinyl alcohol hydrogels by microcontact printing, which in turn effectively patterned the growth of several cell types (HeLa, human embryonic kidney, human umbilical vein endothelial cells (HUVEC) and prostate cancer). The cells could be patterned in geometries down to single-cell confinement, and it was demonstrated that cell patterns could be maintained for at least 3 weeks. Furthermore, polydopamine could be used to modify poly(vinyl alcohol) in situ using a cell-compatible deposition buffer (1 mg mL(-1) dopamine in 25 mM tris with a physiological salt balance). The treatment switched the PVA hydrogel from cell repellent to cell adhesive. Finally, by combining microcontact printing and in situ deposition of polydopamine, patterned co-cultures of the same cell type (HeLa/HeLa) and dissimilar cell types (HeLa/HUVEC) were realized through simple chemistry and could be studied over time. The combination of polyvinyl alcohol and polydopamine was shown to be an attractive route to versatile, patterned cell culture experiments with minimal infrastructure requirements and low complexity. 10.1088/1758-5082/5/4/045009
    Microcontact Peeling: A Cell Micropatterning Technique for Circumventing Direct Adsorption of Proteins to Hydrophobic PDMS. Yokoyama Sho,Matsui Tsubasa S,Deguchi Shinji Current protocols in cell biology Microcontact printing (μCPr) is one of the most popular techniques used for cell micropatterning. In conventional μCPr, a polydimethylsiloxane (PDMS) stamp with microfeatures is used to adsorb extracellular matrix (ECM) proteins onto the featured surface and transfer them onto particular areas of a cell culture substrate. However, some types of functional proteins other than ECM have been reported to denature upon direct adsorption to hydrophobic PDMS. Here we describe a detailed protocol of an alternative technique--microcontact peeling (μCPe)--that allows for cell micropatterning while circumventing the step of adsorbing proteins to bare PDMS. This technique employs microfeatured materials with a relatively high surface energy such as copper, instead of using a microfeatured PDMS stamp, to peel off a cell-adhesive layer present on the surface of substrates. Consequently, cell-nonadhesive substrates are exposed at the specific surface that undergoes the physical contact with the microfeatured material. Thus, although μCPe and μCPr are apparently similar, the former does not comprise a process of transferring biomolecules through hydrophobic PDMS. © 2017 by John Wiley & Sons, Inc. 10.1002/cpcb.22
    Extracellular Matrix in Kidney Fibrosis: More Than Just a Scaffold. Bülow Roman David,Boor Peter The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society Kidney fibrosis is the common histological end-point of progressive, chronic kidney diseases (CKDs) regardless of the underlying etiology. The hallmark of renal fibrosis, similar to all other organs, is pathological deposition of extracellular matrix (ECM). Renal ECM is a complex network of collagens, elastin, and several glycoproteins and proteoglycans forming basal membranes and interstitial space. Several ECM functions beyond providing a scaffold and organ stability are being increasingly recognized, for example, in inflammation. ECM composition is determined by the function of each of the histological compartments of the kidney, that is, glomeruli, tubulo-interstitium, and vessels. Renal ECM is a dynamic structure undergoing remodeling, particularly during fibrosis. From a clinical perspective, ECM proteins are directly involved in several rare renal diseases and indirectly in CKD progression during renal fibrosis. ECM proteins could serve as specific non-invasive biomarkers of fibrosis and scaffolds in regenerative medicine. The gold standard and currently only specific means to measure renal fibrosis is renal biopsy, but new diagnostic approaches are appearing. Here, we discuss the localization, function, and remodeling of major renal ECM components in healthy and diseased, fibrotic kidneys and the potential use of ECM in diagnostics of renal fibrosis and in tissue engineering. 10.1369/0022155419849388
    High-Resolution Patterned Cellular Constructs by Droplet-Based 3D Printing. Graham Alexander D,Olof Sam N,Burke Madeline J,Armstrong James P K,Mikhailova Ellina A,Nicholson James G,Box Stuart J,Szele Francis G,Perriman Adam W,Bayley Hagan Scientific reports Bioprinting is an emerging technique for the fabrication of living tissues that allows cells to be arranged in predetermined three-dimensional (3D) architectures. However, to date, there are limited examples of bioprinted constructs containing multiple cell types patterned at high-resolution. Here we present a low-cost process that employs 3D printing of aqueous droplets containing mammalian cells to produce robust, patterned constructs in oil, which were reproducibly transferred to culture medium. Human embryonic kidney (HEK) cells and ovine mesenchymal stem cells (oMSCs) were printed at tissue-relevant densities (10 cells mL) and a high droplet resolution of 1 nL. High-resolution 3D geometries were printed with features of ≤200 μm; these included an arborised cell junction, a diagonal-plane junction and an osteochondral interface. The printed cells showed high viability (90% on average) and HEK cells within the printed structures were shown to proliferate under culture conditions. Significantly, a five-week tissue engineering study demonstrated that printed oMSCs could be differentiated down the chondrogenic lineage to generate cartilage-like structures containing type II collagen. 10.1038/s41598-017-06358-x
    Impact of Three-dimensional Printing in Urology: State of the Art and Future Perspectives. A Systematic Review by ESUT-YAUWP Group. Cacciamani Giovanni E,Okhunov Zhamshid,Meneses Aurus Dourado,Rodriguez-Socarras Moises Elias,Rivas Juan Gomez,Porpiglia Francesco,Liatsikos Evangelos,Veneziano Domenico European urology CONTEXT:Three-dimensional (3D) printing has profoundly impacted biomedicine. It has been used to pattern cells; replicate tissues or full organs; create surgical replicas for planning, counseling, and training; and build medical device prototypes and prosthetics, and in numerous other applications. OBJECTIVE:To assess the impact of 3D printing for surgical planning, training and education, patient counseling, and costs in urology. EVIDENCE ACQUISITION:A systematic literature review was performed according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses statement. EVIDENCE SYNTHESIS:After screening, 4026 publications were identified for detailed review, of which 52 were included in the present systematic review: two papers reported the use of 3D-printing modeling for adrenal cancer, two papers for urethrovesical anastomosis, 24 papers for kidney transplantation and renal cancer, 13 papers for prostate cancer, seven papers for pelvicalyceal system procedures, and three papers for ureteral stents, and three papers reported 3D-printed biological scaffold development. CONCLUSIONS:Three-dimensional printing shows revolutionary potentials for patient counseling, pre- and intraoperative surgical planning, and education in urology. Together with the "patient-tailored" presurgical planning, it puts the basis for 3D-bioprinting technology. Although costs and "production times" remain the major concerns, this kind of technology may represent a step forward to meet patients' and surgeons' expectations. PATIENT SUMMARY:Three-dimensional printing has been used for several purposes to help the surgeon better understand anatomy, sharpen his/her skills, and guide the identification of lesions and their relationship with surrounding structures. It can be used for surgical planning, education, and patient counseling to improve the decision-making process. 10.1016/j.eururo.2019.04.044
    Solid organ fabrication: comparison of decellularization to 3D bioprinting. Jung Jangwook P,Bhuiyan Didarul B,Ogle Brenda M Biomaterials research Solid organ fabrication is an ultimate goal of Regenerative Medicine. Since the introduction of Tissue Engineering in 1993, functional biomaterials, stem cells, tunable microenvironments, and high-resolution imaging technologies have significantly advanced efforts to regenerate in vitro culture or tissue platforms. Relatively simple flat or tubular organs are already in (pre)clinical trials and a few commercial products are in market. The road to more complex, high demand, solid organs including heart, kidney and lung will require substantive technical advancement. Here, we consider two emerging technologies for solid organ fabrication. One is decellularization of cadaveric organs followed by repopulation with terminally differentiated or progenitor cells. The other is 3D bioprinting to deposit cell-laden bio-inks to attain complex tissue architecture. We reviewed the development and evolution of the two technologies and evaluated relative strengths needed to produce solid organs, with special emphasis on the heart and other tissues of the cardiovascular system. 10.1186/s40824-016-0074-2
    Corrigendum: Microtissues Enhance Smooth Muscle Differentiation and Cell Viability of hADSCs for Three Dimensional Bioprinting. Jin Yipeng,Xu Yongde,Wu Yuanyi,Sun Jilei,Guo Jiaxiang,Gao Jiangping,Yang Yong Frontiers in physiology [This corrects the article on p. 534 in vol. 8, PMID: 28790931.]. 10.3389/fphys.2017.00703
    Toward human organ printing: Charleston Bioprinting Symposium. Mironov Vladimir ASAIO journal (American Society for Artificial Internal Organs : 1992) The First Annual Charleston Bioprinting Symposium was organized by the Bioprinting Research Center of the Medical University of South Carolina (MUSC) and convened July 21, 2006, in Charleston, South Carolina. In broad terms, bioprinting is the application of rapid prototyping technology to the biomedical field. More specifically, it is defined as the layer by layer deposition of biologically relevant material. The 2006 Symposium included four sessions: Computer-aided design and Bioprinting, Bioprinting Technologies; Hydrogel for Bioprinting and, finally, a special session devoted to ongoing research projects at the MUSC Bioprinting Research Center. The Symposium highlight was the presentation of the multidisciplinary Charleston Bioengineered Kidney Project. This symposium demonstrated that bioprinting or robotic biofabrication is one of the most exciting and fast-emerging branches in the tissue engineering field. Robotic biofabrication will eventually lead to industrial production of living human organs suitable for clinical transplantation. The symposium demonstrated that although there are still many technological challenges, organ printing is a rapidly evolving feasible technology. 10.1097/01.mat.0000248999.25334.6a
    Three-dimensional printing of surgical anatomy. Powers Mary K,Lee Benjamin R,Silberstein Jonathan Current opinion in urology PURPOSE OF REVIEW:Over the past decade, three-dimensional printing for the medical field has been expanding rapidly throughout all of medicine. This manuscript reviews the current and potential applications for three-dimensional printing, including education, presurgical planning, surgical simulation, bioprinting, and printed surgical equipment. RECENT FINDINGS:Three-dimensional printing has proved most relevant in the fields of craniofacial, plastic, orthopedics, and especially, urologic surgery. This review focuses on several examples of how three-dimensional printing can be utilized, with emphasis on renal models for renal cell carcinoma, ureteral stents, and staghorn calculus. From an education standpoint, both patients and residents can benefit from the use of three-dimensional printed models, and even skilled surgeons report better understanding of complex procedures by using printed models. SUMMARY:Three-dimensional printing in the field of medicine is growing quickly, and will soon be incorporated into the way residents are taught and patients are educated. For surgical simulation in a variety of disease processes, this will be particularly useful for urologic surgery. 10.1097/MOU.0000000000000274
    Bioprinted three dimensional human tissues for toxicology and disease modeling. Nguyen Deborah G,Pentoney Stephen L Drug discovery today. Technologies The high rate of attrition among clinical-stage therapies, due largely to an inability to predict human toxicity and/or efficacy, underscores the need for in vitro models that better recapitulate in vivo human biology. In much the same way that additive manufacturing has revolutionized the production of solid objects, three-dimensional (3D) bioprinting is enabling the automated production of more architecturally and functionally accurate in vitro tissue culture models. Here, we provide an overview of the most commonly used bioprinting approaches and how they are being used to generate complex in vitro tissues for use in toxicology and disease modeling research. 10.1016/j.ddtec.2017.03.001
    Advanced Polymers for Three-Dimensional (3D) Organ Bioprinting. Wang Xiaohong Micromachines Three-dimensional (3D) organ bioprinting is an attractive scientific area with huge commercial profit, which could solve all the serious bottleneck problems for allograft transplantation, high-throughput drug screening, and pathological analysis. Integrating multiple heterogeneous adult cell types and/or stem cells along with other biomaterials (e.g., polymers, bioactive agents, or biomolecules) to make 3D constructs functional is one of the core issues for 3D bioprinting of bioartificial organs. Both natural and synthetic polymers play essential and ubiquitous roles for hierarchical vascular and neural network formation in 3D printed constructs based on their specific physical, chemical, biological, and physiological properties. In this article, several advanced polymers with excellent biocompatibility, biodegradability, 3D printability, and structural stability are reviewed. The challenges and perspectives of polymers for rapid manufacturing of complex organs, such as the liver, heart, kidney, lung, breast, and brain, are outlined. 10.3390/mi10120814
    3D-bioprinting a genetically inspired cartilage scaffold with GDF5-conjugated BMSC-laden hydrogel and polymer for cartilage repair. Sun Ye,You Yongqing,Jiang Wenbo,Zhai Zanjin,Dai Kerong Theranostics Articular cartilage injury is extremely common in congenital joint dysplasia patients. Genetic studies have identified Growth differentiation factor 5 (GDF5) as a shared gene in joint dysplasia and OA progression across different populations. However few studies have employed GDF5 in biological regeneration for articular cartilage repair. In the present study, we report identified genetic association between GDF5 loci and hip joint dysplasia with genome-wide association study (GWAS). GWAS and replication studies in separate populations achieved significant signals for GDF5 loci. GDF5 expression was dysregulated with allelic differences in hip cartilage of DDH and upregulated in the repaired cartilage in a rabbit cartilage defect model. GDF5 in vitro enhanced chondrogenesis and migration of bone marrow stem cells (BMSCs), GDF5 was tested in ectopic cartilage generation with BMSCs by GDF5 in nude mice in vivo. Genetically inspired, we further generated functional knee articular cartilage construct for cartilage repair by 3d-bioprinting a GDF5-conjugated BMSC-laden scaffold. GDF5-conjugated scaffold showed better cartilage repairing effects compared to control. Meanwhile, transplantation of the 3D-bioprinted GDF5-conjugated BMSC-laden scaffold in rabbit knees conferred long-term chondroprotection. In conclusion, we report identified genetic association between GDF5 and DDH with combined GWAS and replications, which further inspired us to generate a ready-to-implant GDF5-conjugated BMSC-laden scaffold with one-step 3d-bioprinting for cartilage repair. 10.7150/thno.38061
    Current applications of three-dimensional printing in urology. Chen Michael Y,Skewes Jacob,Desselle Mathilde,Wong Cynthia,Woodruff Maria A,Dasgupta Prokar,Rukin Nicholas J BJU international Three-dimensional (3D) printing or additive manufacturing is a new technology that has seen rapid development in recent years with decreasing costs. 3D printing allows the creation of customised, finely detailed constructs. Technological improvements, increased printer availability, decreasing costs, improved cell culture techniques, and biomaterials have enabled complex, novel and individualised medical treatments to be developed. Although the long-term goal of printing biocompatible organs has not yet been achieved, major advances have been made utilising 3D printing in biomedical engineering. In this literature review, we discuss the role of 3D printing in relation to urological surgery. We highlight the common printing methods employed and show examples of clinical urological uses. Currently, 3D printing can be used in urology for education of trainees and patients, surgical planning, creation of urological equipment, and bioprinting. In this review, we summarise the current applications of 3D-printing technology in these areas of urology. 10.1111/bju.14928
    High throughput miniature drug-screening platform using bioprinting technology. Rodríguez-Dévora Jorge I,Zhang Bimeng,Reyna Daniel,Shi Zhi-dong,Xu Tao Biofabrication In the pharmaceutical industry, new drugs are tested to find appropriate compounds for therapeutic purposes for contemporary diseases. Unfortunately, novel compounds emerge at expensive prices and current target evaluation processes have limited throughput, thus leading to an increase of cost and time for drug development. This work shows the development of the novel inkjet-based deposition method for assembling a miniature drug-screening platform, which can realistically and inexpensively evaluate biochemical reactions in a picoliter-scale volume at a high speed rate. As proof of concept, applying a modified Hewlett Packard model 5360 compact disc printer, green fluorescent protein expressing Escherichia coli cells along with alginate gel solution have been arrayed on a coverslip chip under a repeatable volume of 180% ± 26% picoliters per droplet; subsequently, different antibiotic droplets were patterned on the spots of cells to evaluate the inhibition of bacteria for antibiotic screening. The proposed platform was compared to the current screening process, validating its effectiveness. The viability and basic function of the printed cells were evaluated, resulting in cell viability above 98% and insignificant or no DNA damage to human kidney cells transfected. Based on the reduction of investment and compound volume used by this platform, this technique has the potential to improve the actual drug discovery process at its target evaluation stage. 10.1088/1758-5082/4/3/035001
    3D bioprinting of urethra with PCL/PLCL blend and dual autologous cells in fibrin hydrogel: An in vitro evaluation of biomimetic mechanical property and cell growth environment. Zhang Kaile,Fu Qiang,Yoo James,Chen Xiangxian,Chandra Prafulla,Mo Xiumei,Song Lujie,Atala Anthony,Zhao Weixin Acta biomaterialia OBJECTIVE:Urethral stricture is a common condition seen after urethral injury. The currently available treatments are inadequate and there is a scarcity of substitute materials used for treatment of urethral stricture. The traditional tissue engineering of urethra involves scaffold design, fabrication and processing of multiple cell types. METHODS:In this study, we have used 3D bioprinting technology to fabricate cell-laden urethra in vitro with different polymer types and structural characteristics. We hypothesized that use of PCL and PLCL polymers with a spiral scaffold design could mimic the structure and mechanical properties of natural urethra of rabbits, and cell-laden fibrin hydrogel could give a better microenvironment for cell growth. With using an integrated bioprinting system, tubular scaffold was formed with the biomaterials; meanwhile, urothelial cells (UCs) and smooth muscle cells (SMCs) were delivered evenly into inner and outer layers of the scaffold separately within the cell-laden hydrogel. RESULTS:The PCL/PLCL (50:50) spiral scaffold demonstrated mechanical properties equivalent to the native urethra in rabbit. Evaluation of the cell bioactivity in the bioprinted urethra revealed that UCs and SMCs maintained more than 80% viability even at 7days after printing. Both cell types also showed active proliferation and maintained the specific biomarkers in the cell-laden hydrogel. CONCLUSION:These results provided a foundation for further studies in 3D bioprinting of urethral constructs that mimic the natural urethral tissue in mechanical properties and cell bioactivity, as well a possibility of using the bioprinted construct for in vivo study of urethral implantation in animal model. SIGNIFICANCE OF STATEMENTS:The 3D bioprinting is a new technique to replace traditional tissue engineering. The present study is the first demonstration that it is feasible to create a urethral construct. Two kinds of biomaterials were used and achieved mechanical properties equivalent to that of native rabbit urethra. Bladder epithelial cells and smooth muscle cells were loaded in hydrogel and maintained sufficient viability and proliferation in the hydrogel. The highly porous scaffold could mimic a natural urethral base-membrane, and facilitate contacts between the printed epithelial cells and smooth muscle cells on both sides of the scaffold. These results provided a strong foundation for future studies on 3D bioprinted urethra. 10.1016/j.actbio.2016.12.008
    Microtissues Enhance Smooth Muscle Differentiation and Cell Viability of hADSCs for Three Dimensional Bioprinting. Yipeng Jin,Yongde Xu,Yuanyi Wu,Jilei Sun,Jiaxiang Guo,Jiangping Gao,Yong Yang Frontiers in physiology Smooth muscle differentiated human adipose derived stem cells (hADSCs) provide a crucial stem cell source for urinary tissue engineering, but the induction of hADSCs for smooth muscle differentiation still has several issues to overcome, including a relatively long induction time and equipment dependence, which limits access to abundant stem cells within a short period of time for further application. Three-dimensional (3D) bioprinting holds great promise in regenerative medicine due to its controllable construction of a designed 3D structure. When evenly mixed with bioink, stem cells can be spatially distributed within a bioprinted 3D structure, thus avoiding drawbacks such as, stem cell detachment in a conventional cell-scaffold strategy. Notwithstanding the advantages mentioned above, cell viability is often compromised during 3D bioprinting, which is often due to pressure during the bioprinting process. The objective of our study was to improve the efficiency of hADSC smooth muscle differentiation and cell viability of a 3D bioprinted structure. Here, we employed the hanging-drop method to generate hADSC microtissues in a smooth muscle inductive medium containing human transforming growth factor β1 and bioprinted the induced microtissues onto a 3D structure. After 3 days of smooth muscle induction, the expression of α-smooth muscle actin and smoothelin was higher in microtissues than in their counterpart monolayer cultured hADSCs, as confirmed by immunofluorescence and western blotting analysis. The semi-quantitative assay showed that the expression of α-smooth muscle actin (α-SMA) was 0.218 ± 0.077 in MTs and 0.082 ± 0.007 in Controls; smoothelin expression was 0.319 ± 0.02 in MTs and 0.178 ± 0.06 in Controls. Induced MTs maintained their phenotype after the bioprinting process. Live/dead and cell count kit 8 assays showed that cell viability and cell proliferation in the 3D structure printed with microtissues were higher at all time points compared to the conventional single-cell bioprinting strategy (mean cell viability was 88.16 ± 3.98 vs. 61.76 ± 15% for microtissues and single-cells, respectively). These results provide a novel way to enhance the smooth muscle differentiation of hADSCs and a simple method to maintain better cell viability in 3D bioprinting. 10.3389/fphys.2017.00534
    Future of 3D printing: How 3D bioprinting technology can revolutionize healthcare? Morris Steven Birth defects research Currently there are more than 2,000 children on the transplant waiting list-and more than 100,000 Americans nationwide-awaiting a matching organ. Most children aged one through 10 are awaiting a kidney, liver, or heart. As with any transplant, there are two ways to find an organ-someone can donate or someone can die. Unfortunately, the supply falls far short of the demand, leaving people to die every day waiting for a second chance at life. Scientific and medical experts, however, continue to develop promising technology like 3D bioprinting that could save thousands of lives without the need of a donor. 10.1002/bdr2.1351
    Bioprinting of 3D Convoluted Renal Proximal Tubules on Perfusable Chips. Homan Kimberly A,Kolesky David B,Skylar-Scott Mark A,Herrmann Jessica,Obuobi Humphrey,Moisan Annie,Lewis Jennifer A Scientific reports Three-dimensional models of kidney tissue that recapitulate human responses are needed for drug screening, disease modeling, and, ultimately, kidney organ engineering. Here, we report a bioprinting method for creating 3D human renal proximal tubules in vitro that are fully embedded within an extracellular matrix and housed in perfusable tissue chips, allowing them to be maintained for greater than two months. Their convoluted tubular architecture is circumscribed by proximal tubule epithelial cells and actively perfused through the open lumen. These engineered 3D proximal tubules on chip exhibit significantly enhanced epithelial morphology and functional properties relative to the same cells grown on 2D controls with or without perfusion. Upon introducing the nephrotoxin, Cyclosporine A, the epithelial barrier is disrupted in a dose-dependent manner. Our bioprinting method provides a new route for programmably fabricating advanced human kidney tissue models on demand. 10.1038/srep34845
    In vitro human tissues via multi-material 3-D bioprinting. Kolesky David B,Homan Kimberly A,Skylar-Scott Mark,Lewis Jennifer A Alternatives to laboratory animals : ATLA This paper highlights the foundational research on multi-material 3-D bioprinting of human tissues, for which the Lewis Bioprinting team at Harvard University was awarded the 2017 Lush Science Prize. The team's bioprinting platform enables the rapid fabrication of 3-D human tissues that contain all of the essential components found in their in vivo counterparts: cells, vasculature (or other tubular features) and extracellular matrix. The printed 3-D tissues are housed within a customised perfusion system and are subjected to controlled microphysiological environments over long durations (days to months). As exemplars, the team created a thick, stem cell-laden vascularised tissue that was controllably differentiated toward an osteogenic lineage in situ, and a 3-D kidney tissue that recapitulated the proximal tubule, a subunit of the nephron responsible for solute reabsorption. This highly versatile platform for manufacturing 3-D human tissue in vitro opens new avenues for replacing animal models used to develop next-generation therapies, test toxicity and study disease pathology. 10.1177/026119291804600404
    Applications of three-dimensional printing technology in urological practice. Youssef Ramy F,Spradling Kyle,Yoon Renai,Dolan Benjamin,Chamberlin Joshua,Okhunov Zhamshid,Clayman Ralph,Landman Jaime BJU international A rapid expansion in the medical applications of three-dimensional (3D)-printing technology has been seen in recent years. This technology is capable of manufacturing low-cost and customisable surgical devices, 3D models for use in preoperative planning and surgical education, and fabricated biomaterials. While several studies have suggested 3D printers may be a useful and cost-effective tool in urological practice, few studies are available that clearly demonstrate the clinical benefit of 3D-printed materials. Nevertheless, 3D-printing technology continues to advance rapidly and promises to play an increasingly larger role in the field of urology. Herein, we review the current urological applications of 3D printing and discuss the potential impact of 3D-printing technology on the future of urological practice. 10.1111/bju.13183
    Bioprinting of a functional vascularized mouse thyroid gland construct. Bulanova Elena A,Koudan Elizaveta V,Degosserie Jonathan,Heymans Charlotte,Pereira Frederico DAS,Parfenov Vladislav A,Sun Yi,Wang Qi,Akhmedova Suraya A,Sviridova Irina K,Sergeeva Natalia S,Frank Georgy A,Khesuani Yusef D,Pierreux Christophe E,Mironov Vladimir A Biofabrication Bioprinting can be defined as additive biofabrication of three-dimensional (3D) tissues and organ constructs using tissue spheroids, capable of self-assembly, as building blocks. The thyroid gland, a relatively simple endocrine organ, is suitable for testing the proposed bioprinting technology. Here we report the bioprinting of a functional vascularized mouse thyroid gland construct from embryonic tissue spheroids as a proof of concept. Based on the self-assembly principle, we generated thyroid tissue starting from thyroid spheroids (TS) and allantoic spheroids (AS) as a source of thyrocytes and endothelial cells (EC), respectively. Inspired by mathematical modeling of spheroid fusion, we used an original 3D bioprinter to print TS in close association with AS within a collagen hydrogel. During the culture, closely placed embryonic tissue spheroids fused into a single integral construct, EC from AS invaded and vascularized TS, and epithelial cells from the TS progressively formed follicles. In this experimental setting, we observed formation of a capillary network around follicular cells, as observed during in utero thyroid development when thyroid epithelium controls the recruitment, invasion and expansion of EC around follicles. To prove that EC from AS are responsible for vascularization of the thyroid gland construct, we depleted endogenous EC from TS before bioprinting. EC from AS completely revascularized depleted thyroid tissue. The cultured bioprinted construct was functional as it could normalize blood thyroxine levels and body temperature after grafting under the kidney capsule of hypothyroid mice. Bioprinting of functional vascularized mouse thyroid gland construct represents a further advance in bioprinting technology, exploring the self-assembling properties of tissue spheroids. 10.1088/1758-5090/aa7fdd
    Bioprinting Technologies in Tissue Engineering. Yilmaz Bengi,Tahmasebifar Aydin,Baran Erkan Türker Advances in biochemical engineering/biotechnology Bioprinting technology is a strong tool in producing living functional tissues and organs from cells, biomaterial-based bioinks, and growth factors in computer-controlled platform. The aim of this chapter is to present recent progresses in bioprinting of nerve, skin, cardiac, bone, cartilage, skeletal muscle, and other soft tissues and highlight the challenges in these applications. Various composite bioinks with bioactive ceramic-based scaffolds having patient-specific design and controlled micro-architectures were used at clinical and preclinical applications successfully for regeneration of bone. In nerve tissue engineering, bioprinting of alginate- and gelatin-based gel bioinks by extrusion presented a controllable 3D microstructures and showed satisfactory cytocompatibility and axonal regeneration. Bioprinting of cardiac progenitors in biopolymers resulted in limited success, while the use of bioinks from extracellular matrix induced satisfactory results in cardiac regeneration. Osteochondral scaffold bioprinting is challenging due to the complex hierarchical structure and limited chondral regeneration. Therefore, current approaches focused on osteochondral scaffold with vascular network and mimicking hierarchical structures. The applications of bioprinting in other types of tissues were also studied, and results showed significant potentials in regeneration of tissues such as cornea, liver, and urinary bladder. 10.1007/10_2019_108
    Biofabricated Structures Reconstruct Functional Urinary Bladders in Radiation-Injured Rat Bladders. Imamura Tetsuya,Shimamura Mitsuru,Ogawa Teruyuki,Minagawa Tomonori,Nagai Takashi,Silwal Gautam Sudha,Ishizuka Osamu Tissue engineering. Part A The ability to repair damaged urinary bladders through the application of bone marrow-derived cells is in the earliest stages of development. We investigated the application of bone marrow-derived cells to repair radiation-injured bladders. We used a three-dimensional bioprinting robot system to biofabricate bone marrow-derived cell structures. We then determined if the biofabricated structures could restore the tissues and functions of radiation-injured bladders. The bladders of female 10-week-old Sprague-Dawley (SD) rats were irradiated with 2-Gy once a week for 5 weeks. Adherent and proliferating bone marrow-derived cells harvested from the femurs of male 17-week-old green fluorescence protein-transfected Tg-SD rats were cultured in collagen-coated flasks. Bone marrow-derived cell spheroids were formed in 96-well plates. Three layers of spheroids were assembled by the bioprinter onto a 9 × 9 microneedle array. The assembled spheroids were perfusion cultured for 7 days, and then the microneedle array was removed. Two weeks after the last radiation treatment, the biofabricated structures were transplanted into an incision on the anterior wall of the bladders (n = 10). Control rats received the same surgery but without the biofabricated structures (sham-structure, n = 12). At 2 and 4 weeks after surgery, the sham-structure control bladder tissues exhibited disorganized smooth muscle layers, decreased nerve cells, and significant fibrosis with increased presence of fibrosis-marker P4HB-positive cells and hypoxia-marker hypoxia-induced factor 1α (HIF1α)-positive cells. The transplanted structures survived within the recipient tissues, and blood vessels extended within them from the recipient tissues. The bone marrow-derived cells in the structures differentiated into smooth muscle cells and formed smooth muscle clusters. The recipient tissues near the transplanted structures had distinct smooth muscle layers and reconstructed nerve cells, and only minimal fibrosis with decreased presence of P4HB- and HIF1α-positive cells. At 4 weeks after surgery, the sham-structure control rats exhibited significant urinary frequency symptoms with irregular and short voiding intervals, and low micturition volumes. In contrast, the structure-transplanted rats had regular micturition with longer voiding intervals and higher micturition volumes compared with the control rats. Furthermore, the residual volume of the structure-transplanted rats was lower than for the controls. Therefore, transplantation of biofabricated bone marrow-derived cell structures reconstructed functional bladders. 10.1089/ten.TEA.2017.0533
    Digitally Tunable Microfluidic Bioprinting of Multilayered Cannular Tissues. Pi Qingmeng,Maharjan Sushila,Yan Xiang,Liu Xiao,Singh Bijay,van Genderen Anne Metje,Robledo-Padilla Felipe,Parra-Saldivar Roberto,Hu Ning,Jia Weitao,Xu Changliang,Kang Jian,Hassan Shabir,Cheng Haibo,Hou Xu,Khademhosseini Ali,Zhang Yu Shrike Advanced materials (Deerfield Beach, Fla.) Despite advances in the bioprinting technology, biofabrication of circumferentially multilayered tubular tissues or organs with cellular heterogeneity, such as blood vessels, trachea, intestine, colon, ureter, and urethra, remains a challenge. Herein, a promising multichannel coaxial extrusion system (MCCES) for microfluidic bioprinting of circumferentially multilayered tubular tissues in a single step, using customized bioinks constituting gelatin methacryloyl, alginate, and eight-arm poly(ethylene glycol) acrylate with a tripentaerythritol core, is presented. These perfusable cannular constructs can be continuously tuned up from monolayer to triple layers at regular intervals across the length of a bioprinted tube. Using customized bioink and MCCES, bioprinting of several tubular tissue constructs using relevant cell types with adequate biofunctionality including cell viability, proliferation, and differentiation is demonstrated. Specifically, cannular urothelial tissue constructs are bioprinted, using human urothelial cells and human bladder smooth muscle cells, as well as vascular tissue constructs, using human umbilical vein endothelial cells and human smooth muscle cells. These bioprinted cannular tissues can be actively perfused with fluids and nutrients to promote growth and proliferation of the embedded cell types. The fabrication of such tunable and perfusable circumferentially multilayered tissues represents a fundamental step toward creating human cannular tissues. 10.1002/adma.201706913