Publications by authors named "James J Yoo"

220 Publications

Combinations of photoinitiator and UV absorber for cell-based digital light processing (DLP) bioprinting.

Biofabrication 2021 Apr 30. Epub 2021 Apr 30.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, North Carolina, 27157, UNITED STATES.

Digital light processing (DLP) bioprinting, which provides predominant speed, resolution, and adaptability for fabricating complex cell-laden 3D structures, requires a combination of photoinitiator (PI) and UV absorber (UA) that plays critical roles during the photo-polymerization of bioinks. However, the PI and UA combination has not been highlighted for cell-based DLP bioprinting. In this study, the most used PIs and UAs in cell-based bioprinting were compared to optimize a combination that can ensure the maximum DLP printability, while maintaining the cellular activities during the process. The crosslinking time and printability of PIs were assessed, which are critical in minimizing the cell damage by the UV exposure during the fabrication process. On the other hand, the UAs were evaluated based on their ability to prevent the over-curing of layers beyond the focal layer and the scattering of light, which are required for the desirable crosslinking of a hydrogel and high resolution (25-50 microns) to create a complex 3D cell-laden construct. Lastly, the cytotoxicity of PIs and UAs was assessed by measuring the cellular activity of 2D cultured and 3D bioprinted cells. The optimized PI and UA combination provided high initial cell viability (> 90%) for up to 14 days in culture and could fabricate complex 3D structures like a perfusable heart-shaped construct with open vesicles and atriums. This combination can provide a potential starting condition when preparing the bioink for the cell-based DLP bioprinting in tissue engineering applications.
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http://dx.doi.org/10.1088/1758-5090/abfd7aDOI Listing
April 2021

Regenerative Medicine Approaches in Bioengineering Female Reproductive Tissues.

Reprod Sci 2021 Apr 20. Epub 2021 Apr 20.

Wake Forest Institute for Regenerative Medicine, Wake Forest University School of Medicine, Winston-Salem, NC, 27101, USA.

Diseases, disorders, and dysfunctions of the female reproductive tract tissues can result in either infertility and/or hormonal imbalance. Current treatment options are limited and often do not result in tissue function restoration, requiring alternative therapeutic approaches. Regenerative medicine offers potential new therapies through the bioengineering of female reproductive tissues. This review focuses on some of the current technologies that could address the restoration of functional female reproductive tissues, including the use of stem cells, biomaterial scaffolds, bio-printing, and bio-fabrication of tissues or organoids. The use of these approaches could also be used to address issues in infertility. Strategies such as cell-based hormone replacement therapy could provide a more natural means of restoring normal ovarian physiology. Engineering of reproductive tissues and organs could serve as a powerful tool for correcting developmental anomalies. Organ-on-a-chip technologies could be used to perform drug screening for personalized medicine approaches and scientific investigations of the complex physiological interactions between the female reproductive tissues and other organ systems. While some of these technologies have already been developed, others have not been translated for clinical application. The continuous evolution of biomaterials and techniques, advances in bioprinting, along with emerging ideas for new approaches, shows a promising future for treating female reproductive tract-related disorders and dysfunctions.
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http://dx.doi.org/10.1007/s43032-021-00548-9DOI Listing
April 2021

Optimized culture system to maximize ovarian cell growth and functionality in vitro.

Cell Tissue Res 2021 Feb 13. Epub 2021 Feb 13.

Wake Forest Institute for Regernative Medicine, Wake Forest School of Medicine, Medical Center Boulvard, Winston Salem, NC, 27157, USA.

Ovaries are the primary physiological source of female sex hormones, which play a crucial role in maintaining ovarian cycle, determining secondary sexual characteristics and preparing the endometrium for implantation. In vitro follicle engineering has been used to investigate follicle development, including ovarian hormone production and gamete maturation. To engineer functional follicles, culture and expansion of the primary ovarian cells are essential. However, the phenotypic and functional characteristics of primary ovarian cells are often lost during culture. The objective of this study is to develop an optimized culture system for maintaining ovarian cell growth and functionality. Granulosa cells (GCs) and theca cells (TCs) were isolated from female rats. The addition of follicle-stimulating hormone (FSH) or luteinizing hormone (LH) to the basal culture media significantly enhanced the secretion of estradiol from GCs and androstenedione from TCs. Serum concentrations of 5% and 10% had a similar role in promoting ovarian cell expansion and secretion of estradiol and androstenedione hormones from both types of cells. Growth differentiation factor 9 (GDF9), bone morphogenic protein 15 (BMP15), BMP7 and basic fibroblast growth factor (bFGF) enhanced GC proliferation and estradiol production, respectively. Among them, the effect of bFGF was most significant. bFGF also enhanced TC proliferation. When GCs and TCs were cultured in 5% serum, gonadotropin and bFGF-containing medium, they proliferated exponentially throughout the culture period of up to 40 days while maintaining their functional characteristics. Taken together, these results indicate that our medium formula is optimal for maximizing proliferation of functionally differentiated ovarian cells.
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http://dx.doi.org/10.1007/s00441-021-03415-wDOI Listing
February 2021

Accelerating neovascularization and kidney tissue formation with a 3D vascular scaffold capturing native vascular structure.

Acta Biomater 2021 04 30;124:233-243. Epub 2021 Jan 30.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, NC, USA. Electronic address:

Establishing an adequate vascularization of three-dimensional (3D) bioengineered tissues remains a critical challenge. We previously fabricated a vascular scaffold using the vascular corrosion casting technique, which provides a similar 3D geometry of native kidney vasculature. In this study, we functionalized the collagen vascular scaffold with a controlled release of vascular endothelial growth factor (VEGF vascular scaffold) to further promote vascularization. The VEGF vascular scaffold showed improved angiogenic capability in 2-dimensional (2D) and 3D in vitro settings. Implantation of the VEGF vascular scaffold seeded with human renal cells into a rat kidney demonstrated enhanced implant vascularization and reduced apoptosis of implanted human renal cells. Hybrid renal tubule-like structures composed of implanted human and migrated host renal cells were formed. This work highlights the critical role of early vascularization of the geometrically mimetic vascular scaffold using the VEGF incorporated vascular scaffold in reducing apoptosis of implanted cells as well as the formation of renal tissue structures.
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http://dx.doi.org/10.1016/j.actbio.2021.01.040DOI Listing
April 2021

Engineering Functional Rat Ovarian Spheroids Using Granulosa and Theca Cells.

Reprod Sci 2021 Jan 28. Epub 2021 Jan 28.

Wake Forest Institute for Regenerative Medicine, Wake Forest University School of Medicine, Winston-Salem, NC, 27101, USA.

Although menopausal hormone therapy (MHT) is the most effective approach to managing the loss of ovarian activity, serious side effects have been reported. Cell-based therapy is a promising alternative for MHT. This study constructed engineered ovarian cell spheroids and investigated their endocrine function. Theca and granulosa cells were isolated from ovaries of 10-week-old rats. Two types of engineered ovarian cell spheroids were fabricated through forced aggregation in microwells, multilayered spheroids with centralized granulosa aggregates surrounded by an outer layer of theca cells and mixed ovarian spheroids lacking spatial rearrangement. The ovarian cell spheroids were encapsulated into a collagen gel. Non-aggregated ovarian cells served as controls. The endocrine function of the engineered ovarian spheroids was assessed over 30 days. The structure of the spheroids was well maintained during culture. The secretion of 17β-estradiol from both types of engineered ovarian cell spheroids was higher than in the control group and increased continuously in a time-dependent manner. Secretion of 17β-estradiol in the multi-layered ovarian cell spheroids was higher than in the non-layered constructs. Increased secretion of progesterone was detected in the multi-layered ovarian cell spheroids at day 5 of culture and was sustained during the culture period. The initial secretion level of progesterone in the non-layered ovarian cell spheroids was similar to those from the controls and increased significantly from days 21 to 30. An in vitro rat model of engineered ovarian cell spheroids was developed that was capable of secreting sex steroid hormones, indicating that the hormone secreting function of ovaries can be recapitulated ex vivo and potentially adapted for MHT.
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http://dx.doi.org/10.1007/s43032-020-00445-7DOI Listing
January 2021

Pelvic floor muscle function recovery using biofabricated tissue constructs with neuromuscular junctions.

Acta Biomater 2021 02 13;121:237-249. Epub 2020 Dec 13.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157-1094, USA. Electronic address:

Damages in pelvic floor muscles often cause dysfunction of the entire pelvic urogenital system, which is clinically challenging. A bioengineered skeletal muscle construct that mimics structural and functional characteristics of native skeletal muscle could provide a therapeutic option to restore normal muscle function. However, most of the current bioengineered muscle constructs are unable to provide timely innervation necessary for successful grafting and functional recovery. We previously have demonstrated that post-synaptic acetylcholine receptors (AChR) clusters can be pre-formed on cultured skeletal muscle myofibers with agrin treatment and suggested that implantation of AChR clusters containing myofibers could accelerate innervation and recovery of muscle function. In this study, we develop a 3-dimensional (3D) bioprinted human skeletal muscle construct, consisting of multi-layers bundles with aligned and AChR clusters pre-formed human myofibers, and investigate the effect of pre-formed AChR clusters in bioprinted skeletal muscle constructs and innervation efficiency in vivo. Agrin treatment successfully pre-formed functional AChR clusters on the bioprinted muscle constructs in vitro that increased neuromuscular junction (NMJ) formation in vivo in a transposed nerve implantation model in rats. In a rat model of pelvic floor muscle injury, implantation of skeletal muscle constructs containing the pre-formed AChR clusters resulted in functional muscle reconstruction with accelerated construct innervation. This approach may provide a therapeutic solution to the many challenges associated with pelvic floor reconstruction resulting from the lack of suitable bioengineered tissue for efficient innervation and muscle function restoration.
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http://dx.doi.org/10.1016/j.actbio.2020.12.012DOI Listing
February 2021

Microfluidic Systems for Assisted Reproductive Technologies: Advantages and Potential Applications.

Tissue Eng Regen Med 2020 12 25;17(6):787-800. Epub 2020 Nov 25.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC, 27157, USA.

Microfluidic technologies have emerged as a powerful tool that can closely replicate the in-vivo physiological conditions of organ systems. Assisted reproductive technology (ART), while being able to achieve successful outcomes, still faces challenges related to technical error, efficiency, cost, and monitoring/assessment. In this review, we provide a brief overview of the uses of microfluidic devices in the culture, maintenance and study of ovarian follicle development for experimental and therapeutic applications. We discuss existing microfluidic platforms for oocyte and sperm selection and maintenance, facilitation of fertilization by in-vitro fertilization/intracytoplastimc sperm injection, and monitoring, selection and maintenance of resulting embryos. Furthermore, we discuss the possibility of future integration of these technologies onto a single platform and the limitations facing the development of these systems. In spite of these challenges, we envision that microfluidic systems will likely evolve and inevitably revolutionize both fundamental, reproductive physiology/toxicology research as well as clinically applicable ART.
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http://dx.doi.org/10.1007/s13770-020-00311-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7710813PMC
December 2020

A photo-crosslinkable cartilage-derived extracellular matrix bioink for auricular cartilage tissue engineering.

Acta Biomater 2021 02 21;121:193-203. Epub 2020 Nov 21.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157, USA. Electronic address:

Three-dimensional (3D) bioprinting of patient-specific auricular cartilage constructs could aid in the reconstruction process of traumatically injured or congenitally deformed ear cartilage. To achieve this, a hydrogel-based bioink is required that recapitulates the complex cartilage microenvironment. Tissue-derived decellularized extracellular matrix (dECM)-based hydrogels have been used as bioinks for cell-based 3D bioprinting because they contain tissue-specific ECM components that play a vital role in cell adhesion, growth, and differentiation. In this study, porcine auricular cartilage tissues were isolated and decellularized, and the decellularized cartilage tissues were characterized by histology, biochemical assay, and proteomics. This cartilage-derived dECM (cdECM) was subsequently processed into a photo-crosslinkable hydrogel using methacrylation (cdECMMA) and mixed with chondrocytes to create a printable bioink. The rheological properties, printability, and in vitro biological properties of the cdECMMA bioink were examined. The results showed cdECM was obtained with complete removal of cellular components while preserving major ECM proteins. After methacrylation, the cdECMMA bioinks were printed in anatomical ear shape and exhibited adequate mechanical properties and structural integrity. Specifically, auricular chondrocytes in the printed cdECMMA hydrogel constructs maintained their viability and proliferation capacity and eventually produced cartilage ECM components, including collagen and glycosaminoglycans (GAGs). The potential of cell-based bioprinting using this cartilage-specific dECMMA bioink is demonstrated as an alternative option for auricular cartilage reconstruction.
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http://dx.doi.org/10.1016/j.actbio.2020.11.029DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7855948PMC
February 2021

Applications of Organoids for Tissue Engineering and Regenerative Medicine.

Tissue Eng Regen Med 2020 12 12;17(6):729-730. Epub 2020 Nov 12.

Department of Molecular Medicine, Ewha Womans University Medical College, Seoul, 07804, Republic of Korea.

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http://dx.doi.org/10.1007/s13770-020-00315-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7710832PMC
December 2020

Synergistic effect of CNTF and GDNF on directed neurite growth in chick embryo dorsal root ganglia.

PLoS One 2020 5;15(10):e0240235. Epub 2020 Oct 5.

Wake Forest Institute for Regenerative Medicine, Winston Salem, NC, United States of America.

It is often critical to improve the limited regenerative capacity of the peripheral nerves and direct neural growth towards specific targets, such as surgically implanted bioengineered constructs. One approach to accomplish this goal is to use extrinsic neurotrophic factors. The candidate factors first need to be identified and characterized in in vitro tests for their ability to direct the neurite growth. Here, we present a simple guidance assay that allows to assess the chemotactic effect of signaling molecules on the growth of neuronal processes from dorsal root ganglia (DRG) using only standard tissue culture materials. We used this technique to quantitatively determine the combined and individual effects of the ciliary neurotrophic factor (CNTF) and glial cell line-derived neurotrophic factor (GDNF) on neurite outgrowth. We demonstrated that these two neurotrophic factors, when applied in a 1:1 combination, but not individually, induced directed growth of neuronal processes towards the source of the gradient. This chemotactic effect persists without significant changes over a wide (10-fold) concentration range. Moreover, we demonstrated that other, more general growth parameters that do not evaluate growth in a specific direction (such as, neurite length and trajectory) were differentially affected by the concentration of the CNTF/GNDF mixture. Furthermore, GDNF, when applied individually, did not have any chemotactic effect, but caused significant neurite elongation and an increase in the number of neurites per ganglion.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0240235PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7535060PMC
December 2020

The Influence of Printing Parameters and Cell Density on Bioink Printing Outcomes.

Tissue Eng Part A 2020 Dec 14;26(23-24):1349-1358. Epub 2020 Oct 14.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA.

Bioink printability persists as a limiting factor toward many bioprinting applications. Printing parameter selection is largely user-dependent, and the effect of cell density on printability has not been thoroughly investigated. Recently, methods have been developed to give greater insight into printing outcomes. This study aims to further advance those methods and apply them to study the effect of printing parameters (feedrate and flowrate) and cell density on printability. Two printed structures, a crosshatch and five-layer tube, were established as printing standards and utilized to determine the printing outcomes. Acellular bioinks were printed using a testing matrix of feedrates of 37.5, 75, 150, 300, and 600 mm/min and flowrates of 21, 42, 84, 168, and 336 mm/min. Structures were also printed with cell densities of 5, 10, 20, and 40 × 10 cell/mL at 150 mm/min and 84 mm/min. Only speed ratios (defined as flowrate divided by feedrate) from 0.07 to 2.24 mm were suitable for analysis. Increasing speed ratio dramatically increased the height, width, and wall thickness of tubular structures, but did not influence radial accuracy. For crosshatch structures, the area of pores and the frequency of broken filaments were decreased without impacting pore shape (). Within speed ratios, feedrate and flowrate had negligible, inconsistent effects. Cell density did not affect any printing outcomes despite slight rheological changes. Printing outcomes were dominated by the speed ratio, with feedrate, flowrate, and cell density having little impact on printing outcomes when controlling for speed ratio within the ranges tested. The relevance of these results to other bioinks and printing conditions requires continued investigation by the bioprinting community, as well as highlight speed ratio as a key variable to report and suggest that rheology is a more sensitive measure than printing outcomes.
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http://dx.doi.org/10.1089/ten.TEA.2020.0210DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7780841PMC
December 2020

Solid Organ Bioprinting: Strategies to Achieve Organ Function.

Chem Rev 2020 Oct 4;120(19):11093-11127. Epub 2020 Sep 4.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Richard H. Dean Biomedical Building, 391 Technology Way, Winston-Salem, North Carolina 27101, United States.

The field of tissue engineering has advanced over the past decade, but the largest impact on human health should be achieved with the transition of engineered solid organs to the clinic. The number of patients suffering from solid organ disease continues to increase, with over 100 000 patients on the U.S. national waitlist and approximately 730 000 deaths in the United States resulting from end-stage organ disease annually. While flat, tubular, and hollow nontubular engineered organs have already been implanted in patients, in vitro formation of a fully functional solid organ at a translatable scale has not yet been achieved. Thus, one major goal is to bioengineer complex, solid organs for transplantation, composed of patient-specific cells. Among the myriad of approaches attempted to engineer solid organs, 3D bioprinting offers unmatched potential. This review highlights the structural complexity which must be engineered at nano-, micro-, and mesostructural scales to enable organ function. We showcase key advances in bioprinting solid organs with complex vascular networks and functioning microstructures, advances in biomaterials science that have enabled this progress, the regulatory hurdles the field has yet to overcome, and cutting edge technologies that bring us closer to the promise of engineered solid organs.
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http://dx.doi.org/10.1021/acs.chemrev.0c00145DOI Listing
October 2020

NIR fluorescence for monitoring in vivo scaffold degradation along with stem cell tracking in bone tissue engineering.

Biomaterials 2020 11 6;258:120267. Epub 2020 Aug 6.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC, 27157, USA. Electronic address:

Stem cell-based tissue engineering has the potential to use as an alternative for autologous tissue grafts; however, the contribution of the scaffold degradation along with the transplanted stem cells to in vivo tissue regeneration remains poorly understood. Near-infrared (NIR) fluorescence imaging has great potential to monitor implants while avoiding autofluorescence from the adjacent host tissue. To utilize NIR imaging for in vivo monitoring of scaffold degradation and cell tracking, we synthesized 800-nm emitting NIR-conjugated PCL-ran-PLLA-ran-PGA (ZW-PCLG) copolymers with three different degradation rates and labeled 700-nm emitting lipophilic pentamethine (CTNF127) on the human placental stem cells (CT-PSCs). The 3D bioprinted hybrid constructs containing the CT-PSC-laden hydrogel together with the ZW-PCLG scaffolds demonstrate that NIR fluorescent imaging enables tracking of in vivo scaffold degradation and stem cell fate for bone regeneration in a rat calvarial bone defect model. This NIR-based monitoring system can be effectively utilized to study cell-based tissue engineering applications.
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http://dx.doi.org/10.1016/j.biomaterials.2020.120267DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7484145PMC
November 2020

Decellularized Skin Extracellular Matrix (dsECM) Improves the Physical and Biological Properties of Fibrinogen Hydrogel for Skin Bioprinting Applications.

Nanomaterials (Basel) 2020 Jul 29;10(8). Epub 2020 Jul 29.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157, USA.

Full-thickness skin wounds are a significant clinical burden in the United States. Skin bioprinting is a relatively new technology that is under investigation as a new treatment for full-thickness injuries, and development of hydrogels with strong physical and biological characteristics are required to improve both structural integrity of the printed constructs while allowing for a more normal extracellular matrix milieu. This project aims to evaluate the physical and biological characteristics of fibrinogen hydrogel supplemented with decellularized human skin-derived extracellular matrix (dsECM). The hybrid hydrogel improves the cell viability and structural strength of bioprinted skin constructs. Scanning electron microscopy demonstrates that the hybrid hydrogel is composed of both swelling bundles interlocked in a fibrin network, similar to healthy human skin. This hybrid hydrogel has improved rheological properties and shear thinning properties. Extrusion-based printing of the fibrinogen hydrogel + dsECM demonstrates significant improvement in crosshatch pore size. These findings suggest that incorporating the properties of dsECM and fibrinogen hydrogels will improve in vivo integration of the bioprinted skin constructs and support of healthy skin wound regeneration.
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http://dx.doi.org/10.3390/nano10081484DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7466410PMC
July 2020

Self-Assembling Peptide Solution Accelerates Hemostasis.

Adv Wound Care (New Rochelle) 2021 04 10;10(4):191-203. Epub 2020 Sep 10.

Bio-Materials and Technology Lab, Grain Science and Industry, Kansas State University, Manhattan, Kansas, USA.

One of the leading causes of death following traumatic injury is exsanguination. Biological material-based hemostatic agents such as fibrin, thrombin, and albumin have a high risk for causing infection. Synthetic peptide-based hemostatic agents offer an attractive alternative. The objective of this study is to explore the potential of h9e peptide as an effective hemostatic agent in both and models. blood coagulation kinetics in the presence of h9e peptide was determined as a function of gelation time using a dynamic rheometer. hemostatic effects were studied using the Wistar rat model. Results were compared to those of the commercial hemostatic product Celox™, a chitosan-based product. Adhesion of h9e peptide was evaluated using the platelet adhesion test. Biocompatibility of h9e peptide was studied using a mouse model. After h9e peptide solution was mixed with blood, gelation started immediately, increased rapidly with time, and reached more than 100 Pa within 3 s. Blood coagulation strength increased as h9e peptide wt% concentration increased. In the rat model, h9e peptide solution at 5% weight concentration significantly reduced both bleeding time and blood loss, outperforming Celox. Preliminary pathological studies indicate that h9e peptide solution is biocompatible and did not have negative effects when injected subcutaneously in a mouse model. For the first time, h9e peptide was found to have highly efficient hemostatic effects by forming nanoweb-like structures, which act as a preliminary thrombus and a surface to arrest bleeding 82% faster compared to the commercial hemostatic agent Celox. This study demonstrates that h9e peptide is a promising hemostatic biomaterial, not only because of its greater hemostatic effect than commercial product Celox but also because of its excellent biocompatibility based on the mouse model study.
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http://dx.doi.org/10.1089/wound.2019.1109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7906870PMC
April 2021

Administration of secretome from human placental stem cell-conditioned media improves recovery of erectile function in the pelvic neurovascular injury model.

J Tissue Eng Regen Med 2020 10 31;14(10):1394-1402. Epub 2020 Jul 31.

Wake Forest Institute for Regenerative Medicine, Winston-Salem, NC, USA.

Human placental stem cells (PSCs) enhance histological and functional recovery in a rodent erectile dysfunction (ED) model. We tested the hypothesis that bioactive factors secreted by PSC (i.e., the secretome) mediate functional recovery and that acellular-conditioned media (CM) from PSC culture (PSC-CM) could be used independently to facilitate functional and histological recovery. To identify factors relative to efficacy of PSC, a comparison of CM from PSC and three additional human stem cell populations was performed. CM from human PSC, amniotic fluid stem cells (AFSCs), adipose-derived stem cells (ADSC), and human umbilical vein endothelial cells (HUVECs) was assayed using a semi-quantitative human cytokine antibody array. Male rats, after surgically created ED by neurovascular injury, were randomly divided into four groups: vehicle control (phosphate-buffered saline [PBS]), PSC, PSC-CM, and serum-free media control (SFM) as control. Functional data on intracorporal and mean arterial pressure were obtained, and histological architecture was examined 6 weeks after single injection. PSCs were found to secrete at least 27 cytokines and growth factors at a significantly higher level than the other three cell types. Either single injection of PSC-CM or PSC significantly improved erectile functional recovery and histological architecture compared with SFM or PBS. Injection of the secretome isolated from human PSC improves erectile functional recovery and histological structure in a rat model of neurovascular injury-induced ED. Further characterization of the unique protein expression within the PSC-CM may help to identify the potential for a novel injectable cell-free therapeutic for applicable patients.
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http://dx.doi.org/10.1002/term.3105DOI Listing
October 2020

A tissue-engineered uterus supports live births in rabbits.

Nat Biotechnol 2020 11 29;38(11):1280-1287. Epub 2020 Jun 29.

Wake Forest Institute for Regenerative Medicine, Wake Forest University School of Medicine, Winston-Salem, NC, USA.

Bioengineered uterine tissue could provide a treatment option for women with uterine factor infertility. In large animal models, reconstruction of the uterus has been demonstrated only with xenogeneic tissue grafts. Here we use biodegradable polymer scaffolds seeded with autologous cells to restore uterine structure and function in rabbits. Rabbits underwent a subtotal uterine excision and were reconstructed with autologous cell-seeded constructs, with nonseeded scaffolds or by suturing. At 6 months postimplantation, only the cell-seeded engineered uteri developed native tissue-like structures, including organized luminal/glandular epithelium, stroma, vascularized mucosa and two-layered myometrium. Only rabbits with cell-seeded constructs had normal pregnancies (four in ten) in the reconstructed segment of the uterus and supported fetal development to term and live birth. With further development, this approach may provide a regenerative medicine solution to uterine factor infertility.
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http://dx.doi.org/10.1038/s41587-020-0547-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7641977PMC
November 2020

The Role of the Microenvironment in Controlling the Fate of Bioprinted Stem Cells.

Chem Rev 2020 Oct 19;120(19):11056-11092. Epub 2020 Jun 19.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina 27157, United States.

The field of tissue engineering and regenerative medicine has made numerous advances in recent years in the arena of fabricating multifunctional, three-dimensional (3D) tissue constructs. This can be attributed to novel approaches in the bioprinting of stem cells. There are expansive options in bioprinting technology that have become more refined and specialized over the years, and stem cells address many limitations in cell source, expansion, and development of bioengineered tissue constructs. While bioprinted stem cells present an opportunity to replicate physiological microenvironments with precision, the future of this practice relies heavily on the optimization of the cellular microenvironment. To fabricate tissue constructs that are useful in replicating physiological conditions in laboratory settings, or in preparation for transplantation to a living host, the microenvironment must mimic conditions that allow bioprinted stem cells to proliferate, differentiate, and migrate. The advances of bioprinting stem cells and directing cell fate have the potential to provide feasible and translatable approach to creating complex tissues and organs. This review will examine the methods through which bioprinted stem cells are differentiated into desired cell lineages through biochemical, biological, and biomechanical techniques.
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http://dx.doi.org/10.1021/acs.chemrev.0c00126DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7676498PMC
October 2020

The effect of BMP-mimetic peptide tethering bioinks on the differentiation of dental pulp stem cells (DPSCs) in 3D bioprinted dental constructs.

Biofabrication 2020 07 1;12(3):035029. Epub 2020 Jul 1.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Winston-Salem, NC 27157, United States of America.

The goal of this study was to use 3D bioprinting technology to create a bioengineered dental construct containing human dental pulp stem cells (hDPSCs). To accomplish this, we first developed a novel bone morphogenetic protein (BMP) peptide-tethering bioink formulation and examined its rheological properties, its printability, and the structural stability of the bioprinted construct. Second, we evaluated the survival and differentiation of hDPSCs in the bioprinted dental construct by measuring cell viability, proliferation, and gene expression, as well as histological and immunofluorescent analyses. Our results showed that the peptide conjugation into the gelatin methacrylate-based bioink formulation was successfully performed. We determined that greater than 50% of the peptides remained in the bioprinted construct after three weeks in vitro cell culture. Human DPSC viability was >90% in the bioprinted constructs immediately after the printing process. Alizarin Red staining showed that the BMP peptide construct group exhibited the highest calcification as compared to the growth medium, osteogenic medium, and non-BMP peptide construct groups. In addition, immunofluorescent and quantitative reverse transcription-polymerase chain reaction analyses showed robust expression of dentin sialophosphoprotein and osteocalcin in the BMP peptide dental constructs. Together, these results strongly suggested that BMP peptide-tethering bioink could accelerate the differentiation of hDPSCs in 3D bioprinted dental constructs.
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http://dx.doi.org/10.1088/1758-5090/ab9492DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7641314PMC
July 2020

Neural cell integration into 3D bioprinted skeletal muscle constructs accelerates restoration of muscle function.

Nat Commun 2020 02 24;11(1):1025. Epub 2020 Feb 24.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Winston-Salem, NC, 27157, USA.

A bioengineered skeletal muscle construct that mimics structural and functional characteristics of native skeletal muscle is a promising therapeutic option to treat extensive muscle defect injuries. We previously showed that bioprinted human skeletal muscle constructs were able to form multi-layered bundles with aligned myofibers. In this study, we investigate the effects of neural cell integration into the bioprinted skeletal muscle construct to accelerate functional muscle regeneration in vivo. Neural input into this bioprinted skeletal muscle construct shows the improvement of myofiber formation, long-term survival, and neuromuscular junction formation in vitro. More importantly, the bioprinted constructs with neural cell integration facilitate rapid innervation and mature into organized muscle tissue that restores normal muscle weight and function in a rodent model of muscle defect injury. These results suggest that the 3D bioprinted human neural-skeletal muscle constructs can be rapidly integrated with the host neural network, resulting in accelerated muscle function restoration.
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http://dx.doi.org/10.1038/s41467-020-14930-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7039897PMC
February 2020

Reno-protection of Urine-derived Stem Cells in A Chronic Kidney Disease Rat Model Induced by Renal Ischemia and Nephrotoxicity.

Int J Biol Sci 2020 1;16(3):435-446. Epub 2020 Jan 1.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC, USA.

Drug-induced nephrotoxicity can occur in patients with pre-existing renal dysfunction or renal ischemia, potentially leading to chronic kidney disease (CKD) and end-stage renal disease (ESRD). Prompt treatment of CKD and the related side effects is critical in preventing progression to ESRD. The goal of this study was to demonstrate the therapeutic potential of urine-derived stem cells (USC) to treat chronic kidney disease-induced by nephrotoxic drugs and renal ischemia. Human USC were collected, expanded and characterized by flow cytometry. A CKD model was induced by creating an ischemia-reperfusion injury and gentamicin administration. Twenty-eight adult immunodeficient rats were divided into three groups: PBS-treated group (n=9), USC-treated group (n=9), and sham group with age-matched control animals (n=10). Cell suspension of USC (5 x 10 / 100µl / kidney) or PBS was injected bilaterally into the renal parenchyma 9 weeks after CKD model creation. Renal function was evaluated by collection blood and urine samples to measure serum creatinine and glomerulus filtration rate. The kidneys were harvested 12 weeks after cell injection. Histologically, the extent of glomerulosclerosis and tubular atrophy, the amount of collagen deposition, interstitial fibrosis, inflammatory monocyte infiltration, and expression of transforming growth factor beta 1 (TGF-ß1), and superoxide dismutase 1 (SOD-1) were examined. USC expressed renal parietal epithelial cells (CD24, CD29 and CD44). Renal function, measured by GFR and serum Cr in USC-treated group were significantly improved compared to PBS-treated animals (p<0.05). The degree of glomerular sclerosis and atrophic renal tubules, the amount of fibrosis, and monocyte infiltration significantly decreased in USC-treated group compared to the PBS group (p<0.05). The level of TGF-ß1 expression in renal tissues was also significantly lower in the PBS group, while the level of SOD-1 expression was significantly elevated in the USC group, compared to PBS group (p<0.05). The present study demonstrates the nephron-protective effect of USC on renal function via anti-inflammatory, anti-oxidative stress, and anti-fibrotic activity in a dual-injury CKD rat model. This provides an alternative treatment for CKD in certain clinical situations, such as instances where CKD is due to drug-induced nephrotoxicity and renal ischemia.
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http://dx.doi.org/10.7150/ijbs.37550DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6990904PMC
January 2021

Dynamic Changes in Erectile Function and Histological Architecture After Intracorporal Injection of Human Placental Stem Cells in a Pelvic Neurovascular Injury Rat Model.

J Sex Med 2020 03 27;17(3):400-411. Epub 2020 Jan 27.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Winston-Salem, NC, USA. Electronic address:

Introduction: The human placenta provides a bountiful and noncontroversial source of stem cells which have the potential for regeneration of injured tissue. These cells may restore erectile function after neurovascular tissue injury such as that seen in radical pelvic surgeries and pelvic trauma.

Aim: To determine the effect of human placenta-derived stem cells on erectile function recovery and histological changes at various time points in a cavernous nerve injury rat model and to study the fate of injected stem cells throughout the regenerative process.

Methods: Human placental stem cells (PSCs) were dual labeled with monomeric Katushka far red fluorescent protein (mKATE)-renLUC using a lentivirus vector. A pelvic neurovascular injury-induced erectile dysfunction model was established in male, athymic rats by crushing the cavernous nerves and ligating the internal pudendal neurovascular bundles, bilaterally. At the time of defect creation, nonlabeled PSCs were injected into the corpus cavernosum at a concentration of 2.5 × 10 cells/0.2 mL. The phosphate-buffered saline-treated group served as the negative control group, and age-matched rats (age-matched controls) were used as the control group. Erectile function, histomorphological analyses, and Western blot were assessed at 1, 6, and 12 weeks after model creation. The distribution of implanted, dual-labeled PSCs was monitored using an in vivo imaging system (IVIS). Implanted cells were further tracked by detection of mKATE fluorescence in histological sections.

Main Outcome Measure: The main outcome measure includes intracavernous pressure/mean arterial pressure ratio, neural, endothelial, smooth muscle cell regeneration, mKATE fluorescence, and IVIS imaging.

Results: The ratio of intracavernous pressure to mean arterial pressure significantly increased in PSC-injected rats compared with phosphate-buffered saline controls (P < 0.05) at the 6- and 12-week time points, reaching 72% and 68% of the age-matched control group, respectively. Immunofluorescence staining and Western blot analysis showed significant increases in markers of neurons (84.3%), endothelial cells (70.2%), and smooth muscle cells (70.3%) by 6 weeks in treatment groups compared with negative controls. These results were maintained through 12 weeks. IVIS analysis showed luminescence of implanted PSCs in the injected corpora immediately after injection and migration of cells to the sites of injury, including the incision site and periprostatic vasculature by day 1. mKATE fluorescence data revealed the presence of PSCs in the penile corpora and major pelvic ganglion at 1 and 3 days postoperatively. At 7 days, immunofluorescence of penile PSCs had disappeared and was diminished in the major pelvic ganglion.

Clinical Implications: Placenta-derived stem cells may represent a future "off-the-shelf" treatment to mitigate against development of erectile dysfunction after radical prostatectomy or other forms of pelvic injury.

Strength & Limitations: Single dose injection of PSCs after injury resulted in maximal functional recovery and tissue regeneration at 6 weeks, and the results were maintained through 12 weeks. Strategies to optimize adult stem cell therapy might achieve more effective outcomes for human clinical trials.

Conclusion: Human PSC therapy effectively restores the erectile tissue and function in this animal model. Thus, PSC therapy may provide an attractive modality to lessen the incidence of erectile dysfunction after pelvic neurovascular injury. Further improvement in tissue regeneration and functional recovery may be possible using multiple injections or systemic introduction of stem cells. Gu X, Thakker PU, Matz EL, et al. Dynamic Changes in Erectile Function and Histological Architecture After Intracorporal Injection of Human Placental Stem Cells in a Pelvic Neurovascular Injury Rat Model. J Sex Med 2020;17:400-411.
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http://dx.doi.org/10.1016/j.jsxm.2019.12.002DOI Listing
March 2020

Clinical Council Roundtable Discussion: Opportunities and Challenges in Clinical Translation.

Tissue Eng Part A 2020 02 23;26(3-4):113-119. Epub 2020 Jan 23.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina.

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http://dx.doi.org/10.1089/ten.tea.2019.0037DOI Listing
February 2020

Assessment methodologies for extrusion-based bioink printability.

Biofabrication 2020 02 19;12(2):022003. Epub 2020 Feb 19.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Winston-Salem, NC, United States of America. School of Biomedical Engineering and Sciences, Wake Forest University-Virginia Tech, Winston-Salem, NC, United States of America.

Extrusion-based bioprinting is one of the leading manufacturing techniques for tissue engineering and regenerative medicine. Its primary limitation is the lack of materials, known as bioinks, which are suitable for the bioprinting process. The degree to which a bioink is suitable for bioprinting has been described as its 'printability.' However, a lack of clarity surrounding the methodologies used to evaluate a bioink's printability, as well as the usage of the term itself, have hindered the field. This article presents a review of measures used to assess the printability of extrusion-based bioinks in an attempt to assist researchers during the bioink development process. Many different aspects of printability exist and many different measurements have been proposed as a consequence. Researchers often do not evaluate a new bioink's printability at all, while others simply do so qualitatively. Several quantitative measures have been presented for the extrudability, shape fidelity, and printing accuracy of bioinks. Different measures have been developed even within these aspects, each testing the bioink in a slightly different way. Additionally, other relevant measures which had little or no examples of quantifiable methods are also to be considered. Looking forward, further work is needed to improve upon current assessment methodologies, to move towards a more comprehensive view of printability, and to standardize these printability measurements between researchers. Better assessment techniques will naturally lead to a better understanding of the underlying mechanisms which affect printability and better comparisons between bioinks. This in turn will help improve upon the bioink development process and the bioinks available for use in bioprinting.
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http://dx.doi.org/10.1088/1758-5090/ab6f0dDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7039534PMC
February 2020

Decellularization and recellularization strategies for translational medicine.

Methods 2020 01 28;171:1-2. Epub 2019 Dec 28.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27101, USA.

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http://dx.doi.org/10.1016/j.ymeth.2019.12.005DOI Listing
January 2020

Bioprinted Skin Recapitulates Normal Collagen Remodeling in Full-Thickness Wounds.

Tissue Eng Part A 2020 05 28;26(9-10):512-526. Epub 2020 Jan 28.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA.

Over 1 million burn injuries are treated annually in the United States, and current tissue engineered skin fails to meet the need for full-thickness replacement. Bioprinting technology has allowed fabrication of full-thickness skin and has demonstrated the ability to close full-thickness wounds. However, analysis of collagen remodeling in wounds treated with bioprinted skin has not been reported. The purpose of this study is to demonstrate the utility of bioprinted skin for epidermal barrier formation and normal collagen remodeling in full-thickness wounds. Human keratinocytes, melanocytes, fibroblasts, dermal microvascular endothelial cells, follicle dermal papilla cells, and adipocytes were suspended in fibrinogen bioink and bioprinted to form a tri-layer skin structure. Bioprinted skin was implanted onto 2.5 × 2.5 cm full-thickness excisional wounds on athymic mice, compared with wounds treated with hydrogel only or untreated wounds. Total wound closure, epithelialization, and contraction were quantified, and skin samples were harvested at 21 days for histology. Picrosirius red staining was used to quantify collagen fiber orientation, length, and width. Immunohistochemical (IHC) staining was performed to confirm epidermal barrier formation, dermal maturation, vascularity, and human cell integration. All bioprinted skin treated wounds closed by day 21, compared with open control wounds. Wound closure in bioprinted skin treated wounds was primarily due to epithelialization. In contrast, control hydrogel and untreated groups had sparse wound coverage and incomplete closure driven primarily by contraction. Picrosirius red staining confirmed a normal basket weave collagen organization in bioprinted skin-treated wounds compared with parallel collagen fibers in hydrogel only and untreated wounds. IHC staining at day 21 demonstrated the presence of human cells in the regenerated dermis, the formation of a stratified epidermis, dermal maturation, and blood vessel formation in bioprinted skin, none of which was present in control hydrogel treated wounds. Bioprinted skin accelerated full-thickness wound closure by promoting epidermal barrier formation, without increasing contraction. This healing process is associated with human cells from the bioprinted skin laying down a healthy, basket-weave collagen network. The remodeled skin is phenotypically similar to human skin and composed of a composite of graft and infiltrating host cells. Impact statement We have demonstrated the ability of bioprinted skin to enhance closure of full-thickness wounds through epithelialization and normal collagen remodeling. To our knowledge, this article is the first to quantify collagen remodeling by bioprinted skin in full-thickness wounds. Our methods and results can be used to guide further investigation of collagen remodeling by tissue engineered skin products to improve ongoing and future bioprinting skin studies. Ultimately, our skin bioprinting technology could translate into a new treatment for full-thickness wounds in human patients with the ability to recapitulate normal collagen remodeling in full-thickness wounds.
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http://dx.doi.org/10.1089/ten.TEA.2019.0319DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7249461PMC
May 2020

Efficient myotube formation in 3D bioprinted tissue construct by biochemical and topographical cues.

Biomaterials 2020 02 19;230:119632. Epub 2019 Nov 19.

Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC, 27157, USA; Department of Biomechatronic Engineering, College of Biotechnology and Bioengineering, Sungkyunkwan University (SKKU), Suwon, 16419, South Korea. Electronic address:

Biochemical and biophysical cues directly affect cell morphology, adhesion, proliferation, and phenotype, as well as differentiation; thus, they have been commonly utilized for designing and developing biomaterial systems for tissue engineering applications. To bioengineer skeletal muscle tissues, the efficient and stable formation of aligned fibrous multinucleated myotubes is essential. To achieve this goal, we employed a decellularized extracellular matrix (dECM) as a biochemical component and a modified three-dimensional (3D) cell-printing process to produce an in situ uniaxially aligned/micro-topographical structure. The dECM was derived from the decellularization of porcine skeletal muscles and chemically modified by methacrylate process to enhance mechanical stability. By using this ECM-based material and the 3D printing capability, we were able to produce a cell-laden dECM-based structure with unique topographical cues. The myoblasts (C2C12 cell line) laden in the printed structure were aligned and differentiated with a high degree of myotube formation, owing to the synergistic effect of the skeletal muscle-specific biochemical and topographical cues. In particular, the increase of the gene-expression levels of the dECM structure with topographical cues was approximately 1.5-1.8-fold compared with those of a gelatin methacrylate (GelMA)-based structure with the same topographical cues and a dECM-based structure without topographical cues. According to these in vitro cellular responses, the 3D printed dECM-based structures with topographical cues have the potential for bioengineering functional skeletal muscle tissues, and this strategy can be extended for many musculoskeletal tissues, such as tendons and ligaments and utilized for developing in vitro tissue-on-a-chip models in drug screening and development.
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http://dx.doi.org/10.1016/j.biomaterials.2019.119632DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7141931PMC
February 2020

Multimaterial Dual Gradient Three-Dimensional Printing for Osteogenic Differentiation and Spatial Segregation.

Tissue Eng Part A 2020 03 27;26(5-6):239-252. Epub 2019 Dec 27.

Department of Bioengineering, Rice University, Houston, Texas.

In this study of three-dimensional (3D) printed composite β-tricalcium phosphate (β-TCP)-/hydroxyapatite/poly(ɛ-caprolactone)-based constructs, the effects of vertical compositional ceramic gradients and architectural porosity gradients on the osteogenic differentiation of rabbit bone marrow-derived mesenchymal stem cells (MSCs) were investigated. Specifically, three different concentrations of β-TCP (0, 10, and 20 wt%) and three different porosities (33% ± 4%, 50% ± 4%, and 65% ± 3%) were examined to elucidate the contributions of chemical and physical gradients on the biochemical behavior of MSCs and the mineralized matrix production within a 3D culture system. By delaminating the constructs at the gradient transition point, the spatial separation of cellular phenotypes could be specifically evaluated for each construct section. Results indicated that increased concentrations of β-TCP resulted in upregulation of osteogenic markers, including alkaline phosphatase activity and mineralized matrix development. Furthermore, MSCs located within regions of higher porosity displayed a more mature osteogenic phenotype compared to MSCs in lower porosity regions. These results demonstrate that 3D printing can be leveraged to create multiphasic gradient constructs to precisely direct the development and function of MSCs, leading to a phenotypic gradient. Impact Statement In this study, three-dimensional (3D) printed ceramic/polymeric constructs containing discrete vertical gradients of both composition and porosity were fabricated to precisely control the osteogenic differentiation of mesenchymal stem cells. By making simple alterations in construct architecture and composition, constructs containing heterogenous populations of cells were generated, where gradients in scaffold design led to corresponding gradients in cellular phenotype. The study demonstrates that 3D printed multiphasic composite constructs can be leveraged to create complex heterogeneous tissues and interfaces.
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http://dx.doi.org/10.1089/ten.TEA.2019.0204DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7133451PMC
March 2020

Kidney regeneration approaches for translation.

World J Urol 2020 Sep 6;38(9):2075-2079. Epub 2019 Nov 6.

Wake Forest School of Medicine, Wake Forest Institute for Regenerative Medicine, Medical Center Boulevard, Winston-Salem, NC, 27157, USA.

The increase in the incidence of chronic kidney diseases that progress to end-stage renal disease has become a significant health problem worldwide. While dialysis can maintain and prolong survival, the only definitive treatment that can restore renal function is transplantation. Unfortunately, many of these patients die waiting for transplantable kidneys due to the severe shortage of donor organs. Tissue engineering and regenerative medicine approaches have been applied in recent years to develop viable therapies that could provide solutions to these patients. Cell-based and cell-free approaches have been proposed to address the challenges associated with chronic kidney diseases. Strategies and progress toward developing alternative therapeutic options will be reviewed.
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http://dx.doi.org/10.1007/s00345-019-02999-xDOI Listing
September 2020

Structure establishment of three-dimensional (3D) cell culture printing model for bladder cancer.

PLoS One 2019 22;14(10):e0223689. Epub 2019 Oct 22.

Department of Urology, College of Medicine, Chung-Ang University, Seoul, Republic of Korea.

Purpose: Two-dimensional (2D) cell culture is a valuable method for cell-based research but can provide unpredictable, misleading data about in vivo responses. In this study, we created a three-dimensional (3D) cell culture environment to mimic tumor characteristics and cell-cell interactions to better characterize the tumor formation response to chemotherapy.

Materials And Methods: We fabricated the 3D cell culture samples using a 3D cell bio printer and the bladder cancer cell line 5637. T24 cells were used for 2D cell culture. Then, rapamycin and Bacillus Calmette-Guérin (BCG) were used to examine their cancer inhibition effects using the two bladder cancer cell lines. Cell-cell interaction was measured by measuring e-cadherin and n-cadherin secreted via the epithelial-mesenchymal transition (EMT).

Results: We constructed a 3D cell scaffold using gelatin methacryloyl (GelMA) and compared cell survival in 3D and 2D cell cultures. 3D cell cultures showed higher cancer cell proliferation rates than 2D cell cultures, and the 3D cell culture environment showed higher cell-to-cell interactions through the secretion of E-cadherin and N-cadherin. Assessment of the effects of drugs for bladder cancer such as rapamycin and BCG showed that the effect in the 2D cell culture environment was more exaggerated than that in the 3D cell culture environment.

Conclusions: We fabricated 3D scaffolds with bladder cancer cells using a 3D bio printer, and the 3D scaffolds were similar to bladder cancer tissue. This technique can be used to create a cancer cell-like environment for a drug screening platform.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0223689PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6804961PMC
March 2020