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AUTHOR Wei, Zhengxi and Liu, Xue and Ooka, Masato and Zhang, Li and Song, Min Jae and Huang, Ruili and Kleinstreuer, Nicole C. and Simeonov, Anton and Xia, Menghang and Ferrer, Marc
Title Two-Dimensional Cellular and Three-Dimensional Bio-Printed Skin Models to Screen Topical-Use Compounds for Irritation Potential [Abstract]
Year 2020
Journal/Proceedings Frontiers in Bioengineering and Biotechnology
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Assessing skin irritation potential is critical for the safety evaluation of topical drugs and other consumer products such as cosmetics. The use of advanced cellular models, as an alternative to replace animal testing in the safety evaluation for both consumer products and ingredients, is already mandated by law in the European Union (EU) and other countries. However, there has not yet been a large-scale comparison of the effects of topical-use compounds in different cellular skin models. This study assesses the irritation potential of topical-use compounds in different cellular models of the skin that are compatible with high throughput screening (HTS) platforms. A set of 451 topical-use compounds were first tested for cytotoxic effects using two-dimensional (2D) monolayer models of primary neonatal keratinocytes and immortalized human keratinocytes. Forty-six toxic compounds identified from the initial screen with the monolayer culture systems were further tested for skin irritation potential on reconstructed human epidermis (RhE) and full thickness skin (FTS) three-dimensional (3D) tissue model constructs. Skin irritation potential of the compounds was assessed by measuring tissue viability, trans-epithelial electrical resistance (TEER), and secretion of cytokines interleukin 1 alpha (IL-1α) and interleukin 18 (IL-18). Among known irritants, high concentrations of methyl violet and methylrosaniline decreased viability, lowered TEER, and increased IL-1α secretion in both RhE and FTS models, consistent with irritant properties. However, at low concentrations, these two compounds increased IL-18 secretion without affecting levels of secreted IL-1α, and did not reduce tissue viability and TEER, in either RhE or FTS models. This result suggests that at low concentrations, methyl violet and methylrosaniline have an allergic potential without causing irritation. Using both HTS-compatible 2D cellular and 3D tissue skin models, together with irritation relevant activity endpoints, we obtained data to help assess the irritation effects of topical-use compounds and identify potential dermal hazards.
AUTHOR Derr, Kristy and Zou, Jinyun and Luo, Keren and Song, Min Jae and Sittampalam, G. Sitta and Zhou, Chao and Michael, Samuel and Ferrer, Marc and Derr, Paige
Title Fully 3D Bioprinted Skin Equivalent Constructs with Validated Morphology and Barrier Function [Abstract]
Year 2019
Journal/Proceedings Tissue Engineering Part C: Methods
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Development of high throughput, reproducible, three-dimensional bioprinted skin equivalents that are morphologically and functionally comparable to native skin tissue is advancing research in skin diseases, and providing a physiologically relevant platform for the development of therapeutics, transplants for regenerative medicine, and testing of skin products like cosmetics. Current protocols for the production of engineered skin rafts are limited in their ability to control three dimensional geometry of the structure and contraction leading to variability of skin function between constructs. Here we describe a method for the biofabrication of skin equivalents that are fully bioprinted using an open market bioprinter, made with commercially available primary cells and natural hydrogels. The unique hydrogel formulation allows for the production of a human-like skin equivalent with minimal lateral tissue contraction in a multiwell plate format, thus making them suitable for high throughput bioprinting in a single print with fast print and relatively short incubation times. The morphology and barrier function of the fully three-dimensional bioprinted skin equivalents are validated by immunohistochemistry staining, optical coherence tomography, and permeation assays.
AUTHOR Liu, Xue and Michael, Samuel and Bharti, Kapil and Ferrer, Marc and Song, Min Jae
Title A biofabricated vascularized skin model of atopic dermatitis for preclinical studies [Abstract]
Year 2020
Journal/Proceedings Biofabrication
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Three-dimensional (3D) biofabrication techniques enable the production of multicellular tissue models as assay platforms for drug screening. The increased cellular and physiological complexity in these 3D tissue models should recapitulate the relevant biological environment found in the body. Here we describe the use of 3D bioprinting techniques to fabricate skin equivalent tissues of varying physiological complexity, including human epidermis, non-vascularized and vascularized full-thickness skin tissue equivalents, in a multi-well platform to enable drug screening. Human keratinocytes, fibroblasts, and pericytes, and induced pluripotent stem cell (iPSC)-derived endothelial cells were used in the biofabrication process to produce the varying complexity. The skin equivalents exhibit the correct structural markers of dermis and epidermis stratification, with physiological functions of the skin barrier. The robustness, versatility and reproducibility of the biofabrication techniques are further highlighted by the generation of atopic dermatitis (AD)-disease like tissues. These AD models demonstrate several clinical hallmarks of the disease, including: (i) spongiosis and hyperplasia; (ii) early and terminal expression of differentiation proteins; and (iii) increases in levels of pro-inflammatory cytokines. We show the pre-clinical relevance of the biofabricated AD tissue models to correct disease phenotype by testing the effects of dexamethasone, an anti-inflammatory corticosteroid, and three Janus Kinase inhibitors from clinical trials for AD. This study demonstrates the development of a versatile and reproducible bioprinting approach to create human skin equivalents with a range of cellular complexity for disease modelling. In addition, we establish several assay readouts that are quantifiable, robust, AD relevant, and can be scaled up for compound screening. The results show that the cellular complexity of the tissues develops a more physiologically relevant AD disease model. Thus, the skin models in this study offer an in vitro approach for the rapid understanding of pathological mechanisms, and testing for efficacy of action and toxic effects of drugs.
AUTHOR Mestre, Rafael and Patiño, Tania and Barceló, Xavier and Anand, Shivesh and Pérez-Jiménez, Ariadna and Sánchez, Samuel
Title Force Modulation and Adaptability of 3D-Bioprinted Biological Actuators Based on Skeletal Muscle Tissue [Abstract]
Year 2019
Journal/Proceedings Advanced Materials Technologies
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Abstract The integration of biological systems into robotic devices might provide them with capabilities acquired from natural systems and significantly boost their performance. These abilities include real-time bio-sensing, self-organization, adaptability, or self-healing. As many muscle-based bio-hybrid robots and bio-actuators arise in the literature, the question of whether these features can live up to their expectations becomes increasingly substantial. Herein, the force generation and adaptability of skeletal-muscle-based bio-actuators undergoing long-term training protocols are analyzed. The 3D-bioprinting technique is used to fabricate bio-actuators that are functional, responsive, and have highly aligned myotubes. The bio-actuators are 3D-bioprinted together with two artificial posts, allowing to use it as a force measuring platform. In addition, the force output evolution and dynamic gene expression of the bio-actuators are studied to evaluate their degree of adaptability according to training protocols of different frequencies and mechanical stiffness, finding that their force generation could be modulated to different requirements. These results shed some light into the fundamental mechanisms behind the adaptability of muscle-based bio-actuators and highlight the potential of using 3D bioprinting as a rapid and cost-effective tool for the fabrication of custom-designed soft bio-robots.
AUTHOR Visscher, Dafydd O. and Bos, Ernst J. and Peeters, Mirte and Kuzmin, Nikolay V. and Groot, Marie Louise and Helder, Marco N. and van Zuijlen, Paul P. M.
Title Cartilage Tissue Engineering: Preventing Tissue Scaffold Contraction Using a 3D-Printed Polymeric Cage. [Abstract]
Year 2016
Journal/Proceedings Tissue engineering Part C: Methods
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Scaffold contraction is a common but underestimated problem in the field of tissue engineering. It becomes particularly problematic when creating anatomically complex shapes such as the ear. The aim of this study was to develop a contraction-free biocompatible scaffold construct for ear cartilage tissue engineering. To address this aim, we used three constructs: (i) a fibrin/hyaluronic acid (FB/HA) hydrogel, (ii) a FB/HA hydrogel combined with a collagen I/III scaffold, and (iii) a cage construct containing (ii) surrounded by a 3D-printed poly-varepsilon-caprolactone mold. A wide range of different cell types were tested within these constructs, including chondrocytes, perichondrocytes, adipose-derived mesenchymal stem cells, and their combinations. After in vitro culturing for 1, 14, and 28 days, all constructs were analyzed. Macroscopic observation showed severe contraction of the cell-seeded hydrogel (i). This could be prevented, in part, by combining the hydrogel with the collagen scaffold (ii) and prevented in total using the 3D-printed cage construct (iii). (Immuno)histological analysis, multiphoton laser scanning microscopy, and biomechanical analysis showed extracellular matrix deposition and increased Young's modulus and thereby the feasibility of ear cartilage engineering. These results demonstrated that the 3D-printed cage construct is an adequate model for contraction-free ear cartilage engineering using a range of cell combinations.