Scaffold-Free Multicellular 3D Tissue Constructs Utilizing Bio-orthogonal Click Strategy.

Multicellular 3D tissue constructs (MTCs) are important in biomedical research due to their capacity to accurately mimic the structure and variation found in real tissues. This study presents a novel bio-orthogonal engineering strategy (BIEN), a transformative scaffold-free approach, to create advanced MTCs. BIEN harnesses the cellular biosynthetic machinery to incorporate bio-orthogonal azide reporters into cell surface glycoconjugates, followed by a click reaction with multiarm PEG, resulting in rapid assembly of MTCs. The implementation of this cutting-edge strategy culminates in the formation of uniform, heterogeneous spheroids, characterized by a high degree of intercellular junction and pluripotency. Remarkably, MTCs simulate tumor features, ensure cell heterogeneity, and significantly improve the subcutaneous xenograft model after transplantation, thereby bolstering both in vitro and in vivo research models. In conclusion, the utilization of the bio-orthogonal engineering strategy as a scaffold-free method to generate superior MTCs holds promising potential for driving advancements in cancer research.

Quantitative Photothermal Characterization with Bioprinted 3D Complex Tissue Constructs for Early-Stage Breast Cancer Therapy Using Gold Nanorods.

Plasmonic photothermal therapy (PPTT) using gold nanoparticles (AuNPs) has shown great potential for use in selective tumor treatment, because the AuNPs can generate destructive heat preferentially upon irradiation. However, PPTT using AuNPs has not been added to practice, owing to insufficient heating methods and tissue temperature measurement techniques, leading to unreliable and inaccurate treatments. Because the photothermal properties of AuNPs vary with laser power, particle optical density, and tissue depth, the accurate prediction of heat generation is indispensable for clinical treatment. In this report, bioprinted 3D complex tissue constructs comprising processed gel obtained from porcine skin and human decellularized adipose tissue are presented for characterization of the photothermal properties of gold nanorods (AuNRs) having an aspect ratio of 3.7 irradiated by a near-infrared laser. Moreover, an analytical function is suggested for achieving PPTT that can cause thermal damage selectively on early-stage human breast cancer by regulating the heat generation of the AuNRs in the tissue.

Investigating Tissue Mechanics in vitro Using Untethered Soft Robotic Microdevices.

This paper presents the design, fabrication, and operation of a soft robotic compression device that is remotely powered by laser illumination. We combined the rapid and wireless response of hybrid nanomaterials with state-of-the-art microengineering techniques to develop machinery that can apply physiologically relevant mechanical loading. The passive hydrogel structures that constitute the compliant skeleton of the machines were fabricated using single-step in situ polymerization process and directly incorporated around the actuators without further assembly steps. Experimentally validated computational models guided the design of the compression mechanism. We incorporated a cantilever beam to the prototype for life-time monitoring of mechanical properties of cell clusters on optical microscopes. The mechanical and biochemical compatibility of the chosen materials with living cells together with the on-site manufacturing process enable seamless interfacing of soft robotic devices with biological specimen.

Minimally Invasive Delivery of 3D Shape Recoverable Constructs with Ordered Structures for Tissue Repair.

Minimally invasive procedures are becoming increasingly more common in surgery. However, the biomaterials capable of delivering biomimetic, three-dimensional (3D) functional tissues in a minimally invasive manner and exhibiting ordered structures after delivery are lacking. Herein, we reported the fabrication of gelatin methacryloyl (GelMA)-coated, 3D expanded nanofiber scaffolds, and their potential applications in minimally invasive delivery of 3D functional tissue constructs with ordered structures and clinically appropriate sizes (4 cm × 2 cm × 1.5 mm). GelMA-coated, expanded 3D nanofiber scaffolds produced by combining electrospinning, gas-foaming expansion, hydrogel coating, and cross-linking are extremely shape recoverable after release of compressive strain, displaying a superelastic property. Such scaffolds can be seeded with various types of cells, including dermal fibroblasts, bone marrow-derived mesenchymal stem cells, and human neural stem/precursor cells to form 3D complex tissue constructs. Importantly, the developed 3D tissue constructs can be compressed and loaded into a 4 mm diameter glass tube for minimally invasive delivery without compromising the cell viability. Taken together, the method developed in this study could hold great promise for transplantation of biomimetic, 3D functional tissue constructs with well-organized structures for tissue repair and regeneration using minimally invasive procedures like laparoscopy and thoracoscopy.

Organoids and Microphysiological Systems: New Tools for Ophthalmic Drug Discovery.

Organoids are adept at preserving the inherent complexity of a given cellular environment and when integrated with engineered micro-physiological systems (MPS) present distinct advantages for simulating a precisely controlled geometrical, physical, and biochemical micro-environment. This then allows for real-time monitoring of cell-cell interactions. As a result, the two aforementioned technologies hold significant promise and potential in studying ocular physiology and diseases by replicating specific eye tissue microstructures in vitro. This miniaturized review begins with defining the science behind organoids/MPS and subsequently introducing methods for generating organoids and engineering MPS. Furthermore, we will discuss the current state of organoids and MPS models in retina, cornea surrogates, and other ocular tissue, in regards to physiological/disease conditions. Finally, future prospective on organoid/MPS will be covered here. Organoids and MPS technologies closely recapture the in vivo microenvironment and thusly will continue to provide new understandings in organ functions and novel approaches to drug development.

Effect of Pulsed Electromagnetic Fields on Human Mesenchymal Stem Cells Using 3D Magnetic Scaffolds.

Alternative bone regeneration strategies that do not rely on harvested tissue or exogenous growth factors are needed. One of the major challenges in tissue reconstruction is recreating the bone tissue microenvironment using the appropriate combination of cells, scaffold, and stimulation to direct differentiation. This study presents a bone regeneration formulation that involves the use of human adipose-derived mesenchymal stem cells (hASCs) and a three-dimensional (3D) hydrogel scaffold based on self-assembled RADA16 peptides containing superparamagnetic iron oxide nanoparticles (NPs). Although superparamagnetic NPs could be used as stimulus to manipulate the cell proliferation and differentiation, in this paper their use is explored for assisting osteogenic differentiation of hASCs in conjunction with direct stimulation by extremely low-frequency pulsed electromagnetic fields (pEMFs). Cellular morphology, proliferation, and viability, as well as alkaline phosphatase activity, calcium deposition, and osteogenic capacity were monitored for cells cultured up to 21 days in the 3D construct. The results show that the pEMFs and NPs do not have any negative effect on cell viability, but instead distinctly induced early differentiation of hASCs to an osteoblastic phenotype, when compared with cells without biophysical stimulation. This effect is attributed to synergy between the pEMFs and NPs, which may have stimulated mechanotransduction pathways, which, in turn activated biochemical signals between cells to differentiate or proliferate. This approach may offer a safe and effective option for the treatment of non-union bone fractures. Bioelectromagnetics. © 2020 The Authors. Bioelectromagnetics published by Wiley Periodicals, Inc.

Full-Thickness Human Skin Equivalent Models of Atopic Dermatitis.

Atopic dermatitis is a chronic inflammatory skin disease caused by complex multifactorial etiology. In the recent years, there have been significant advances in tissue engineering and the generation of in vitro skin models representative of healthy and diseased states. This chapter describes the methodology for the fabrication of in vitro human skin equivalent (HSE) from human keratinocytes and fibroblasts using a fibrin-based dermal matrix and serum-free culture conditions. Modification of the culture conditions with the supplementation of Th2 cytokines such as interleukin-4 induces the development of atopic dermatitis-like skin model. The chapter also describes the histological and immunohistochemical tools for characterization of the HSE model. The reconstruction of tissue-engineered HSE models that recapitulate the essential features of atopic dermatitis provides powerful tools for deeper understanding of the underlying pathological mechanisms on epidermal level, identification and testing of novel treatment options, and safety and toxicological evaluation in a pathophysiologically relevant system.

A Single-Step Self-Assembly Approach for the Fabrication of Aligned and Multilayered Three-Dimensional Tissue Constructs Using Multidomain Peptide Hydrogel.

Hydrogels are homogenous materials that are limited in their ability to form oriented multilayered architecture in three-dimensional (3D) tissue constructs. Current techniques have led to advancements in this area. Such techniques often require extra devices and/or involve complex processes that are inaccessible to many laboratories. Here is described a one-step methodology that permits reliable alignment of cells into multiple layers using a self-assembling multidomain peptide (MDP) hydrogels. We characterized the structural features, viability, and molecular properties of dental pulp cells fabricated with MDP and demonstrated that manipulation of the layering of cells in the scaffolds was achieved by decreasing the weight by volume percentage (w/v%) of MDP contained within the scaffold. This approach allows cells to remodel their environment and enhanced various gene expression profiles, such as cell proliferation, angiogenesis, and extracellular matrix (ECM) remodeling-related genes. We further validated our approach for constructing various architectural configurations of tissues by fabricating cells into stratified multilayered and tubular structures. Our methodology provides a simple, rapid way to generate 3D tissue constructs with multilayered architectures. This method shows great potential to mimic in vivo microenvironments for cells and may be of benefit in modeling more complex tissues in the field of regenerative medicine.

Three-dimensional direct cell bioprinting for tissue engineering.

Bioprinting is a relatively new technology where living cells with or without biomaterials are printed layer-by-layer in order to create three-dimensional (3D) living structures. In this article, novel bioprinting methodologies are developed to fabricate 3D biological structures directly from computer models using live multicellular aggregates. Multicellular aggregates made out of at least two cell types from fibroblast, endothelial and smooth muscle cells are prepared and optimized. A novel bioprinting approach is proposed in order to continuously extrude cylindrical multicellular aggregates through the bioprinter's glass microcapillaries. The multicellular aggregates are first aspirated into a capillary and then compressed to form a continuous cylindrical multicellular bioink. To overcome surface tension-driven droplet formation, the required compression ratio is calculated. Based on the developed bioprinting strategies, multicellular aggregates and their support structures are bioprinted to form 3D tissue constructs with predefined shapes. The effect of the bioprinting process was examined for fusion, cell viability at different compression ratios, and f-actin cytoskeletal organization. The results show that the bioprinted 3D constructs fuse rapidly and have high cell viability after printing. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 2530-2544, 2017.