Construction of brain-like spheroids containing endothelial tubular networks by an adhesive culture system.

Spheroids which are composed of several types of cells have been widely studied in the pharmaceutical field as their structure and functions are similar to human organs. Three-dimensional brain-like tissues are one of the most important tissues for the development of medicines to treat brain diseases and for in vitro brain models. In this study, spheroids mainly containing neurons, astrocytes, and endothelial cells were fabricated using a novel 3D culture plate, "MicoCell™" to construct a brain mimicking tissue.

3D-cultured small size adipose-derived stem cell spheroids promote bone regeneration in the critical-sized bone defect rat model.

Adipose-derived stem cells (ADSCs), due to their regenerative ability, have beneficial effects on bone and cartilage defects. In addition, spheroid formation of ADSCs obtained using three-dimensional (3D) culture accelerates the regenerative ability of ADSCs. The study investigated the regenerative effect of 3D-cultured small size ADSC spheroids without a scaffold in rats with defects in the critical-sized calvarial bone.

Fabrication of Spheroids with Dome-Shaped Endothelial Tube Networks by an Adhesive Culture System.

3D functional tissues, such as spheroids fabricated by mesenchymal stem cells (MSCs), which can mimic parts of tissues and organs, have recently been extensively studied in the fields of regenerative medicine and drug discovery. In this study, spheroids containing endothelial tubular structures are fabricated by use of a novel 3D culture plate, "MicoCell." As MicoCell has a mild cell adhesive surface and multicavity structures, it can provide multiple attached spheroids at the same time (about ≈10 to ≈10 spheroids).

3D-Printing of Structure-Controlled Antigen Nanoparticles for Vaccine Delivery.

Targeted delivery of antigens to immune cells using micro/nanocarriers may serve as a therapeutic application for vaccination. However, synthetic carriers have potential drawbacks including cytotoxicity, low encapsulation efficiency of antigen, and lack of a morphological design, which limit the translation of the delivery system to clinical use. Here, we report a carrier-free and three-dimensional (3D)-shape-designed antigen nanoparticle by multiphoton lithography-based 3D-printing.

Micro Vacuum Chuck and Tensile Test System for Bio-Mechanical Evaluation of 3D Tissue Constructed of Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes (hiPS-CM).

In this report, we propose a micro vacuum chuck (MVC) which can connect three-dimensional (3D) tissues to a tensile test system by vacuum pressure. Because the MVC fixes the 3D tissue by vacuum pressure generated on multiple vacuum holes, it is expected that the MVC can fix 3D tissue to the system easily and mitigate the damage which can happen by handling during fixing. In order to decide optimum conditions for the size of the vacuum holes and the vacuum pressure, various sized vacuum holes and vacuum pressures were applied to a normal human cardiac fibroblast 3D tissue.

Engraftment and morphological development of vascularized human iPS cell-derived 3D-cardiomyocyte tissue after xenotransplantation.

One of the major challenges in cell-based cardiac regenerative medicine is the in vitro construction of three-dimensional (3D) tissues consisting of induced pluripotent stem cell-derived cardiomyocyte (iPSC-CM) and a blood vascular network supplying nutrients and oxygen throughout the tissue after implantation. We have successfully built a vascularized iPSC-CM 3D-tissue using our validated cell manipulation technique. In order to evaluate an availability of the 3D-tissue as a biomaterial, functional morphology of the tissues was examined by light and transmission electron microscopy through their implantation into the rat infarcted heart.

Fabrication of Orientation-Controlled 3D Tissues Using a Layer-by-Layer Technique and 3D Printed a Thermoresponsive Gel Frame.

Herein, we report the fabrication of orientation-controlled tissues similar to heart and nerve tissues using a cell accumulation and three-dimensional (3D) printing technique. We first evaluated the 3D shaping ability of hydroxybutyl chitosan (HBC), a thermoresponsive polymer, by using a robotic dispensing 3D printer. HBC polymer could be laminated to a height of 1124 ± 14 μm.

Construction and histological analysis of a 3D human arterial wall model containing vasa vasorum using a layer-by-layer technique.

There is considerable global demand for three-dimensional (3D) functional tissues which mimic our native organs and tissues for use as in vitro drug screening systems and in regenerative medicine. In particular, there has been an increasing number of patients who suffer from arterial diseases such as arteriosclerosis. As such, in vitro 3D arterial wall models that can evaluate the effects of novel medicines and a novel artificial graft for the treatment are required.

Effect of Hydrophobic Side Chains in the Induction of Immune Responses by Nanoparticle Adjuvants Consisting of Amphiphilic Poly(γ-glutamic acid).

The new generation vaccines are safe but poorly immunogenic, and thus they require the use of adjuvants. Adjuvants that can control the balance and induction level of cellular and humoral immunities are urgently required for the treatment of and/or protection from infectious diseases and cancers. However, there are no adjuvants which can achieve these requirements.

The hydrophobic effect of nanoparticles composed of amphiphilic poly(γ-glutamic acid) on the degradability of the encapsulated proteins.

For the development of safe and effective next-generation vaccine carriers, their physicochemical properties (size, shape, surface charge, and hydrophobic/hydrophilic balance) are crucial to control their interactions (cellular uptake, intracellular degradability of the loaded antigen, and intracellular localization) with immune cells. Recently, the hydrophobicity of carriers affected the cellular uptake and immune response, which demonstrated that hydrophobicity is one of the most important factors to control the behaviors of the loaded antigens and carriers. In this study, we investigated the effect of the hydrophobicity of nanoparticles (NPs) composed of amphiphilic poly(γ-glutamic acid)-graft-phenylalanine ethyl ester (γ-PGA-Phe) with various grafting degrees of hydrophobic side chains on cellular uptake of the encapsulated antigens, their degradability, and their release behavior in the endosomal environment.

Biodegradable nanoparticles composed of enantiomeric poly(γ-glutamic acid)-graft-poly(lactide) copolymers as vaccine carriers for dominant induction of cellular immunity.

The design of particulate materials with controlled degradation at desired sites is important in applications for drug/vaccine/gene delivery systems. Amphiphilic biodegradable polymeric nanoparticles are promising vaccine delivery carriers due to their ability to stably maintain antigens, provide tailored release kinetics, effectively target, and function as adjuvants. In this study, we report that stereocomplex nanoparticles (SC NPs) composed of enantiomeric poly(γ-glutamic acid)-graft-poly(lactide) (γ-PGA-PLA) copolymers are excellent protein delivery carriers for vaccines that can deliver antigenic proteins to dendritic cells (DCs) and elicit potent immune responses.

The role of hydrophobicity in the disruption of erythrocyte membrane by nanoparticles composed of hydrophobically modified poly(γ-glutamic acid).

Polymeric nanoparticles (NPs) prepared from biocompatible polymers have been studied extensively as carriers for the targeted and controlled delivery of antigens to develop safe and effective vaccines. Especially, the endosomal escape of antigens is essential for the induction of antigen-specific potent immune responses, and the NPs which can control the endosomal escape are urgently required. It has been reported that the hydrophobicity of polymers affected the interactions between the polymer and the membranes, but there have no reports about investigating the effect of the hydrophobicity of the NPs on the membrane disruptive property.

Manipulating the antigen-specific immune response by the hydrophobicity of amphiphilic poly(γ-glutamic acid) nanoparticles.

The new generation vaccines are safe but poorly immunogenic, and thus they require the use of adjuvants. However, conventional vaccine adjuvants fail to induce potent cellular immunity, and their toxicity and side-effects hinder the clinical use. Therefore, a vaccine adjuvant which is safe and can induce an antigen-specific cellular immunity-biased immune response is urgently required.

Size effect of amphiphilic poly(γ-glutamic acid) nanoparticles on cellular uptake and maturation of dendritic cells in vivo.

We prepared size-regulated nanoparticles (NPs) composed of amphiphilic poly(γ-glutamic acid) (γ-PGA). In this study, 40, 100 and 200 nm γ-PGA-graft-l-phenylalanine ethylester (γ-PGA-Phe) NPs were employed. The size of NPs significantly influenced the uptake and activation behaviors of antigen-presenting cells (APCs).

Synergistic stimulation of antigen presenting cells via TLR by combining CpG ODN and poly(γ-glutamic acid)-based nanoparticles as vaccine adjuvants.

CpG oligodeoxynucleotide (ODN) encapsulated poly(γ-glutamic acid)-graft-l-phenylalanine ethyl ester (γ-PGA-Phe) nanoparticles (NPs) employing polycations were prepared to develop vaccine delivery and adjuvant systems. The CpG ODN was stably encapsulated into the NPs when protamine was used as the polycation. The CpG ODN-encapsulated γ-PGA-Phe NPs were taken up by macrophages and CpG ODN which was encapsulated into the NPs internalized into endo/lysosomes, where the toll-like receptor (TLR) 9, which recognizes CpG ODN, is expressed.

Uptake of biodegradable poly(γ-glutamic acid) nanoparticles and antigen presentation by dendritic cells in vivo.

Poly(γ-glutamic acid) (γ-PGA) nanoparticles (NPs) carrying antigens have been shown to induce potent antigen-specific immune responses. However, in vivo delivery of γ-PGA NPs to dendritic cells (DCs), a key regulator of immune responses, still remains unclear. In this study, γ-PGA NPs were examined for their uptake by DCs and subsequent migration from the skin to the regional lymph nodes (LNs) in mice.

Comparative activity of biodegradable nanoparticles with aluminum adjuvants: antigen uptake by dendritic cells and induction of immune response in mice.

Biodegradable poly(γ-glutamic acid) (γ-PGA) nanoparticles (NPs) are considered to be an excellent antigen carrier. Antigen-carrying γ-PGA NPs were examined for their uptake by murine dendritic cells (DCs) and subsequent induction of antigen-specific immune responses in mice and compared with aluminum (AL) adjuvants. Ovalbumin (OVA)-carrying NPs (FITC-OVA-NPs) were taken up much more efficiently by DCs than OVA alone or its AL-associated form.

Intracellular degradation and distribution of protein-encapsulated amphiphilic poly(amino acid) nanoparticles.

Physicochemical properties, such as particle size, shape, molecular weight, surface charge and composition, play a key role in the cellular uptake of polymeric nanoparticles. Antigen-encapsulated biodegradable nanoparticles have considerable potential for use in vaccine delivery systems. Although it is accepted that particle size is important for the induction of antigen-specific immune responses in vivo, little is known about how their size affects their intracellular fate.