Structure–property relationships between material properties and stem cell behaviour are investigated using high-throughput methods. The data identify the optimal substrates within a range of different polymeric surfaces to support the growth and self-renewal of human embryonic stem cells from fully dissociated single cells.

Combinatorial Development of Biomaterials for Clonal Growth of Human Pluripotent Stem Cells

The ultimate goal of most biomedical research is to gain greater insight into mechanisms of human disease or to develop new and improved therapies or diagnostics. Although great advances have been made in terms of developing disease models in animals, such as transgenic mice, many of these models fail to faithfully recapitulate the human condition. In addition, it is difficult to identify critical cellular and molecular contributors to disease or to vary them independently in whole-animal models. This challenge has attracted the interest of engineers, who have begun to collaborate with biologists to leverage recent advances in tissue engineering and microfabrication to develop novel in vitro models of disease. As these models are synthetic systems, specific molecular factors and individual cell types, including parenchymal cells, vascular cells, and immune cells, can be varied independently while simultaneously measuring system-level responses in real time. In this article, we provide some examples of these efforts, including engineered models of diseases of the heart, lung, intestine, liver, kidney, cartilage, skin and vascular, endocrine, musculoskeletal, and nervous systems, as well as models of infectious diseases and cancer. We also describe how engineered in vitro models can be combined with human inducible pluripotent stem cells to enable new insights into a broad variety of disease mechanisms, as well as provide a test bed for screening new therapies.

Engineered in vitro disease models.

We report the first investigation into the bioprinting of human induced pluripotent stem cells (hiPSCs), their response to a valve-based printing process as well as their post-printing differentiation into hepatocyte-like cells (HLCs). HLCs differentiated from both hiPSCs and human embryonic stem cells (hESCs) sources were bioprinted and examined for the presence of hepatic markers to further validate the compatibility of the valve-based bioprinting process with fragile cell transfer. Examined cells were positive for nuclear factor 4 alpha and were demonstrated to secrete albumin and have morphology that was also found to be similar to that of hepatocytes. Both hESC and hiPSC lines were tested for post-printing viability and pluripotency and were found to have negligible difference in terms of viability and pluripotency between the printed and non-printed cells. hESC-derived HLCs were 3D printed using alginate hydrogel matrix and tested for viability and albumin secretion during the remaining differentiation and were found to be hepatic in nature. 3D printed with 40-layer of HLC-containing alginate structures reached peak albumin secretion at day 21 of the differentiation protocol. This work demonstrates that the valve-based printing process is gentle enough to print human pluripotent stem cells (hPSCs) (both hESCs and hiPSCs) while either maintaining their pluripotency or directing their differentiation into specific lineages. The ability to bioprint hPSCs will pave the way for producing organs or tissues on demand from patient specific cells which could be used for animal-free drug development and personalized medicine.

https://iopscience.iop.org/article/10.1088/1758-5090/7/4/044102/pdf

Bioprinting of human pluripotent stem cells and their directed differentiation into hepatocyte-like cells for the generation of mini-livers in 3D

Ultrafast laser nanostructuring of photopolymers: a decade of advances

Stereolithography is a solid freeform technique (SFF) that was introduced in the late 1980s Although many other techniques have been developed since then, stereolithography remains one of the most powerful and versatile of all SFF techniques It has the highest fabrication accuracy and an increasing number of materials that can be processed is becoming available In this paper we discuss the characteristic features of the stereolithography technique and compare it to other SFF techniques The biomedical applications of stereolithography are reviewed, as well as the biodegradable resin materials that have been developed for use with stereolithography Finally, an overview of the application of stereolithography in preparing porous structures for tissue engineering is given

A review on stereolithography and its applications in biomedical engineering

This study reports on the production of high-resolution 3D structures of polylactide-based materials via multi-photon polymerization and explores their use as neural tissue engineering scaffolds. To achieve this, a liquid polylactide resin was synthesized in house and rendered photocurable via attaching methacrylate groups to the hydroxyl end groups of the small molecular weight prepolymer. This resin cures easily under UV irradiation, using a mercury lamp, and under femtosecond IR irradiation. The results showed that the photocurable polylactide (PLA) resin can be readily structured via direct laser write (DLW) with a femtosecond Ti:sapphire laser and submicrometer structures can be produced. The maximum resolution achieved is 800 nm. Neuroblastoma cells were grown on thin films of the cured PLA material, and cell viability and proliferation assays revealed good biocompatibility of the material. Additionally, PC12 and NG108-15 neuroblastoma growth on bespoke scaffolds was studied in more detail to assess potential applications for neuronal implants of this material.

https://iopscience.iop.org/article/10.1088/1758-5082/3/4/045005/pdf

Direct laser writing of 3D scaffolds for neural tissue engineering applications.

We report on the fabrication of high resolution 3D scaffolds of polylactide-based materials using direct laser writing and we explore their use as neural tissue engineering scaffolds.

Direct Laser Writing of 3D scaffolds for neural tissue engineering applications

In this study, we explore the production of well-defined macroscopic scaffolds with two-photon polymerization (2PP) and their use as neural tissue engineering scaffolds. We also demonstrate that these 3D scaffolds can be replicated via soft lithography, which increases production efficiency. Photopolymerizable polylactic acid (PLA) was used to produce scaffolds by 2PP and soft lithography. We assessed the biocompatibility of these scaffolds using an SH-SY5Y human neuronal cell line and primary cultured rat Schwann cells (of direct relevance to the repair of nerve injuries). A Comet assay with SH-SY5Y human neuronal cells revealed minimal DNA damage after washing the photocured material for 7 days in ethanol. Additionally, thin films and 3D scaffolds of the photocured PLA sustained a high degree of Schwann cell purity (99%), enabled proliferation over 7 days and provided a suitable substrate for supporting Schwann cell adhesion such that bi-polar and tri-polar morphologies were observed. Evidence of orthogonally aligned and organized actin thin filaments and the formation of focal contacts were observed for the majority of Schwann cells. In summary, this work supports the use of PLA as a suitable material for supporting Schwann cell growth and in turn use of 3D soft lithography for the synthesis of neural scaffolds in nerve repair.

https://iopscience.iop.org/article/10.1088/1758-5082/4/2/025005/pdf

Two-photon polymerization-generated and micromolding-replicated 3D scaffolds for peripheral neural tissue engineering applications

In this study, we report the production of amine functionalized nanodiamond. The amine functionalized nanodiamond forms a conformal monolayer on a negatively charged surface produced via plasma polymerization of acrylic acid. Nanodiamond terminated surfaces were studied as substrates for neuronal cell culture. NG108-15 neuroblastoma-glyoma hybrid cells were successfully cultured upon amine functionalized nanodiamond coated surfaces for between 1 and 7 d. Additionally, primary dorsal root ganglion (DRG) neurons and Schwann cells isolated from Wistar rats were also successfully cultured over a period of 21 d illustrating the potential of the coating for applications in the treatment of peripheral nerve injury.

/pdf/amine-functionalized-nanodiamond-promotes-cellular-adhesion-qstagr9w0b.pdf

Amine functionalized nanodiamond promotes cellular adhesion, proliferation and neurite outgrowth.

The aim of this study was to develop a synthetic alternative to the human corneal limbus for use initially as an ex vivo model in which to study corneal stem cell function within a niche environment and ultimately to develop an implantable limbus for future clinical use. Microstereolithography was used for the fabrication of polyethylene glycol diacrylate (PEGDA) based rings on a macroscopic (1.2 cm) scale containing unique microfeatures (pockets) which were then modified with fibronectin to promote cell adhesion. These rings were designed to mimic the limbal area of the eye containing structures of the approximate size and shape of the stem cell microenvironments found in the palisades of Vogt. The attachment of rabbit limbal fibroblasts and rabbit limbal epithelial cells to the PEGDA rings was increased by pretreating the microfabricated structures with biotinylated fibronectin. Cell outgrowth from fibronectin coated microfabricated structures was 50% greater than from rings without structures or fibronectin coating. The cell loaded rings were then placed on an ex vivo wounded cornea model and the outgrowth of cells to form a multilayered epithelium was observed. We suggest this is a new approach to investigating limbal stem cells niches and the first steps towards a new approach for corneal regeneration.

http://iopscience.iop.org/article/10.1088/1758-5082/5/2/025008/pdf

A.A. Gill

Papers

Direct laser writing of 3D scaffolds for neural tissue engineering applications.

Direct Laser Writing of 3D scaffolds for neural tissue engineering applications

Two-photon polymerization-generated and micromolding-replicated 3D scaffolds for peripheral neural tissue engineering applications

Amine functionalized nanodiamond promotes cellular adhesion, proliferation and neurite outgrowth.

Development of a microfabricated artificial limbus with micropockets for cell delivery to the cornea.