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3D Hydrogel Microwell Arrays to Control Cardiac Differentiation of Human Pluripotent Stem Cells in a High Throughput Manner

Final Report Summary - STEM CELL HYDROGELS (3D Hydrogel Microwell Arrays to Control Cardiac Differentiation of Human Pluripotent Stem Cells in a High Throughput Manner)

Synthetic dental biomaterials for repair and regeneration
Dental disease is one of the leading causes of pain and infection worldwide. The global economic burden of dental disease was estimated to be $442 billion in 2010. If untreated, dental decay can progress further into the tooth and infect the vital pulp tissue, which is treated by surgically removing the pulp tissue (root canal therapy). Current dental restorative materials are toxic to cells and cannot be used in direct contact with pulp tissue. The pulp tissue contains various cell types including a stem cell population called dental pulp stem cells (DPSCs). DPSCs are mulitpotent and can differentiate to form odontoblasts, which in turn produce dentin. The objective of this project is to develop a synthetic biomaterial that can support DPSC attachment, proliferation and differentiation to repair and regenerate damaged pulp tissue and restore dentin.

High throughput screening (HTS) using polymer microarrays was used to discover new materials for pulp tissue regeneration. DPSCs were cultured on chemically-diverse polymer microarrays to find materials that support DPSC attachment (Figure 1).

[Figure 1. (a) Polymer microarray and (b) a single biomaterial from the microarray with adhered DPSCs. scale bar: 100 μm.]

The best performing polymers were scaled up for in vitro studies and were able to support attachment, proliferation and differentiation of DPSCs in cultureware. Furthermore, these materials outperformed the commercial dental restorative material (BisGMA) which is toxic to cells. After 21 days culture, genes associated with odontoblastic differentiation were found to be upregulated versus TCP control indicating that these materials could support odontoblastic differentiation and dentin production in vivo.

Physicochemical data was acquired and combined with biological data to determine mechanisms of DPSC attachment, proliferation and differentiation in response to the synthetic dental biomaterials. Specifically, quantitative polymerase chain reaction (qPCR) was used to determine genetic changes in DPSCs that had been exposed to different synthetic materials (Figure 2). This revealed that the regenerative polymers provide a supportive niche for DPSC attachment, proliferation and differentiation by maintaining Collagen I expression (Figure 2a). Conversely, commercial dental materials such methacrylates upregulate non-natural collagen genes (Figure 2b and c) and genes associated with cell stress (Figure 2d-f). Furthermore, DPSCs require integrin-ß1 to attach to the triacrylates dental biomaterials (data not shown).

This regenerative approach could significantly impact the practice of dentistry and, thus, could establish a new paradigm for improving dental treatments. Furthermore, these materials could be employed as a preventative care measure to reduce overall healthcare costs globally.

Hydrogel-based tissue adhesives
Adhesives that bond to wet and dynamic surfaces such as biological tissues would have broad applications in various fields ranging from drug delivery to biomedical devices. However, existing bioadhesives do not perform well in these conditions and this remains a significant challenge. The objective of this project was to design a material that could bond strongly to wet and dynamic surfaces while being biocompatible for use as an adhesive for tissue repair and closure.

Hydrogels have been widely used in fields such as tissue engineering and regenerative medicine as they mimic some properties of the extracellular matrix (ECM) such as elastic modulus and high water content. However, strong adhesion of hydrogels to biological tissues has not been achieved and this was used as biocompatible substrate for adhesion. The hydrogels used in this project were designed to consist of a dissipative bulk matrix which can undergo large deformation, up to 20 times the initial length, without fracture and an adhesive surface that can form bonds with the dissipative bulk matrix and the target surface (tissue). The former adheres to the substrate by electrostatic interactions, covalent bonds, and physical interpenetration. The latter amplifies the dissipation of energy through hysteresis. The two layers synergistically lead to high adhesion energy on wet surfaces, ranging from tissues such as heart and skin, to hydrogels like polyacrylamide. Adhesion is fast, independent of blood exposure, and compatible with in vivo dynamic movements (e.g. beating heart) (Figure 3).

[Figure 3. Strong adhesion on wet biological tissues. (a) ex vivo porcine skin and (b) in vivo beating heart.]

This design approach for tough adhesives can be extended to a variety of other material systems, and will open opportunities for innovation in applications such as drug delivery, tissue repair, and placement of biomedical devices in the body. It could also have a wider societal impact by reducing complications associated with suturing during surgery and reduce overall healthcare costs.
This technology has been filed in a US patent application and was recently published in the journal Science.