Real-time-monitoring enabled manufacturing of a biophysiochemical 3D in vitro tissue model of the periodontium

Secondary Supervisor(s): Dr Gowsihan Poologasundarampillai

University of Registration: University of Birmingham

BBSRC Research Themes: Integrated Understanding of Health (Regenerative Biology)

Project Outline

The periodontium (tooth-supporting tissues) is a complex organ uniquely exposed to microbial challenge. Because this multi-tissue (epithelium, connective tissue, bone and periodontal ligament) environment is prone to infection and inflammation, it is particularly important to understand health and homeostasis of the periodontium. Homeostasis is maintained through the intricate interplay of different immune (e.g., neutrophils, macrophages) and structural cells (e.g., fibroblasts, epithelial cells) with each other as well as with commensal bacteria which inhabit the oral cavity. The direct demonstration and characterisation of dynamic tissue turnover and cell-to-cell communication has not been possible due to a lack of in-vitro and ex-vivo 3D organotypic models of this multicellular tissue environment. Their availability would enable a deeper understanding of how a healthy state is maintained despite microbial challenge. In the longer term, the development of novel targeted interventions to support the healthy state and to prevent inflammation may thus be possible. Furthermore, we envisage the creation of an in vitro testing platform for pharmaceutical applications, which could reduce the need for animal models in periodontal research.

Aims and Objectives

Our goal is to create a 3D printable organotypic model of the periodontium, by assembling complex vascularised cultures in 3D hydrogels mimicking hard and soft tissues, to which various microbiological, physiochemical and inflammatory challenges can be applied. Real-time behaviour of immune cells will be reproduced in the model. Specifically, the proposed work has the following aims:

1. To develop an additive manufacturing method to create cell bearing tissue-like structures, enabling monitoring of matrix parameters such as pH, high throughput assaying of protein synthesis, and inflammatory mediator release.

2. To build complex 3D in vitro models using this process to recreate the periodontal tissue environment including structural cells, extracellular tissue matrix and blood vessels, vascularised bone and microbial biofilms.

3. To use these models to investigate migration of immune cells and their activation under in real-time.

To achieve this, we plan to combine bioprinting, organ-on-a-chip technologies and light sheet-based real-time imaging. These will take place in phases, starting with the formulation of complex soft-solid hydrogels (work package [WP] 1) to imitate extracellular matrix of periodontal tissues. This will be followed by incorporation of structural and immune cells, and the assessment of cell-hydrogel interactions (WP2). Existing protocols will be refined to manufacture blood vessels (WP3), through which immune cells can be delivered into the 3D matrix. Subsequently, bone organoids will be developed and incorporated. These will contain active bone remodelling units, which mimic the alveolar bone needed to retain teeth (WP4). Finally, bacteria will be introduced into the model. This will allow for mechanistic studies of cell behaviour for the first time, creating fundamental knowledge of this living system in health (WP5).

References

1. Hirschfeld, J. & Chapple, I.L. (editors) 2021, Periodontitis and Systemic Diseases: Clinical Evidence and Biological Plausibility. Quintessence Publishing (Berlin, Tokyo, Chicago). ISBN: 978-1-78.

2. Poologasundarampillai, G., et al. (2021). Real-time imaging and analysis of cell hydrogel interplay within an extrusion-bioprinting capillary. Bioprinting., 23, e00144.