Faculty of Science and Engineering, The University of Manchester

We are pleased to introduce the collection Frontiers in Medical Technology – Highlights in Regenerative Technologies 2021/22. This collection will welcome and showcase a selection of high impact articles authored by leaders in the field. The work presented here highlights the broad diversity of research performed across the Regenerative Technologies section, and aims to put a spotlight on the main areas of interest. This collection aims to further support Frontiers’ strong community by shining a spotlight on our authors' highly impactful research.

The University of Sheffield

Lancaster University

Spanish National Research Council (CSIC)

Frontiers in Medical Technology

Regenerative Technologies

Highlights in Regenerative Technologies 2021/22

Allogeneic T cells are key immune therapeutic cells to fight cancer and other clinical indications. High T cell dose per patient and increasing patient numbers result in clinical demand for a large number of allogeneic T cells. This necessitates a manufacturing platform that can be scaled up while retaining cell quality. Here we present a closed and scalable platform for T cell manufacturing to meet clinical demand. Upstream manufacturing steps of T cell activation and expansion are done in-vessel, in a stirred-tank bioreactor. T cell selection, which is necessary for CAR-T-based therapy, is done in the bioreactor itself, thus maintaining optimal culture conditions through the selection step. Platform's attributes of automation and performing the steps of T cell activation, expansion, and selection in-vessel, greatly contribute to enhancing process control, cell quality, and to the reduction of manual labor and contamination risk. In addition, the viability of integrating a closed, automated, downstream process of cell concentration, is demonstrated. The presented T cell manufacturing platform has scale-up capabilities while preserving key factors of cell quality and process control.

Allogeneic T cells are key immune therapeutic cells to fight cancer and other clinical indications. High T cell dose per patient and increasing patient numbers result in clinical demand for a large number of allogeneic T cells. This necessitates a manufacturing platform that can be scaled up while retaining cell quality. Here we present a closed and scalable platform for T cell manufacturing to meet clinical demand. Upstream manufacturing steps of T cell activation and expansion are done in-vessel, in a stirred-tank bioreactor. T cell selection, which is necessary for CAR-T-based therapy, is done in the bioreactor itself, thus maintaining optimal culture conditions through the selection step. Platform&apos;s attributes of automation and performing the steps of T cell activation, expansion, and selection in-vessel, greatly contribute to enhancing process control, cell quality, and to the reduction of manual labor and contamination risk. In addition, the viability of integrating a closed, automated, downstream process of cell concentration, is demonstrated. The presented T cell manufacturing platform has scale-up capabilities while preserving key factors of cell quality and process control.

Enabling Allogeneic T Cell-Based Therapies: Scalable Stirred-Tank Bioreactor Mediated Manufacturing

Decellularized extracellular matrix (dECM) deposited by mesenchymal stromal cells (MSCs) has emerged as a promising substrate for improved expansion of MSCs. To date, essentially all studies that have produced dECM for MSC expansion have done so on tissue culture plastic or glass. However, substrate surface chemistry has a profound impact on the adsorption of proteins that mediate cell-material interactions, and different surface chemistries can cause changes in cell behavior, ECM deposition, and the in vivo response to a material. This study tested the hypothesis that substrate surface chemistry impacts the deposition of ECM and its subsequent bioactivity. This hypothesis was tested by producing glass surfaces with various surface chemistries (amine, carboxylic acid, propyl, and octyl groups) using silane chemistry. ECM was deposited by an immortalized MSC line, decellularized, and characterized through SDS-PAGE and immunofluorescence microscopy. No significant difference was observed in dECM composition or microarchitecture on the different surfaces. The decellularized surfaces were seeded with primary MSCs and their proliferation and differentiation were assessed. The presence of dECM improved the proliferation of primary MSCs by ~100% in comparison to surface chemistry controls. Additionally, the adipogenesis increased by 50–90% on all dECM surfaces in comparison to surface chemistry controls, and the osteogenesis increased by ~50% on the octyl-modified surfaces when dECM was present. However, no statistically significant differences were observed within the set of dECM surfaces or control surfaces. These results support the null hypothesis, meaning surface chemistry (over the range tested in this work) is not a key regulator of the composition or bioactivity of MSC-derived dECM. These results are significant because they provide an important insight into regenerative engineering technologies. Specifically, the utilization of dECM in stem cell manufacturing and tissue engineering applications would require the dECM to be produced on a wide variety of substrates. This work indicates that it can be produced on materials with a range of surface chemistries without undesired changes in the bioactivity of the dECM.

Decellularized extracellular matrix (dECM) deposited by mesenchymal stromal cells (MSCs) has emerged as a promising substrate for improved expansion of MSCs. To date, essentially all studies that have produced dECM for MSC expansion have done so on tissue culture plastic or glass. However, substrate surface chemistry has a profound impact on the adsorption of proteins that mediate cell-material interactions, and different surface chemistries can cause changes in cell behavior, ECM deposition, and the in vivo response to a material. This study tested the hypothesis that substrate surface chemistry impacts the deposition of ECM and its subsequent bioactivity. This hypothesis was tested by producing glass surfaces with various surface chemistries (amine, carboxylic acid, propyl, and octyl groups) using silane chemistry. ECM was deposited by an immortalized MSC line, decellularized, and characterized through SDS-PAGE and immunofluorescence microscopy. No significant difference was observed in dECM composition or microarchitecture on the different surfaces. The decellularized surfaces were seeded with primary MSCs and their proliferation and differentiation were assessed. The presence of dECM improved the proliferation of primary MSCs by ~100% in comparison to surface chemistry controls. Additionally, the adipogenesis increased by 50–90% on all dECM surfaces in comparison to surface chemistry controls, and the osteogenesis increased by ~50% on the octyl-modified surfaces when dECM was present. However, no statistically significant differences were observed within the set of dECM surfaces or control surfaces. These results support the null hypothesis, meaning surface chemistry (over the range tested in this work) is not a key regulator of the composition or bioactivity of MSC-derived dECM. These results are significant because they provide an important insight into regenerative engineering technologies. Specifically, the utilization of dECM in stem cell manufacturing and tissue engineering applications would require the dECM to be produced on a wide variety of substrates. This work indicates that it can be produced on materials with a range of surface chemistries without undesired changes in the bioactivity of the dECM.

Improvement of Mesenchymal Stromal Cell Proliferation and Differentiation via Decellularized Extracellular Matrix on Substrates With a Range of Surface Chemistries

Electrical stimulation (ES) is a well-known method for guiding the behaviour of nerve cells in in vitro systems based on the response of these cells to an electric field. From this perspective, understanding how the electrochemical stimulus can be tuned for the design of a desired cell response is of great importance. Most biomedical studies propose the application of an electrical potential to cell culture arrays while examining the cell response regarding viability, morphology, and gene expression. Conversely, various studies failed to evaluate how the fine physicochemical properties of the materials used for cell culture influence the observed behaviours. Among the various materials used for culturing cells under ES, conductive polymers (CPs) are widely used either in pristine form or in addition to other polymers. CPs themselves do not possess the optimal surface for cell compatibility because of their hydrophobic nature, which leads to poor protein adhesion and, hence, poor bioactivity. Therefore, understanding how to tailor the chemical properties on the material surface will determine the obtention of improved ES platforms. Moreover, the structure of the material, either in a thin film or in porous electrospun scaffolds, also affects the biochemical response and needs to be considered. In this review, we examine how materials based on CPs influence cell behaviour under ES, and we compile the various ES setups and physicochemical properties that affect cell behaviour. This review concerns the culture of various cell types, such as neurons, fibroblasts, osteoblasts, and Schwann cells, and it also covers studies on stem cells prone to ES. To understand the mechanistic behaviour of these devices, we also examine studies presenting a more detailed biomolecular level of interaction. This review aims to guide the design of future ES setups regarding the influence of material properties and electrochemical conditions on the behaviour of in vitro cell studies.

Electrical stimulation (ES) is a well-known method for guiding the behaviour of nerve cells in in vitro systems based on the response of these cells to an electric field. From this perspective, understanding how the electrochemical stimulus can be tuned for the design of a desired cell response is of great importance. Most biomedical studies propose the application of an electrical potential to cell culture arrays while examining the cell response regarding viability, morphology, and gene expression. Conversely, various studies failed to evaluate how the fine physicochemical properties of the materials used for cell culture influence the observed behaviours. Among the various materials used for culturing cells under ES, conductive polymers (CPs) are widely used either in pristine form or in addition to other polymers. CPs themselves do not possess the optimal surface for cell compatibility because of their hydrophobic nature, which leads to poor protein adhesion and, hence, poor bioactivity. Therefore, understanding how to tailor the chemical properties on the material surface will determine the obtention of improved ES platforms. Moreover, the structure of the material, either in a thin film or in porous electrospun scaffolds, also affects the biochemical response and needs to be considered. In this review, we examine how materials based on CPs influence cell behaviour under ES, and we compile the various ES setups and physicochemical properties that affect cell behaviour. This review concerns the culture of various cell types, such as neurons, fibroblasts, osteoblasts, and Schwann cells, and it also covers studies on stem cells prone to ES. To understand the mechanistic behaviour of these devices, we also examine studies presenting a more detailed biomolecular level of interaction. This review aims to guide the design of future ES setups regarding the influence of material properties and electrochemical conditions on the behaviour of in vitro cell studies.

Electrical Stimulation and Conductive Polymers as a Powerful Toolbox for Tailoring Cell Behaviour in vitro

This review will focus on the targeted design, synthesis and application of redox polymers for use in regenerative medicine and tissue engineering. We define redox polymers to encompass a variety of polymeric materials, from the multifunctional conjugated conducting polymers to graphene and its derivatives, and have been adopted for use in the engineering of several types of stimulus responsive tissues. We will review the fundamental properties of organic conducting polymers (OCPs) and graphene, and how their properties are being tailored to enhance material - biological interfacing. We will highlight the recent development of high-resolution 3D fabrication processes suitable for biomaterials, and how the fabrication of intricate scaffolds at biologically relevant scales is providing exciting opportunities for the application of redox polymers for both in-vitro and in-vivo tissue engineering. We will discuss the application of OCPs in the controlled delivery of bioactive compounds, and the electrical and mechanical stimulation of cells to drive behaviour and processes towards the generation of specific functional tissue. We will highlight the relatively recent advances in the use of graphene and the exploitation of its physicochemical and electrical properties in tissue engineering. Finally, we will look forward at the future of organic conductors in tissue engineering applications, and where the combination of materials development and fabrication processes will next unite to provide future breakthroughs.

This review will focus on the targeted design, synthesis and application of redox polymers for use in regenerative medicine and tissue engineering. We define redox polymers to encompass a variety of polymeric materials, from the multifunctional conjugated conducting polymers to graphene and its derivatives, and have been adopted for use in the engineering of several types of stimulus responsive tissues. We will review the fundamental properties of organic conducting polymers (OCPs) and graphene, and how their properties are being tailored to enhance material - biological interfacing. We will highlight the recent development of high-resolution 3D fabrication processes suitable for biomaterials, and how the fabrication of intricate scaffolds at biologically relevant scales is providing exciting opportunities for the application of redox polymers for both in-vitro and in-vivo tissue engineering. We will discuss the application of OCPs in the controlled delivery of bioactive compounds, and the electrical and mechanical stimulation of cells to drive behaviour and processes towards the generation of specific functional tissue. We will highlight the relatively recent advances in the use of graphene and the exploitation of its physicochemical and electrical properties in tissue engineering. Finally, we will look forward at the future of organic conductors in tissue engineering applications, and where the combination of materials development and fabrication processes will next unite to provide future breakthroughs.