Effect of collagen-glycosaminoglycan scaffold pore size on matrix mineralization and cellular behavior in different cell types

Significance Statement

Bone tissue engineering has emerged as one of the leading fields in tissue engineering and regenerative medicine. Its success relies on understanding the interplay between progenitor cells, regulatory signals, and the biomaterials/scaffolds used to deliver them. Subtle changes in scaffold architecture can have significant effects on cellular activity. Optimising the design of bioactive scaffolds is guided by an understanding of the behaviour and responses of cells to their surrounding environment. Pore size is an essential architectural consideration in construct development; therefore, it is crucial to identify the optimal pore size for augmented tissue formation.

Using a series of collagen-glycosaminoglycan (CG) scaffolds with a homogenous mean pore size ranging from 85 µm up to 325 µm, we identified key differences in osteoblast and mesenchymal stem cell (MSC) behaviour in response to pore size. Scaffolds with the largest pore size (325 µm) facilitated superior osteoblast attachment, migration, scaffold infiltration and matrix deposition. MSC response was similar to osteoblasts but cell motility, proliferation, and scaffold infiltration was reduced. This was associated with differences in the profile of integrin subunits (α2) and collagen receptors (CD44), indicating that osteoblasts have a stronger affinity for collagen-glycosaminoglycan scaffolds compared to MSCs.

This study, for the first time within the literature, compares two very different cell types head to head to investigate individual cell behaviour in response to a single parameter. The findings elucidate fundamental mechanisms underlying the differences between the two cell types and highlight the importance of tailoring scaffold micro-architecture and cell type for cell-specific applications.

About the author

Dr. Ciara Murphy received her PhD in area of bone tissue engineering from the Royal College of Surgeons in Ireland (RCSI) in 2010. Subsequently, she joined the Orthopaedic & Biotechnology Research (ORB) Group in the Children’s Hospital at Westmead, Sydney, Australia, where she focused her post-doctoral research on developing biologic delivery systems that utilised tissue engineering technologies, including collagen-based scaffolds, as novel therapies for bone healing.

In 2014, she was awarded the New Investigator Recognition Award (NIRA) at the International Orthopaedic Research Society (ORS) for her postdoctoral work. She returned to Ireland in 2015 joining University College Dublin (UCD) as an Assistant Professor in the School of Medicine and a Principal Investigator in the UCD Centre for Biomedical Engineering. Her research focuses on developing advanced biomaterials as innovative platforms for targeted therapeutic delivery, disease model systems and 3-D studies of cell-matrix interactions.

About the author

A/Prof Garry Duffy leads a multidisciplinary team of biomaterials, stem cell and drug delivery scientists within the Tissue Engineering Research Group (TERG), based in the Royal College of Surgeons in Ireland, with a large focus on chronic diseases. The long-term goal of his lab is to develop advanced biomaterials to facilitate targeted delivery and future clinical translation of cell based therapeutics. As well as the DRIVE project, Garry also leads the Advanced Materials for Cardiac Regeneration (AMCARE) project, an €8.6 million FP7-funded research programme with the goal of using smart biomaterials and minimally-invasive surgical devices for targeted delivery of stem cells to treat the infarcted myocardium.

About the author

A/Prof Aaron Schindeler is a Senior Research Scientist at The Children’s Hospital at Westmead and the Director of Basic Research in the Centre for Children’s Bone & Musculoskeletal Health (CCBMH). He joined the orthopaedic research department in 2003 and since then has tackled a range of research questions looking at traumatic bone injuries and genetic diseases affecting. Aaron leads a multidisciplinary team of scientists, engineers, and medical and allied health professionals. Key research areas for him include reducing the risk and impact of fracture and implant infection, cell and genetic therapies for brittle bone disease, studying the metabolic muscle weakness associated with neurofibromatosis type 1, and bone tissue engineering using novel biomaterials and 3D printing.

About the author

Prof Fergal O’Brien is a leading innovator in the development of advanced biomaterials for drug delivery and tissue repair. He is Professor of Bioengineering & Regenerative Medicine, Deputy Director for Research and heads the Tissue Engineering Research Group based in the Royal College of Surgeons in Ireland. He is also a PI and Deputy Director of the €58 million SFI-funded Advanced Materials and Bioengineering Research (AMBER) Centre. He is currently a member of the World Council of Biomechanics, Biomaterials Topic Chair for the Orthopaedic Research Society and President of the Section of Bioengineering of the Royal Academy of Medicine in Ireland.

Since his faculty appointment in 2003, he has published over 150 journal articles in leading peer-reviewed international journals and supervised 30 doctoral candidates to completion. He has a current h-index of 47. Accolades include a Fulbright Scholarship (2001), New Investigator Recognition Award by the Orthopaedic Research Society (2002), Science Foundation Ireland, President of Ireland Young Researcher Award (€1.1. million, 2004), Engineers Ireland Chartered Engineer of the Year (2005), European Research Council (ERC) Investigator Award (€2 million, 2009), Anatomical Society New Fellow of the Year (2014) and Fellowship of Engineers Ireland (2013) and the European Alliance for Medical & Biological Engineering Science (2016).

Journal Reference

J Biomed Mater Res A. 2016;104(1):291-304.

Murphy CM^1,2,3, Duffy GP^2,3,4, Schindeler A^5,6, O’brien FJ^2,3,4.

[expand title=”Show Affiliations”]

School of Medicine & Medical Science, University College Dublin, Dublin, Ireland.
Tissue Engineering Research Group, Department of Anatomy, Royal College of Surgeons in Ireland (RCSI), Dublin, Ireland.
Trinity Centre for Bioengineering, Trinity College Dublin (TCD), Dublin, Ireland.
Advanced Materials and Bioengineering Research Centre (AMBER) RCSI & TCD, Dublin, Ireland.
Orthopaedic Research & Biotechnology Unit the Children’s Hospital at Westmead.
Discipline of Paediatrics and Child Health, University of Sydney, Sydney, Australia.

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Abstract

We have previously examined osteoblast behavior on porous collagen-glycosaminoglycan (CG) scaffolds with a range of mean pore sizes demonstrating superior cell attachment and migration in scaffolds with the largest pores (325 μm). Scaffolds provide a framework for construct development; therefore, it is crucial to identify the optimal pore size for augmented tissue formation. Utilizing the same range of scaffolds (85 μm – 325 μm), this study aimed to examine the effects of mean pore size on subsequent osteoblast differentiation and matrix mineralization, and to understand the mechanism by which pore size influences behavior of different cell types. Consequently, primary mesenchymal stem cells (MSCs) were assessed and their behavior compared to osteoblasts.

Results demonstrated that scaffolds with the largest pore size (325 μm) facilitated improved osteoblast infiltration, earlier expression of mature bone markers osteopontin (OPN) and osteocalcin (OCN), and increased mineralization. MSCs responded similarly to osteoblasts whereby cell attachment and scaffold infiltration improved with increasing pore size. However, MSCs showed reduced cell motility, proliferation, and scaffold infiltration compared to osteoblasts. This was associated with differences in the profile of integrin subunits (α2) and collagen receptors (CD44), indicating that osteoblasts have a stronger affinity for collagen-glycosaminoglycan scaffolds compared to MSCs.

In summary, these results reveal how larger pores promote improved cell infiltration, essential for construct development, however the optimal scaffold pore size can be cell type specific. As such, this study highlights a necessity to tailor both scaffold micro-architecture and cell-type when designing constructs for successful bone tissue engineering applications.

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