Cell Modification in 3D: a new Paradigm in the Creation of Living Cell-Biomaterial Composites

Lead Research Organisation: University of Southampton
Department Name: Development Origin of Health and Disease


Our aims are to develop new, improved materials and methods which will allow stem cells (mesenchymal; MSCs) to be manipulated to form bone tissue. We will do this using tiny (<100 nm; 1/1000 diameter human hair) calcium phosphate (hydroxyapatite; HAP) particles as vectors to carry specific biological molecules to the cells. To maximize the delivery of these chemical and genetic signals, the whole cell surface of each individual cell will be covered with the vectors (3D coating). Specifically, this will allow us to produce and grow self supporting, living bone tissue, either inside the body at the site of damage, or outside in culture dishes ready for implant. More generally, the improved efficiency and cost effectiveness of this approach will also enhance studies in the generation of other tissue types from MSCs (e.g. nerve and muscle) and in modifying other types of stem cell.Why mesenchymal stem cells?Stem cells have huge potential as therapeutic agents. Embryonic stem cells (ESC) have the potential to form all the major types of cell in the body, and are relatively easy to grow in culture. However, there are ethical and compatibility concerns with there use. Adult stem cells (ASC) can be harvested from specific tissue types (blood, nerves, skin), but the populations need to be expanded to get sufficient material for therapeutic use. In this regard, bone marrow mesenchymal stem cells (MSC) offer great hope for tissue engineering as methods for isolation and rapid cultivation are well established. They are natural precursors to bone, cartilage, fat and fibrous connective tissue formation. Thus they are already intensively studied as components of systems for replacing damaged bone tissues (e.g. restorative surgery). In addition the same person can be donor and recipient, thus alleviating the problems associated with ESCs.Why small hydroxyapatite particles?Hydroxyapatite is the chemical form of calcium phosphate found in bone, so it is compatible with the cells. The crystals of HAP in bone are also of a similar size (<100 nm). In addition, because the crystals are so small as well as coating the cell, some will be transported inside the cell. Thus the particles can be used to deliver information to the cell surface and interior.Why chemical and genetic signals?The key to using stem cells to regenerate tissue is the ability to persuade them to form the required type. There are two ways to manipulate these cells towards bone formation / direct genetic modification of the internal cell nucleus, or the use of indirect external stimuli such as chemicals secreted by other cells or present in the local cell environment, and physical contact with other cells.Why 3D coating?Bone-like cell behaviour is induced indirectly by adding expensive chemicals to the culture medium or adsorbing them onto the substrates. Direct genetic modification requires DNA to be delivered to the cell nucleus, typically by attachment to small carrier particles. For example, standard methods for gene delivery using HAP involve mixing the precursor chemicals together, then allowing the crystals that form to randomly settle on the cells like a snowstorm. There are two main factors which contribute to the inefficiency of both these current approaches; (a) the cells are adhered to a substrate, so not all the cell surface is available, and (b) the cells themselves are not targeted, so the additives are used at higher concentration than necessary. Both these problems should be alleviated by our proposed method.
Description This work set out to examine the generation and development of bone void fillers containing added biological function to aid skeletal tissue formation. This would have implications for improved efficacy in bone replacement surgery. We demonstrate the potential for hydroxyapatite nanoparticles with motifs (small amino acid sequences) that allow cell binding can aid bone matrix formation both in the laboratory and in small animals. Hydroxyapatite nanoparticles were combined with human bone marrow stromal for up to 21 days, either as a monolayer or as a 3D pellet culture system, resulting in significantly increased levels of markers of bone differentiation in comparison with uncoated cells.
Critically, in 3D pellet culture conditions all hydroxyapatite nanoparticle cell complexes promoted bone cell differentiation and animal studies in mice showed new osteoid formation after 21 days, comparable to the extensive areas of mineralized extracellular matrix seen in the laboratory. Thus we have shown that pre-coating of bone marrow stromal cells enables the formation of viable hybrid multicellular 3D constructs with demonstrable activity both in the laboratory and in animals.
Similarly our work using calcium carbonate microspheres (vaterite microspheres) led to bone cell activation and promotion of 3D skeletal tissue formation in the laboratory and in animals using a preclinical model where bone stock is impacted into a construct. These studies demonstrate that bio-inspired calcium carbonate microspheres aid in vivo bone formation in impaction bone grafting. We were able to add functional groups to the microspheres and thus our microspheres with added biological functionality offer innovative therapeutic approaches to activate skeletal populations and enhance bone formation. These studies offer real development opportunities in bone repair
Exploitation Route Our development of innovative hydroxyapatite nanoparticles and calcium carbonate microspheres to aid bone cell differentiation and bone formation in animals offers new approaches to hard tissue repair.
Sectors Healthcare

Description Development of teaching tools and ideas for regenerative medicine demonstration to students and adult populations
First Year Of Impact 2011
Sector Healthcare
Description Stem Cell Mountain at Bestival Music Festival Science Tent 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Stem Cell Mountain at the Bestival Music Festival Science Tent (4 days) - exhibit in shared Science Tent, mixed Festival audience (mainly general public and especially young people 17-25) Measure 1: visitors passing stand; 1400 per day = 5600 Measure 2: visitors interacting with researchers on stand; 500 per day = 2000 Particularly notable was the impact of this activity on the researchers themselves and how they viewed the importance of their own research. For example one researcher reported: "This festival was great to see how amazed people are about the potential of stem cells"
Year(s) Of Engagement Activity 2015