Robert Keatch

Engineering the Bone–Ligament Interface Using Polyethylene Glycol Diacrylate Incorporated with Hydroxyapatite

Ligaments and tendons have previously been tissue engineered. However, without the bone attachment, implantation of a tissue-engineered ligament would require it to be sutured to the remnant of the injured native tissue. Due to slow repair and remodeling, this would result in a chronically weak tissue that may never return to preinjury function. In contrast, orthopaedic autograft reconstruction of the ligament often uses a bone-to-bone technique for optimal repair. Since bone-to-bone repairs heal better than other methods, implantation of an artificial ligament should also occur from bone-to-bone. The aim of this study was to investigate the use of a poly(ethylene glycol) diacrylate (PEGDA) hydrogel incorporated with hydroxyapatite (HA) and the cell-adhesion peptide RGD (Arg-Gly-Asp) as a material for creating an in vitro tissue interface to engineer intact ligaments (i.e., bone-ligament-bone). Incorporation of HA into PEG hydrogels reduced the swelling ratio but increased mechanical strength and stiffness of the hydrogels. Further, HA addition increased the capacity for cell growth and interface formation. RGD incorporation increased the swelling ratio but decreased mechanical strength and stiffness of the material. Optimum levels of cell attachment were met using a combination of both HA and RGD, but this material had no better mechanical properties than PEG alone. Although adherence of the hydrogels containing HA was achieved, failure occurs at about 4 days with 5% HA. Increasing the proportion of HA improved interface formation; however, with high levels of HA, the PEG HA composite became brittle. This data suggests that HA, by itself or with other materials, might be well suited for engineering the ligament-bone interface.

Construction of 3D biological matrices using rapid prototyping technology

Purpose – Hydrogels with low viscosities tend to be difficult to use in constructing tissue engineering (TE) scaffolds used to replace or restore damaged tissue, due to the length of time it takes for final gelation to take place resulting in the scaffolds collapsing due to their mechanical instability. However, recent advances in rapid prototyping have allowed for a new technology called bioplotting to be developed, which aims to circumvent these inherent problems. This paper aims to present details of the process.Design/methodology/approach – The paper demonstrates how by using the bioplotting technique complex 3D geometrical scaffolds with accurate feature sizes and good pore definition can be fabriated for use as biological matrices. PEG gels containing the cell‐adhesive RGD peptide sequence were patterned using this method to produce layers of directional microchannels which have a functionalised bioactive surface. Seeding these gels with C2C12 myoblasts showed that the cells responded to the topogra...

Quantitative evaluation of degenerated tendon model using combined optical coherence elastography and acoustic radiation force method

Abstract. Damage of collagen fibers in tendons is often directly related to changes in a tendon’s mechanical properties. Direct quantitative elasticity measurement of tendons will provide important information in tendon dysfunction diagnosis and treatment assessment. A feasibility study of quantifying the mechanical properties of a degenerated tendon model by a nondestructive imaging modality, which combines optical coherence elastography and acoustic radiation force (ARF) method, is presented. The degenerated tendon model was produced by the partial degradation of chicken tendons through incubation with collagenase at different concentrations and incubation times. A 30-kHz longitudinal ultrasound transducer was used to provide an ARF signal, which was detected by an ultra-high sensitive phase sensitive optical coherence tomography (PhS-OCT) system. The experimental results demonstrate that the combination of ARF method and PhS-OCT can measure the elasticity of tendon quantitatively. The corresponding changes in tendon elasticity due to the application of collagenase have been revealed by this new imaging modality. This method can potentially be used in the assessment of tissue engineering products and in the diagnosis and treatment progression of tendon diseases.

Characterisation of rapid prototyping techniques for studies in cell behaviour

Purpose – The area of microfluidic systems has greatly enhanced the in vitro field of tissue engineering. Microfluidic systems such as microchannelled assays are now widely used for mimicking in vivo cell behaviour and studies into basic biological research. In certain cases engineered tissue cell design use 3D ordered geometrical configurations in vitro (such as microchannel assays) to reproduce native in vivo functions. The most common approach for manufacturing micro‐assays is now rapid prototyping (RP) technology. The choice of assay material is dependent on the proposed cell type and ultimately the tissue application. However, many RP technologies can be unsuitable for cell growth applications because of the construction methods and materials they employ. The purpose of this paper is to describe a comparison between two different RP 3D printing methods of fabrication and investigates the merits of each technology for direct cell culture applications using micro‐assays, while also examining the dispen...

Formed 3D Bio-Scaffolds via Rapid Prototyping Technology

The construction of biomaterial scaffolds for cell seeding is now seen as the most common approach for producing artificial tissue as compared with cell self-assembly and Acellular matrix techniques. This paper describes the use of synthetic and natural polymeric material shaped into 3D biological matrices by using Rapid Prototyping (RP) technology. Recent advances in RP technology have greatly enhanced the range of biomaterials that can now be constructed into scaffolds, also allowing for maximized control of the pore size and architecture. Bioplotting is one such method which allows the dispensing of various biomaterials into a media bath which has similar rheological properties and acts as mechanical support and in most cases a cross-linking agent to produce high quality scaffolds. This method was used to construct scaffolds using agarose and gelatin with tight interconnecting pores which aim to enhance cell growth. Bioplotting was also used to pattern microchanneled layers in one direction with a PEG gel containing cell adhesive RGD peptide sequence, when seeded with C2C12 myoblasts demonstrated that cells responded to their topographical environment and aligned along the direction of the layered microchannels. This result indicates that this technique can be used to produce 3D scaffolds which aid tissue regeneration for physiologically functional tissue.

A microstructure for detecting the stress distribution in thin coatings deposited on to silicon substrates

Abstract A microscopic gauge is described which may be used for detecting and measuring the stress which is present in thin coatings which are deposited on to silicon substrates. An array of these structures may be formed on the substrate by means of microengineering fabrication processes, and this enables the uniformity of the stress to be mapped. Evaluation is made from an SEM photomicrograph.

The Development of a Combined Electrical & Mechanical Stimulation Bioreactor to Improve Tissue Engineered Muscle Function

This paper describes the development of novel bioreactor systems for use in studying the development of tissue engineered muscle constructs. The bioreactor systems combine two environmental stimuli, mechanical loading and electrical pulses which, independently, are known to promote muscle tissue growth, into a single system to investigate whether further functional improvements can be achieved. The bioreactors are reprogrammable to enable different stimulation protocols to be assessed and allow the simultaneous stimulation of up to six tissues in a single system. This allows us to improve experimental throughput and speeds up our understanding of the physiological processes involved in skeletal muscle development.

Microstructure and Cell Adhesion of Hydroxyapatite/Collagen Composites

This paper reports the development of a simple and cost effective method to fabricate hydroxyapatite/collagen nanocomposites. Hydroxyapatite is obtained by simultaneous dropping of calcium containing solution and phosphate containing solution into a reaction vessel. The microstructure of the hydroxyapatite particles exhibits a plate-like morphology under certain pH and temperature conditions. The plate-like morphology has a positive effect on the flexural strength of the composites prepared from precipitated hydroxyapatite and agarose gel. Cell adhesion tests on the precipitated hydroxyapatite and collagen indicate that the composite has excellent biocompatibility.

Micro-gripping of Small Scale Tissues

This paper describes the design and simulation of an integrated micro-electro-mechanical system (MEMS) to be used for small scale tissue manipulation. The microgrippers are to be used to test the mechanical cell adhesion properties of the gut epithelium. In the majority of sporadic colon cancers the Adenomatous Polyopsis Coli (APC) protein is mutated or missing. Mutations of APC occur extremely early in the development of cancer, before formation of polyps. Micro-grippers were designed and finite element analysis (FEA) was used to find actuation displacements, tip temperature and stresses. Monolayers of gut epithelial tissue will be grown on collagen substrates and stretched under tensile force. Ni micro-grippers will be used to grip the substrate due to its high gripping stiffness and force resolution. SU-8 micro-grippers will be used to directly grip on cell membrane to analyze cell adhesion forces with APC present or absent. The following paper shows the design of the system and FEA of micro-grippers.

The Production of High-Aspect-Ratio Microstructures (HARMS)

Abstract Current techniques used for the production of individual hohlraum components require single-point ultra-precision machining. In an effort to reduce cost and time, mass-production techniques adopted from the Microelectronics sector have been developed. These Microengineering processes will allow a variety of materials to be investigated with various geometrical features and surface topographies. Using thick photosensitive polymers, combined with electroplating processes, complex 3-D structures can be fabricated in multiple stages. In this review the production of a hohlraum end-cap used for indirect inertial confinement fusion (ICF) is presented as an example of this procedure.